Toxins doi: 10.3390/toxins18110484

Authors: Shauna Farr-Jones James D. Marks Milan T. Tomic

We have developed highly potent botulinum neurotoxin (BoNT) antitoxins consisting of three mAbs that bind non-overlapping epitopes and act synergistically to neutralize and rapidly clear circulating toxin. The resulting potency permits a low total antibody dose for treatment (34 mg for a seven-serotype product under the assumptions used here). Based on this high potency, the cost of producing the drug substance per dose is relatively low relative to chronic, high-dose antibody therapeutics. Recombinant human IgG also provides a longer serum half-life than equine F(ab’)2 antitoxin and therefore has potential for both treatment and prevention of botulism lasting for months. Here, we analyze the manufacturing costs of the recombinant antitoxin and the development costs of multivalent products and compare them to those of the currently approved equine-derived antitoxin. Approaches to reducing manufacturing complexity, including co-expression and co-purification, formulation, stockpiling, clinical use, analytical control, and multivalent antibody formats, are discussed. Our analysis indicates that recombinant oligoclonal BoNT antitoxins are not intrinsically cost-prohibitive and that development and overall manufacturing costs of Drug Product do not increase linearly with the number of serotypes covered.

Toxins doi: 10.3390/toxins18110483

Authors: Subsai Kongsaengdao Kanoksri Samintharapanya

Comparative evidence across repeated botulinum toxin treatment cycles for hemifacial spasm (HFS) remains limited. This 50-week double-blind randomized crossover trial compared Abobotulinumtoxin A 100 U (Abo-BoNT-A) with Neubotulinumtoxin A 33.33 U (Neu-BoNT-A) across multiple dimensions, including improvement frequency, magnitude, time course, patient impact, harms, and cost in 92 participants. Each formulation was scheduled for two consecutive 12-week periods, with a 2-week washout before crossover. Although Total Intensity Score did not demonstrate Neu-BoNT-A noninferiority, exploratory diary-defined HFS-free improvement during Days 2–14 occurred in 48.0% versus 34.1% of Abo-BoNT-A and Neu-BoNT-A treatment records (adjusted odds ratio, OR 1.89; 95% CI, 1.26–2.84; nominal p = 0.0021). Among responders, time to peak improvement and post-peak duration did not differ significantly. Selected exploratory symptom trajectories differed, whereas cycle-average burden and patient-reported outcomes showed no consistent differences. Eyelid ptosis occurred in 4.3% versus 0.5% of reported exposures (unadjusted paired p = 0.039). Under vial sharing, acquisition costs were USD 81.52 versus USD 30.78 per treatment. These multidimensional findings suggest an exploratory response–safety–cost trade-off beyond average symptom scores. Trial registration: NCT04589364.

Toxins doi: 10.3390/toxins18110482

Authors: Michael Uwe Martin Jürgen Frevert Niamh Corduff Je-Young Park Leihong Xiang Clifton Ming Tay Wei Qi Loh

Botulinum neurotoxin type A (BoNT/A) has become one of the most widely utilized biologics today in both therapeutic and aesthetic applications. Alongside innovative explorations in BoNT/A use, advancements have also been made in manufacturing and formulation practices. The earliest concept of a “pure” BoNT/A referred to the successful isolation of the native neurotoxin complex from Clostridium botulinum, which enabled its clinical development. Subsequently, various definitions of the term “purity” have been applied to BoNT/A, encompassing distinct scientific, regulatory, clinical, and commercial interpretations. Formulation advances first emphasized the removal of non-toxic bacterial components in efforts to reduce protein load and immunogenicity. Increasingly, analytical methods such as SDS-PAGE and ELISA are also commonly cited to support claims for BoNT/A identity and quantity and define purity; however, these provide limited information regarding biological activity. Given that BoNT/A is not a single uniform molecular entity, differences in bacterial strains, manufacturing processes, and the presence or absence of complexing proteins have resulted in distinct product formulations that further complicate direct comparisons relating to purity. This review examines various working definitions of BoNT/A purity, discusses implications for clinicians and patients, and highlights emerging scientific and regulatory efforts to improve product comparability through standardized and biologically relevant measures of potency.

Toxins doi: 10.3390/toxins18110481

Authors: Seyda Senturk Hakan Karaca

Aflatoxins are toxic secondary metabolites; among them, aflatoxin B1 is classified as a Group 1 carcinogen by the International Agency for Research on Cancer. Aflatoxins exhibit fluorescence under ultraviolet light. Kojic acid, another substance produced by Aspergillus species, has also been reported to exhibit fluorescence. These compounds are commonly detected using chromatographic methods and rapid test kits. This study aimed to classify aflatoxin B1 concentration in dried figs by analysing UV images using a convolutional neural network-based image processing approach, thereby reducing dependence on chromatographic methods. Fresh figs were sliced, dried, and inoculated with Aspergillus flavus (A. flavus) Changes in aflatoxin and kojic acid levels were monitored using chromatographic analyses over a 21-day period, at 7-day intervals, while images were acquired concurrently. Training, validation, and test datasets were generated using UV images of different aflatoxin B1 standard solutions at concentrations of 5 to 250 μg/L. The VGG16-based model achieved an overall precision, recall, and F1-score values of 0.96, 0.96, and 0.95, respectively. The model accuracy for fig samples ranged from 0.64 to 1.00, highlighting the potential of image-based analysis as a rapid screening approach. On day 0, no aflatoxin B1 was detected by HPLC, and no fluorescence was observed in the fig samples. Accordingly, the model classified all samples as aflatoxin B1-negative. In conclusion, the developed model demonstrated its ability to classify predefined aflatoxin B1 concentration levels, including the lowest class of 5 μg/L. The model has the potential to contribute significantly to the development of fast, low-cost, image-based analysis systems.

Toxins doi: 10.3390/toxins18110480

Authors: Elise Nacer-Khodja Anouschka Middelkoop Marie Gallissot Kowsigaraj Palanisamy Francesc Molist Regiane R. Santos

This study aimed to determine the effects of a multi-target intervention on intestinal health and production performance in weaned piglets fed diets contaminated with a mixture of Fusarium toxins. To mimic farm conditions, naturally contaminated ingredients were used. In brief, piglets were exposed to four diets contaminated with multiple mycotoxins for 35 days post-weaning. Two maize batches were selected to prepare the experimental diets to obtain low (L_MYC) and high (H_MYC) multi-contaminated diets, either supplemented or not with a blend of bentonite, yeast cell wall, gingko biloba, and silymarin, resulting in a 2 × 2 factorial design. Growth performance of the piglets was assessed for two feeding periods—days 0–14 and days 14–35 post-weaning—as well as for the overall experimental period (days 0–35). Furthermore, in vivo intestinal integrity was assessed on days 7 and 21 post-weaning by inoculating piglets with fluorescein isothiocyanate-dextran (FITC-d) via oral gavage and subsequent blood sampling. Faecal samples were collected at the same time points for untargeted metabolomic analysis. The H_MYC diet impaired the growth performance of the piglets during the first 14 days of the experiment and negatively impacted the intestinal barrier, and also led to intestinal stress on day 21 post-weaning. Dietary supplementation with the blend improved the intestinal integrity on day 21 post-weaning and counteracted the stress caused by the mycotoxins regardless of their levels in the diet.

Toxins doi: 10.3390/toxins18110479

Authors: Miranda K. Gibson Alonso Delgado Aurélien De Jode Marymegan Daly Nicolas Salamin Benjamin M. Titus

Venom evolution has traditionally been investigated using proteomic and transcriptomic approaches, which have provided critical insights into toxin composition and function but remain constrained by uneven taxonomic and geographic sampling. These methods rely on fresh tissue and targeted field collections, biasing venom discovery toward medically relevant, easily accessible, or frequently encountered species, while underrepresenting the extensive ecological and biogeographic diversity present in many venomous lineages. Within Cnidaria, sea anemones exemplify this gap: despite exhibiting at least 16 characterized toxin families with diverse functions, large portions of their phylogenetic diversity remain unsampled. Here, we introduce a complementary framework that leverages whole-genome shotgun sequencing and reference-guided annotation using museum and biodiversity collections, focusing on Clade Stichodactylina as a model system. By standardizing genome-based toxin annotation, we demonstrate high concordance and sequence conservation relative to proteomic and transcriptomic datasets, validating whole-genome data as a robust resource for venom gene discovery. Analyses of phospholipase A2 (PLA2) toxins further reveal high numbers of orthologs coupled with strong purifying selection, rejecting a simple birth-and-death model of venom evolution for this family. Together, our results highlight the value of genome-enabled approaches for expanding venom research beyond traditional sampling paradigms and underscore that evolutionary dynamics of venom genes are highly clade- and family-specific, cautioning against generalized models of venom evolution.

Toxins doi: 10.3390/toxins18110478

Authors: Ronald A. Jenner

Phenotypic plasticity mediates organismal responses to environmental variation and plays an important role in ecological adaptation. Because animal venoms are adaptations that interact directly with the external environment, it seems reasonable to expect that phenotypic plasticity could play a vital role in fine-tuning venoms to changing ecological circumstances. Yet, at first glance, little seems to be known about venom plasticity. A 2017 benchmark paper on scorpions claimed to have found the first evidence of adaptive plasticity in venom composition, but since then no studies that cite this paper have reported other convincing cases of venom plasticity. Adaptive venom plasticity is actually more widespread and more widely studied than is generally recognized. I will argue that this apparent lack of awareness results largely from the historical lack of an integrative venom concept, which has produced a literature that is fractured along taxonomic fault lines and different functional categories of venom. Here I synthesize the disconnected literature on venom plasticity to reveal that our knowledge is both deeper and broader than commonly appreciated. I underline the importance of using the phrase “venom plasticity” only for cases of nongenetic, environmentally induced intraspecific venom variation.

Toxins doi: 10.3390/toxins18110477

Authors: Veronica Ghionna Marilena Donnaloia Biancamaria Ciasca Marisa Florio Paola Bovio Nicola Ungaro Antonio Moretti Antonella Leone Lucrezia Cilenti

Saxitoxins (STXs) are potent neurotoxins associated with harmful algal blooms (HABs) and represent a major concern for seafood safety. While monitoring programs mainly target bivalve molluscs, information on STXs occurrence in other aquatic organisms remains limited. This study investigated STXs occurrence in marine invertebrates from the Lesina and Varano coastal lagoons (southern Italy) and its relationship with phytoplankton composition and environmental conditions. STXs were determined by enzyme-linked immunosorbent assay (ELISA) in the Pacific oyster (Magallana gigas), the Atlantic blue crab (Callinectes sapidus), and gelatinous zooplankton. Potentially STXs-producing dinoflagellates were mainly represented by Gymnodinium spp. and Alexandrium spp., including Alexandrium minutum. Temperature showed the strongest association with the genus Alexandrium, whereas Gymnodinium spp. was negatively associated with salinity, although these relationships were not statistically significant. STXs were detected in all investigated matrices at concentrations below the EU maximum level for bivalve molluscs, which was used only as a contextual reference. Among the investigated matrices, C. sapidus showed both the highest occurrence of STXs-positive samples and the highest toxin concentrations. Tissue-specific analyses revealed STXs in all analysed anatomical compartments, with the highest concentrations detected in the hepatopancreas. These findings suggest that STXs circulation may extend beyond traditional bivalve vectors and provide preliminary evidence supporting a multi-compartment approach for investigating toxin dynamics in Mediterranean transitional ecosystems.

Toxins doi: 10.3390/toxins18110476

Authors: Negar Taheriashtiani Arash Zamyadi

Improving water quality through the support of natural biological and biogeochemical processes has attracted increasing interest in aquatic system management. It is hypothesised that phytoplankton community restructuring intervention using amorphous silica and adsorbed micronutrients could support biological processes involved in nutrient uptake and transformation. This study evaluated ecological responses following silica-adsorbed micronutrient formulation (SAMF) dosing at two field sites in Australia and New Zealand using nutrient and phytoplankton indicators. Site A comprised a reservoir monitored across pre-dosing, dosing, and post-dosing phases, whereas Site B comprised a constructed wetland with treated and untreated monitoring locations. Temporal and regression analyses were used to examine nutrient dynamics and phytoplankton responses. At Site A, the full-dosing period coincided with lower cyanobacterial abundance, increased diatom abundance, and generally lower total nitrogen concentrations relative to pre-dosing conditions. Regression analyses indicated that, relative to pre-dosing conditions, the full-dosing period was associated with approximately 76% lower cyanobacterial cell counts (95% CI: 43.9–89.9%) and 35% lower cyanobacterial biovolume (95% CI: 16.8–48.6%), while diatom abundance was statistically significantly higher during the full-dosing period. In contrast, Site B exhibited weaker and less consistent responses, with no significant difference in cyanobacterial abundance between treated and control locations in the main model (p = 0.691), despite temporal variation in nutrient and phytoplankton indicators among monitoring locations. Overall, the findings provide field-based evidence that SAMF dosing may be associated with changes in nutrient dynamics and phytoplankton community structure under real-world conditions. However, responses were not consistent across sites, suggesting that treatment performance is likely influenced by local environmental conditions.

Toxins doi: 10.3390/toxins18110475

Authors: Leticia Machado de Araújo Nicole Coelho Lopes Igor Moreira de Almeida Kelma Maria Maia Gutemberg do Nascimento Oliveira Homero Vasconcelos Neto Karla do Nascimento Magalhães Rodrigo Castellari Gonzalez Gdayllon Cavalcante Meneses Elizabeth De Francesco Daher Camila Justa Furtado Geraldo Bezerra da Silva Junior Polianna Lemos Moura Moreira Albuquerque Alice Maria Costa Martins

Scorpion envenoming is primarily recognized as a neurotoxic syndrome, but increasing evidence suggests that inflammatory responses may contribute to its systemic effects. The clinical relevance of these responses in patients with mild envenoming remains poorly characterized. This study investigated serum interleukin-6 (IL-6) and its association with angiopoietin-1 (Ang-1) and angiopoietin-2 (Ang-2) in adults with mild Tityus stigmurus envenoming. A cross-sectional study was conducted in 80 adults treated for scorpion envenoming at a referral hospital in Northeast Brazil, between August 2025 and April 2026. Clinical, hematological, biochemical, and serum biomarker parameters were evaluated. IL-6, Ang-1, and Ang-2 were quantified by enzyme-linked immunosorbent assay. Patients were classified according to detectable or undetectable serum IL-6, using 7 pg/mL as the assay detection threshold. All patients had mild envenoming, no systemic clinical manifestations, and favorable outcomes. IL-6 was detectable in 17 patients (21.3%). Patients with detectable IL-6 had higher Ang-2 concentrations than those with undetectable IL-6 (3.94 vs. 1.03 ng/mL, p = 0.006) and a higher Ang-2/Ang-1 ratio (0.091 vs. 0.027, p = 0.003). Ang-1 concentrations did not differ between groups (p = 0.524). IL-6 showed a moderate positive correlation with Ang-2 (ρ = 0.549, p = 0.022). Conventional hematological, renal, hepatic, and pancreatic parameters did not differ significantly according to IL-6 status. These findings indicate that mild T. stigmurus envenoming may induce detectable inflammatory activation despite the absence of systemic clinical manifestations. The association between IL-6 and Ang-2 suggests a link between inflammatory and endothelial responses and supports IL-6 as a candidate marker for investigating the clinical effects of scorpion venom.

Toxins doi: 10.3390/toxins18110474

Authors: Atul Patel Bilal Chughtai Ahunna Ukah Nina L. Alesci Kimberly Becker Ifantides Grace Forde

OnabotulinumtoxinA (onabotA) is a neurotoxin that is FDA approved to treat multiple indications. However, data regarding the burden of these indications and onabotA utilization among older adults are limited. This analysis examined the prevalence of onabotA-approved indications and the proportion of those patients treated with onabotA in adults ≥65 years of age in Medicare Fee-for-Service (FFS; 2023) and Medicare Advantage (MA; 2022). Proportions of patients diagnosed with and treated for ≥2 indications were also estimated. Among FFS enrollees, 6.1% of patients were diagnosed with ≥1 indication and 7.0% of diagnosed patients received onabotA. Urological indications (3.2%) and spasticity (2.5%) were most commonly diagnosed, and <5% of patients diagnosed with either of these indications received onabotA. Among patients diagnosed with ≥1 onabotA-approved indication, 7.4% were diagnosed with ≥2 indications. Concomitant diagnosis of urological indications and spasticity was common (>80%), and concomitant treatment of these indications was among the lowest (~5%). Differences in diagnosis and onabotA treatment rates by age, sex, and Medicaid eligibility were observed. OnabotA treatment rates among MA enrollees were lower. Among older adults, the most commonly diagnosed onabotA-approved indications had the lowest onabotA treatment rates, suggesting a potential gap between disease burden and real-world treatment utilization.

Toxins doi: 10.3390/toxins18110473

Authors: Stephen W. Miller Ashifa Al Juwaiser Sakthivel Vaiyapuri

Bothrops lanceolatus, a viper species endemic to Martinique, is unusual among lancehead pit vipers in producing delayed thrombosis rather than venom-induced consumption coagulopathy. The molecular basis of the thrombotic phenotype of this venom, and the role of impaired fibrinolysis in particular, remains incompletely characterised. Therefore, this study assessed various aspects of the venom’s effects on blood clotting. B. lanceolatus venom was assessed by rotational thromboelastometry, a 96-well halo clot-lysis assay using tissue plasminogen activator (tPA), a cell-surface plasminogen activation assay, and platelet aggregometry. The impact of varespladib (a phospholipase A2 inhibitor), marimastat (a metalloprotease inhibitor), and the Fab antivenom (Crofab) in mitigating venom-induced effects was evaluated. The venom produced concentration-dependent procoagulant activity that was partially reversed by varespladib, marimastat, and Crofab. The venom did not lyse preformed whole-blood clots, but preincubation with tPA significantly impaired tPA-mediated fibrinolysis. In the cell-surface plasminogen activation assay, the venom unexpectedly enhanced rather than inhibited substrate cleavage, an effect attributed in part to non-specific venom protease activity. The venom did not induce platelet aggregation but selectively inhibited collagen-induced aggregation. B. lanceolatus venom impairs fibrinolysis at the level of polymerised fibrin while leaving the cell-surface plasminogen-activation pathway intact. Combined with its procoagulant activity, this fibrinolytic impairment provides a coherent mechanistic basis for the delayed thrombotic complications observed clinically. Small-molecule toxin inhibitors warrant further evaluation as adjuncts to species-specific antivenom.

Toxins doi: 10.3390/toxins18110472

Authors: Dominic Reisig Heba Abdelgaffar Dawson Kerns George David Buntin Tim B. Bryant Isaac L. Esquivel Scott H. Graham Jeremy K. Greene Anders Huseth Alana Jacobson Silvana Paula-Moraes Amanda Quadrel Francis P. F. Reay-Jones Phillip M. Roberts Ron Smith Sally V. Taylor Juan Luis Jurat-Fuentes

Helicoverpa zea Boddie (corn earworm or bollworm) is a primary target of transgenic crops expressing insecticidal Cry and Vip3Aa proteins from Bacillus thuringiensis (Bt). Widespread resistance to Cry toxins has increased reliance on Vip3Aa. We evaluated Vip3Aa susceptibility in 71 field populations of H. zea collected across the southeastern USA from 2018 through 2025 using diet-overlay bioassays of F1 progeny. Vip3Aa1 was used in 2018 and Vip3Aa39 from 2019 through 2025. Median lethal concentrations (LC50) were estimated and resistance ratios calculated relative to two susceptible laboratory colonies, a field-collected susceptible and the commercially available Benzon colony. Resistance ratios depended strongly on the susceptible reference used. Three populations evaluated in 2018 had resistance ratios ≥ 10 relative to the field-collected susceptible colony, whereas all Benzon-referenced resistance ratios were <1.3. Benzon-referenced resistance ratios increased significantly over time, whereas field-collected-susceptible-referenced resistance ratios decreased. In contrast, field-population LC50 values showed no temporal trend from 2019 through 2025, when all populations were evaluated with Vip3Aa39. These results provide no evidence of a sustained regional decline in Vip3Aa susceptibility in this region, but they highlight the importance of stable susceptible benchmarks, protein quality control, and complementary approaches for long-term resistance monitoring.

Toxins doi: 10.3390/toxins18110471

Authors: Ningning Li Caihong Zhang Zaw Lin Naing Ei Thinzar Soe Huan Yu Xing Liu Lizhen Chen Gemei Liang

Large-scale cultivation of transgenic Bt crops has led to target pest resistance to Bt proteins, especially Cry1Ac. Under persistent selection pressure, Helicoverpa armigera has evolved resistance to Bt proteins in both laboratory and field populations, highlighting an urgent need to comprehensively elucidate the molecular mechanisms underlying pest Bt resistance. Previous studies have confirmed that the eukaryotic translation initiation factor eIF2 plays a critical role in regulating Bt resistance. Our prior RNA-seq data showed significant differences in the expression level of eIF3L between susceptible and Cry1Ac-resistant strains of H. armigera. In this study, we further observed that the mRNA level of eIF3L was significantly reduced in larvae surviving Cry1Ac exposure. More importantly, heterologous expression of eIF3L increased the susceptibility of Sf9 cells to Cry1Ac. Consistently, knockdown of eIF3L was associated with reduced mortality in H. armigera larvae exposed to Cry1Ac. Together, our in vivo and in vitro results suggest that eIF3L is involved in Cry1Ac toxicity. Furthermore, eIF3L affects the expression of the Cry1Ac receptor genes CAD, ABCC2, and ABCC3. Collectively, these results indicate that down-regulation of eIF3L is associated with decreased susceptibility of H. armigera to Cry1Ac, providing novel insights into the molecular mechanisms underlying insect resistance to Bt proteins.

Toxins doi: 10.3390/toxins18110470

Authors: Jeongmi Moon ByeongJo Chun Eujene Jung Dongki Kim Yeonji Seong

Long-term renal outcomes following snake envenomation may vary across regions, yet remain poorly characterized in South Korea. We investigated the long-term risk of chronic kidney disease (CKD) following snake envenomation and snakebite-related acute kidney injury (AKI). This nationwide population-based cohort study used data from the Korean National Health Information Database (2012–2023). We matched 729 adults hospitalized for snake envenomation who received antivenom to 2916 individuals without snake envenomation (1:4). Using a 1-year landmark design, we assessed incident CKD with Cox proportional hazards models and Kaplan–Meier analysis. CKD occurred in 48 patients with snake envenomation and in 156 controls, with incidence rates of 7.16 and 5.59 per 1000 person-years, respectively. Snake envenomation was not significantly associated with incident CKD (adjusted hazard ratio [aHR], 1.15; 95% confidence interval [CI], 0.85–1.55). Among patients with snake envenomation, 73 patients (10.0%) had coded AKI during the index hospitalization. Snakebite-related AKI was associated with incident CKD (aHR, 4.25; 95% CI, 2.15–8.40), whereas the association with severe envenomation did not reach statistical significance. CKD developed in 17 of the 73 patients with AKI (23.3%). Thus, no statistically significant association was detected between snake envenomation and incident CKD overall, whereas coded AKI was associated with subsequent CKD among envenomated patients. New CKD events continued to occur throughout follow-up.

Toxins doi: 10.3390/toxins18110469

Authors: Regina L. Radan Raphael M. Kudela

Alexandrium catenella, a harmful algal species responsible for paralytic shellfish poisoning (PSP), is frequently found along the California coast. Although it is prevalent, the environmental and climatic variables underlying long-term variability in A. catenella abundance, bloom occurrence, and paralytic shellfish toxins (PSTs) in Monterey Bay, California, remain poorly understood. This study analyzed a 24-year dataset (2001–2025) collected weekly at the Santa Cruz Municipal Wharf in Santa Cruz, California. Principal component analysis (PCA) and generalized linear models (GLMs) were used to examine the relationships among environmental variables, basin-scale climate indices, and the presence of A. catenella and PSTs. Year-round (annual and seasonal (spring/summer, fall, winter) models indicated that the occurrence of A. catenella was associated with warmer, lower-nutrient conditions and lower discharge rates from the San Lorenzo River. Long-term variations in A. catenella and PST occurrences were also influenced by basin-scale climate patterns, particularly the PDO-ONI and NPGO climate modes. These results show that the dynamics of A. catenella in Monterey Bay are affected by environmental conditions, seasonal oceanographic variability, and large-scale climate processes.

Toxins doi: 10.3390/toxins18110468

Authors: Aashna Uppal Hams Bashir Parinita Arora Paul Kingpriest Trudie Lang

Snakebite envenoming is the world’s deadliest neglected tropical disease, predominantly affecting the most resource-limited regions. This scoping review aimed to map the global landscape of studies investigating the epidemiology of snakebite envenoming. Of the 1700 studies included in this review, nearly half were from authors based in high-income countries. While trends in authorship have changed over time, authors from low-income countries were consistently underrepresented in the body of literature. This underscores a critical gap between where the burden of snakebite lies and where the literature is coming from (which may, in part, reflect general geographic inequities in the literature databases). Further, there were variables that were well-reported across the body of literature, like age and sex of the person affected by snakebite envenoming. However, there were gaps related to the reporting of outcomes such as reproductive complications, qualitative outcomes, and costs associated with snakebite care.

Toxins doi: 10.3390/toxins18110466

Authors: Cristian Persu Irina Ciofu Tudor Proca Remus Nicolae Cartas Narcis Chirca Victor Cauni

Intradetrusor abobotulinumtoxinA (aboBoNT-A) is used off-label for refractory idiopathic overactive bladder (OAB), but real-world data on persistence are scarce. This study aimed to describe persistence and reasons for discontinuation in all treated patients and to report outcomes in those who continued treatment. We retrospectively reviewed all toxin-naïve patients who received a first injection of aboBoNT-A (500 Speywood units, 30 sites, trigone included) at a single center between 2013 and 2022; each injection was followed for at least 6 months. Of 232 patients, 97 (41.8%) received at least one repeat injection; the main reasons for stopping were loss to follow-up (17.7%), procedure cost (13.4%), adverse events (9.9%) and lack of efficacy (8.6%). Effectiveness and safety were assessed only in these 97 patients, at their last injection. The International Consultation on Incontinence Questionnaire–Urinary Incontinence Short Form (ICIQ-UI SF) score decreased from 16.7 ± 2.8 to 12.3 ± 2.6 at 2 weeks (p < 0.001). Mean post-void residual volume rose from 16 to 79 mL (p < 0.001); 30.9% had at least 100 mL, none exceeded 200 mL and none required catheterization. The interval to re-injection was under 6 months in 24.7%, 6–12 months in 37.1% and over 12 months in 38.1%. Limitations include the retrospective design, outcome data restricted to continuers and an early outcome assessment. AboBoNT-A appears effective in patients who continue treatment, but most patients discontinue, and cost is a modifiable barrier.

Toxins doi: 10.3390/toxins18110467

Authors: Hiba I. Sheikh Tibor Fülöp Mariana Murea

Kidney assistance therapy, a term that more accurately reflects the limited therapeutic capabilities of dialytic modalities conventionally referred to as kidney replacement therapy, sustains life by removing retained uremic toxins and excess fluid, while operating within a dynamic biologic milieu shaped by comorbidities, intercurrent illness, diet, medications, and the evolving course of residual kidney function [...]

Toxins doi: 10.3390/toxins18110465

Authors: Yerassyl Temirbekov Khaknazar Sadibek Kakima Kastuganova Arman Bolatov Piotr Skrzypacz Kanat Samarkhanov Sabina Marks Natasha S. Barteneva

Harmful algal blooms (HABs) produce potent neurotoxins, such as domoic acid (DA) and saxitoxin (STX), which negatively impact marine mammal health. To contextualize these threats within specific regional ecosystems, we conducted a comprehensive meta-analysis of algal toxin concentrations in stranded and non-stranded seals and sea lions reported between 1990 and 2025. Following a systematic screening of 2377 records, 15 studies met the criteria for inclusion in our systematic review and quantitative analysis. Both toxins displayed substantial variability among studies. Notably, the concentrations of DA found in feces were markedly higher than in other matrices, and feces appears to be a high-yield matrix for detecting recent exposure. Urine and serum showed moderate DA levels, while fetal fluids were among the lowest. Geographically, most studies originated from California and the Alaskan Arctic. Multi-matrix monitoring of algal toxins is required in remote or logistically challenging regions. We propose that exposure to algal and cyanobacterial neurotoxins is a testable contributor to recurrent, unexplained mass mortality events among Caspian seals (Pusa caspica) and that targeted toxicological screening within the Caspian Sea ecosystem is urgently needed.

Toxins doi: 10.3390/toxins18110464

Authors: Ting Cheng Feng Yang Jizhuang Fan Weijia Wu Zekun Huang Fang Li

Highly lethal Vibrio disease (HLVD) is a newly emerging shrimp disease caused by VTc toxin-producing Vibrio strains, designated VHLVD. VTc toxin, composed of VTcA, VTcB, and VTcC subunits, is a novel member of the Tc toxin family. Structural and functional studies of VTc toxin have been hindered by the lack of efficient purification protocols. Here, we established a simple method for purification of milligram-scale bioactive VTc toxin via anion-exchange chromatography. The procedures of cell lysis, sample pretreatment, and anion-exchange chromatography were optimized. This protocol yielded approximately 1.5 mg of high-purity toxin from 40 mL of bacterial culture (VHLVD strain vp-HL-202005). The composition of the product was confirmed by SDS-PAGE and LC-MS/MS. Purified VTc toxin was bioactive, inducing typical HLVD mortality in shrimp post-larvae. Transmission Electron Microscopy (TEM) and native PAGE analysis revealed two distinct structural populations, which may represent the club-shaped holotoxin (VTcA2-VTcB2-VTcC2) and the bell-shaped VTcA2 homopentamer. This simple and efficient method will greatly facilitate research on the fine structure and functional mechanism of VTc toxin and thus expand our knowledge of the Tc toxin family.

Toxins doi: 10.3390/toxins18110463

Authors: Lin Chen Jie Wang Xuezhen Ma Yanling Yang Weilang Lin Hongyun Qian Zhenhong Zhuang

Pseudomonas plecoglossicida is a Gram-negative opportunistic pathogen in aquaculture that infects several economically important fish species, including large yellow croaker, orange-spotted grouper and yellow drum. Due to its high incidence and mortality rates, visceral white spot disease (VWND) caused by P. plecoglossicida has become a major bacterial disease threatening the sustainable development of aquaculture and has caused huge economic losses worldwide. Owing to tremendous technological advances, such as dual RNA-seq, our understanding of P. plecoglossicida pathogenesis and associated host defense mechanisms has been greatly enhanced. Nevertheless, many pathogenecity-related mechanisms of P. plecoglossicida remain uncovered, including the functions of various unannotated bacterial virulence factors. Therefore, there is an urgent need to develop new theoretical and practical approaches to address the challenges of P. plecoglossicida. In this review, we summarize the current research advances in the virulence factors, prevention and control strategies, and host–pathogen interactions of this microbial pathogen. This review will contribute to a more comprehensive understanding of the pathogenesis of P. plecoglossicida and provide new insights and strategies for its prevention and control.

Toxins doi: 10.3390/toxins18110462

Authors: Rui Luo Dong Zhang Tao Li Rong Li Li Xue Xiaojun Ma Xianghong Du Yuxuan He

Zearalenone (ZEA) is an estrogenic mycotoxin that poses a potential threat to male reproductive health. This study investigated the age-dependent testicular toxicity of ZEA and its association with autophagy-related changes in male mice. Male C57BL/6J mice at different developmental stages, TM3 cells, and primary ovine Leydig cells were used to evaluate the effects of ZEA on testicular morphology, testosterone secretion, autophagy-related protein expression, and signaling pathways. ZEA induced seminiferous tubule disorganization and spermatogenic cell sloughing in 4–8-week-old mice, whereas no obvious testicular pathological changes were observed in 2-week-old mice. Serum testosterone levels showed a downward trend following ZEA exposure, although the differences were not statistically significant. Western blotting revealed changes in the expression of autophagy-related proteins in testicular tissue, including decreased levels of Beclin1 and LC3, accompanied by increased phosphorylation of PI3K and AKT at 2, 6, and 8 weeks of age and significantly increased mTOR phosphorylation at 6 and 8 weeks of age. In vitro, 40 μM ZEA significantly reduced the viability of TM3 and primary ovine Leydig cells and altered the expression of autophagy-related proteins. These changes were accompanied by increased phosphorylation of PI3K and AKT, whereas the increase in mTOR phosphorylation was not statistically significant in either cell model. Collectively, these findings indicate that susceptibility to ZEA-associated testicular toxicity varies with developmental stage, with more evident histopathological changes observed in 4–8-week-old mice. ZEA exposure was also associated with alterations in autophagy-related protein expression and PI3K/AKT/mTOR pathway phosphorylation. Further studies are needed to determine whether ZEA alters autophagic flux and whether these signaling changes causally contribute to testicular toxicity and Leydig cell dysfunction.

Toxins doi: 10.3390/toxins18110461

Authors: Oky Setyo Widodo Dhidhi Pambudi Mirni Lamid Seiichi Uno Emiko Kokushi Osamu Yamato Masayasu Taniguchi Mitsuhiro Takagi

Zearalenone (ZEN), an estrogenic mycotoxin affecting cow reproduction, and deoxynivalenol (DON) are cytotoxic and immunotoxic, reducing protein synthesis. Contamination with ZEN and DON decreases the productivity of breeding cows. We evaluated the feasibility of commercial lateral flow immunochromatographic strip tests (LFISTs) as a rapid first-line screening approach for urinary ZEN and DON exposure in cattle. Using 81 samples from 13 Japanese Black (JB) and 14 Holstein Friesian (HF) herds, line intensity ratios were quantified by analyzing photographs of test and control lines within regions of interest (ROIs). LFIST correlated significantly with ELISA (ZEN/Cre: p < 0.001 and τ = 0.35; DON/Cre: p < 0.001 and τ = 0.31) and LC-MS/MS (ZEN/Cre: p = 0.002 and r = 0.35; α-ZOL/Cre: p = 0.013 and r = 0.27; β-ZOL/Cre: p = 0.007 and r = 0.30; ΣZEN/Cre: p = 0.005 and r = 0.31). The relationship between parameters was explored using the Weibull accelerated failure time model analysis. LFIST-ZEN and LFIST-DON were significantly associated with ELISA ZEN/Cre (p = 0.00057) and ELISA DON/Cre (p = 0.00061), respectively. To test whether LFISTs could predict mycotoxin contamination in feed, data from nine other herds (n = 38), with known feed mycotoxin levels, were analyzed. Estimation equations related feed ZEN and DON concentrations to urinary ZEN and DON levels in the herds. Our findings support the potential utility of LFIST as a rapid first-line herd-level screening tool for ZEN and DON exposure, providing screening information with LFIST results obtainable within approximately 15 min after initiation of the assay, while confirmatory laboratory analysis remains necessary when exposure is suspected.

Toxins doi: 10.3390/toxins18110460

Authors: Iara A. Cardoso Sophie Hall Thomas Crasset Mark C. Wilkinson Adam Robinson Bronwyn Rand Dakang Shen Connor E. Webb Richard Stenner David K. T. Waterhouse Ingeborg Hers Renaud Vincentelli Imre Berger Nicholas R. Casewell Loïc Quinton Alastair W. Poole Christiane Schaffitzel

Disintegrins are snake venom toxins that bind integrins and modulate platelet aggregation and cell migration. Isolation from venom often yields insufficient quantities for comprehensive study, making recombinant expression essential for detailed toxin characterisation. Here, we produced and characterised four naturally occurring disintegrins from Echis coloratus and Echis ocellatus in Escherichia coli. These homologous proteins contain distinct integrin-binding motifs: EcDis_RGD, EoDis_RGD, EcDis_KGD, and EcDis_VGD. Functional activities were evaluated using human platelet aggregation and endothelial cell scratch wound-healing assays. RGD-containing disintegrins strongly inhibited platelet aggregation, with EcDis_RGD showing the highest potency. The EcDis_KGD disintegrin inhibited platelet aggregation less potently, whereas EcDis_VGD showed no detectable activity in our assays. In endothelial cells, EcDis_RGD disintegrin markedly impaired cell migration, while EoDis_RGD displayed weaker anti-migratory activity. Neither KGD- nor VGD-containing disintegrins significantly affected wound closure. EoDis_RGD exhibited similar antiplatelet activity to the native, venom-purified disintegrin ocellatusin, validating our recombinant expression strategy. However, the corresponding PII-SVMP preparation displayed reduced inhibitory activity, consistent with incomplete generation of a fully mature disintegrin species. Our findings demonstrate the importance of both integrin-binding motif identity and surrounding sequence context in determining disintegrin function, and support further use of recombinant toxins in toxinology and as therapeutic lead molecules.

Toxins doi: 10.3390/toxins18100459

Authors: William T. Hay Susan P. McCormick Nathan D. Kemp Mark Busman Daren W. Brown Hye-Seon Kim Carson M. Andorf Stephen F. Harding Robert H. Proctor Martha M. Vaughan

Fusarium graminearum is a plant-pathogenic and mycotoxin-producing fungus of major concern to cereal production worldwide. There are genetically distinct populations of the pathogen that are associated with different trichothecene mycotoxin profiles. A recent study has identified key differences in genes associated with light response and circadian regulation among the major North American F. graminearum populations. It is currently unclear how these populations differ in their response to photoperiod (hours of light per day). Therefore, we evaluated the effects of photoperiod on growth and trichothecene production in isolates representing the three most prominent North American populations (NA1, NA2 and NA3) of F. graminearum. Photoperiod had a profound effect on fungal growth, colony morphology, and trichothecene production. This effect varied by substrate (i.e., fungal growth medium or plant host) and among pathogen populations. Photoperiod had a considerable impact on inoculated corn seedlings, and total trichothecene production was substantially greater (2–25×) in inoculated corn compared with wheat. Photoperiod had little effect on trichothecene production in inoculated wheat seedlings, except for NA3. Differences in trichothecene production were not due to differences in fungal biomass, but likely the result of photoperiod-dependent gene expression changes to trichothecene biosynthesis.

Toxins doi: 10.3390/toxins18100458

Authors: Xinxin Zhang Min Xu Bei Chen Shuilin Cai Qilin Huang Kun Qiao Zhiyu Liu

Wild pufferfish are abundant in the coastal waters of southeastern China, yet systematic data on their toxicity remain limited. This study collected 1230 specimens from Fujian, Guangdong, Guangxi, and Hainan and quantified tetrodotoxin (TTX) in nine tissues (eyes, skin, muscle, ovaries/testis, liver, digestive tract, kidneys, and heart) by ELISA. In total, 10 species were identified (Lagocephalus wheeleri, L. gloveri, L. inermis, L. lunaris, Takifugu xanthopterus, T. ocellatus, T. niphobles, T. poecilonotus, T. bimaculatus, T. oblongus). All tissues of L. wheeleri and L. gloveri contained TTX below the safety limit (2.2 μg/g), whereas the remaining eight species showed varying degrees of toxicity. T. bimaculatus and T. oblongus exhibited lethal-grade toxicity, with ovarian TTX reaching 320 μg/g and 393.56 μg/g, respectively. TTX was predominantly concentrated in the ovaries, liver, skin, and digestive tract, with substantial inter-individual variation. Marked sex-dependent differences and seasonal fluctuations were observed, driven by reproductive activity, water temperature, and prey availability. This study is the first to report that the wild T. bimaculatus from this region is a highly toxic species, and confirms the lethal toxicity of T. oblongus, providing a scientific basis for managing the consumption-related risks of wild pufferfish.

Toxins doi: 10.3390/toxins18100456

Authors: Liang Chen Jun Chen

Cyanobacteria, or blue-green algae, the first oxygenic photosynthetic organisms on Earth, play indispensable ecological roles as primary producers and drivers of biogeochemical cycles, and are also sources of human food, nutraceuticals, and biotechnological products. However, cyanobacterial harmful algal blooms (CyanoHABs), intensified by eutrophication and global climate change, threaten water quality, biodiversity, and human health due to their production of diverse secondary metabolites, i.e., taste and odor (T&O) compounds and cyanotoxins. This Special Issue, “Ecology and Toxicology of Cyanobacteria and Cyanotoxins”, compiles 13 papers from 9 countries that collectively advance our understanding of these issues across multiple scales. Cyanotoxins, including microcystins, cylindrospermopsins, anatoxins, and saxitoxins, can cause various hazards to animals and humans, including systemic toxicity, hepatotoxicity, genotoxicity, carcinogenicity, reproductive/developmental toxicity, neurotoxicity, immunotoxicity, and endocrine disruption. CyanoHABs are governed by complex bottom-up and top-down regulation, and blooms and cyanotoxin production are closely linked, but not always consistent/synonymous. Environmental stressors, e.g., phosphorus limitation, cold shock, and grazing pressure, selectively favor toxigenic genotypes, or can increase the cellular cyanotoxin quota, although overall population growth is suppressed. This can be explained by the fact that cyanotoxins can help cyanobacteria to better adapt to environmental stressors. When blooms disperse, demise, or are disrupted by certain interventions, the reduced biomass may increase exposure to dissolved, extra-cellular cyanotoxins if intra-cellular toxins are released faster than they are degraded or removed. We term the uncoupling of biomass and toxicity as the “cyanobacterial biomass–toxicity paradox”. We propose a “3P framework”, i.e., prediction, prevention, and prioritization, for the evidence-based risk management of CyanoHABs and cyanotoxins. We also advocate for more studies using multi-omics and artificial intelligence, as well as multi-disciplinary, multi-sectoral, and multi-stakeholder collaboration under the “One Health” umbrella to translate science into actionable solutions for sustainable development.

Toxins doi: 10.3390/toxins18100457

Authors: João Victor Batista da Silva Jéssica Cristina Leme Giovana Perez Montenegro Deisy Mara Silva Longo Marlei Josiele Augusto Fernando Silva Ramalho Lucas Gabriel Dionisio Freire Carlos Humberto Corassin Fernando Gustavo Tonin Sher Ali Carlos Augusto Fernandes de Oliveira Leandra Náira Zambelli Ramalho

Aflatoxin (AF) exposure during pregnancy may compromise maternal health and fetal development. This study aimed to evaluate the effects of gestational exposure to AF in pregnant Wistar rats and the mitigating effect of a functional yogurt (FY) containing inactivated biomass of Lacticaseibacillus rhamnosus and Saccharomyces cerevisiae. Animals were exposed to total AF (sum AFB1, AFB2, AFG1 and AFG2: 1.0 mg/kg/day) from gestational day (GD) 6 to 15, with or without FY supplementation. Maternal and reproductive parameters, fetal development, cranial ossification and placental growth factor (PlGF) immunostaining were evaluated, along with determination of AF metabolites in maternal and fetal samples. AF exposure reduced maternal weight gain during GD6–GD15 and fetal weight at GD20. Delayed cranial ossification occurred in 51.4 and 15.6% of fetuses in AF and AF + FY groups, respectively, with most severe alterations occurring in the AF group. Serum aflatoxin M1 (AFM1) concentrations were lower (p = 0.002) in AF + FY than in AF groups. AF metabolites were quantified less frequently in maternal liver from FY-supplemented animals (p = 0.015), while placentas or fetal livers from the AF + FY group had no quantifiable levels of AF metabolites. PlGF immunostaining was highest in AF group and lowest in the FY group (p = 0.0007). In conclusion, FY containing L. rhamnosus and S cerevisiae biomasses attenuated some AF-induced effects on fetal development, particularly delayed cranial ossification.

Toxins doi: 10.3390/toxins18100455

Authors: Fernando L. Vera-Baquero Lorena González-Gómez Isabel Sierra Sonia Morante-Zarcero

This review examines recent advances in the enantiomeric separation of tropane alkaloids (TAs), focusing on atropine, a racemic mixture of (+)- and (−)-hyoscyamine whose enantiomers differ significantly in pharmacological activity, with (−)-hyoscyamine being the more potent antimuscarinic agent. Enantiomeric purity and racemisation control are therefore essential in pharmaceutical formulations, biological samples, and food, where TAs are regulated contaminants due to their toxicological effects. The review covers analytical strategies reported from 2010 to 2026, with emphasis on high-performance liquid chromatography (HPLC), often coupled with tandem mass spectrometry (MS/MS) to improve sensitivity and selectivity. Polysaccharide-based chiral stationary phases operated in normal-phase or polar organic modes, usually with amine additives, represent the most widely employed approach. Protein-, cyclodextrin-, and antibiotic-based chiral selectors have also demonstrated successful enantiomeric resolution. Sample preparation is particularly challenging for food matrices, requiring efficient extraction and clean-up while preventing enantiomeric racemisation. Published strategies include solid-phase extraction (SPE), matrix solid-phase dispersion (MSPD), and QuEChERS protocols. Despite significant analytical advances, chiral analysis of TAs in foods remains underexplored. Recent studies have reported (−)-hyoscyamine concentrations exceeding regulatory limits in herbal infusions, underscoring the need for further research to support European Food Safety Authority (EFSA) recommendations and improve consumer risk assessment.

Toxins doi: 10.3390/toxins18100454

Authors: Yıldırım Cesaretli Özcan Özkan

Poison-centre records can inform toxicovigilance, but their interpretation depends on stable definitions and complete fields. We audited scorpion-sting data published by Turkey’s National Poison Control Centre (UZEM) and mapped record-design features in international publications. The detailed 2014–2018 appendix was analysed separately from 2019–2020 tables. It contained 7055 records; 51.0% involved males, 48.8% females, and 0.2% had unreported sex. May–September accounted for 74.1%, with July highest (17.9%). The source’s unequal age intervals initially favoured wider adult categories; after harmonisation into 10-year bands, ages 0–9 became the largest band in every year and represented 21.8% overall. Aggregate tables separately reported 1451 records in 2019 and 1729 in 2020. The audit covered 191 analytic ledger entries across 39 variables and identified numerical conflicts, changing definitions, and gaps in clinical linkage. Clinical tables were available only for 2017–2018, with no symptom recorded for 50.6% and 37.6% of annual records, respectively. Management fields documented recommendations and limited post-antivenom feedback, but not complete linked administration, severity, disposition, or outcomes. We propose a draft field framework prioritising event identity, clinical trajectory, treatment, disposition, and outcome. The findings describe poison-centre reporting activity rather than population risk.

Toxins doi: 10.3390/toxins18100453

Authors: Akın Tuna Ayşenur Botsalı Mustafa Şen Okan Ali Aksoy Özge Banu Öztürk Ercan Çalışkan

Although previous studies have demonstrated that ozone gas can inactivate botulinum toxin in liquid solutions, evidence regarding its effects under in vivo conditions remains limited. This exploratory study investigated whether local ozone injection is associated with earlier functional recovery from abobotulinum toxin A-induced muscle paralysis in a rabbit model. Three New Zealand White rabbits received bilateral intramuscular injections of 15 U abobotulinum toxin A into the anterior auricular muscles. After maximal paralysis was established, the right anterior auricular muscle received 1 mL ozone gas at a concentration of 10 µg/mL twice weekly for two weeks (four sessions in total), while the left side received room air injections as the concurrent control. Ear positions were assessed by twice-weekly imaging using a four-grade visual paralysis scale (Grade 0–III) until baseline position was restored. All ozone-treated ears improved from Grade III to Grade II by day 7 and returned to their baseline positions (Grade 0) by day 10, remaining at Grade 0 thereafter. In contrast, the room-air control ears remained at Grade III through week 3, showed progressive improvement during weeks 4 and 5, and returned to Grade 0 by week 6. Previous findings using the same experimental model without additional intervention reported persistence of paralysis for approximately 16 weeks. This study provides the first in vivo experimental evidence suggesting that local ozone administration may be associated with earlier functional recovery from abobotulinum toxin A-induced paralysis. However, larger, randomized, and objectively assessed studies are required to confirm this observation and clarify the underlying mechanisms.

Toxins doi: 10.3390/toxins18100452

Authors: Josh A. Garber Damian Shea Juliette L. Smith

The co-occurrence of phycotoxins produced by harmful algal blooms is becoming more frequent. To combat this problem, monitoring programs that can track and quantify a wide range of hydrophilic and lipophilic phycotoxins are required. A 24 h, bench-scale study was conducted to provide an initial comparison of apparent sorption and extraction recovery for 24 freshwater and marine phycotoxins from two passive samplers under controlled artificial seawater conditions. Modifications were made to previously published ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) methods to capture all phycotoxin analytes of interest in two injections. This analytical method was also evaluated alongside extraction methods for a commonly used passive sampler, a solid phase adsorption toxin tracker (SPATT), and a novel passive sampler, the composite integrated passive sampler (CIPS). SPATT contained a lipophilic resin, Diaion® HP20, whereas the CIPS is a continuous sheet with hydrophilic and lipophilic regions aimed at allowing for adsorption across a wider range of phycotoxins. A single-step extraction procedure was tested for both samplers to determine apparent extraction recoveries for all 24 phycotoxins. The CIPS seemed to produce overall higher apparent extraction recoveries of analytes, but the receiving phase generally sorbed less ng of phycotoxin during the experimental period than SPATTs and matrix effects were present. Across all lipophilic and hydrophilic phycotoxins, SPATT sorbed more than 90% of each phycotoxin from the seawater within 24 h, for all but four phycotoxins. The CIPS, however, sorbed less of each, but still more than 50% in artificial seawater for all but three phycotoxins. When considering extraction efficiency, SPATT may be better suited for hydrophilic phycotoxins, while the extraction method for CIPS appeared more efficient for lipophilic compounds, but further investigations are required to investigate potential matrix effects. The utility of the novel CIPS was preliminarily demonstrated in the laboratory, with seemingly relatively improved performance for particular phycotoxin classes (Azaspiracids, Dinophysistoxins/Pectenotoxins, Microcystins, and Brevetoxin) as compared to SPATT. The expanded UPLC methods and CIPS must be trialed in the field next to determine their applicability to monitoring, research, and management. The benchtop work, however, already suggests that users should consider the full range of phycotoxins present at an environmental site and create sampling schemes appropriate for their region given the different sorption and extraction efficiencies of these two samplers.

Toxins doi: 10.3390/toxins18100451

Authors: Avra Melina Madella Carlon Catharina Emma van Gerven Elena Sendino Garví Jeroen van Bergenhenegouwen Johan Garssen Rosalinde Masereeuw Saskia Adriana Overbeek

Chronic kidney disease (CKD) is increasingly linked to intestinal barrier dysfunction, which contributes to systemic inflammation and disease progression. We investigated whether 2′-fucosyllactose (2′-FL), a human milk oligosaccharide, could modulate intestinal epithelial barrier function under uremic toxin exposure and examined the role of the aryl hydrocarbon receptor (AhR). Using wild-type (WT) and AhR knockout (AhR KO) Caco-2 cell lines, we assessed barrier function and cellular responses through transepithelial electrical resistance (TEER), paracellular permeability (Papp), tight junction (TJ) gene expression, AhR target gene activation, and metabolic assays. AhR deficiency impaired epithelial maturation, confirming its role in barrier development. Short-term exposure to uremic toxins (UTs) did not significantly affect TEER or paracellular permeability under the conditions tested. However, 2′-FL reduced paracellular permeability in both WT and AhR KO cells in the presence of FCS. These effects occurred without changes in TJ gene expression, indicating mechanisms beyond transcriptional regulation. AhR target genes were not induced by UTs, suggesting limited receptor activation, and metabolic activity was unaffected. Overall, 2′-FL reduced paracellular permeability under selected experimental conditions, while the impaired barrier development observed in AhR-deficient cells supports a role for AhR in intestinal epithelial barrier development.

Toxins doi: 10.3390/toxins18100448

Authors: Woon-Kyong Chung Min Joo Kim

More aesthetic patients now ask for changes in facial contour: a smaller face, a sharper jawline, and less sagging. Botulinum toxin (BoNT) is best known as a treatment for wrinkles, but it can also be used for modifying facial contour. With BoNT alone, a wide face can be made narrower, a heavy lower face can be made lighter, sagging on the jawline can be reduced, and a short chin can be corrected. Many physicians already do this, but each site is taught and studied on its own. This review brings the sites together under the name “contouring BoNT”. It describes seven sites: five muscles (masseter, temporalis, platysma, depressor anguli oris, mentalis) and two salivary glands (parotid, submandibular). The seven sites work in three ways: reducing the volume of a muscle or gland, removing a downward pull so that the face looks lifted, and relaxing an overactive muscle that changes the shape of the chin. All three serve one goal that patients describe in their own words: a smaller and lifted face. For each site, the review explains what it does to the contour, who is a good candidate, who needs caution, the published dose range and the authors’ practice, and which neighboring site changes when the target is treated. It explains which sites are checked together and how each is examined. Two safety points specific to contouring—the immunogenicity of large repeated doses, and salivary gland treatment—are discussed separately. The framework provided is a checklist for assessment, not a rule stating that every patient needs all seven sites to be treated.

Toxins doi: 10.3390/toxins18100449

Authors: Luyao Wang Li Ding Tianxiang Qin Xinyu Liu Chenglan Lou Hongyi Luo Fang Sun Chenhu Qin Xiangdong Ye Xudong Luo Zongyun Chen

Kunitz-type peptides from venomous and blood-sucking animals represent an important natural molecular resource for anticoagulant development towards XIa, a new target for antithrombotic drug discovery. However, the plasmin-inhibitory activity of these Kunitz-type XIa inhibitory anticoagulants limits their application. Here, using the snake Kunitz-type toxin analogue BF9-N17H, an XIa inhibitor with a unique P1-site histidine, as a template, P2′ scanning was performed to enhance its selectivity towards XIa. Ten BF9-N17H analogues were designed using different amino acid classification replacement at the P2′ site, including non-polar amino acids A and P, acidic amino acids D and E, polar amino acids S, N, and Y, and basic amino acids K, R, and H. Coagulation factor inhibitory experiments showed that three BF9-N17H analogues, BF9N17HP2, BF9N17HK8 and BF9N17HR9, lost their XIa inhibitory activity, but seven other analogues, BF9N17HA1, BF9N17HD2, BF9N17HS4, BF9N17HN5, BF9N17HY6, BF9N17HE7 and BF9N17HH10, effectively maintained their inhibitory activity towards XIa, with IC50 values from 61.19 ± 7.04 nM to 4.54 ± 3.65 nM. One analogue, BF9N17HD2, inhibited XIa with an IC50 of 6.19 ± 1.38 nM and had a selectivity index of 142.79 towards plasmin. It only had weak inhibitory activity toward plasmin, with an IC50 of 883.88 ± 148.35 nM. Excitingly, five analogues, BF9N17HS4, BF9N17HN5, BF9N17HY6, BF9N17HE7 and BF9N17HH10, almost lost their plasmin-inhibitory activity completely, indicating their high selectivity towards XIa. To the best of our knowledge, this is the first report of Kunitz-type XIa inhibitors without plasmin-inhibitory activity, and it highlights that the Kunitz-type peptide scaffold might be used as a potential candidate for the development of potent and selective XIa-inhibitory antithrombotic agents.

Toxins doi: 10.3390/toxins18100447

Authors: İsmet Bulut Zeynep Yegin Katran Özge Argın Mustafa Katran Fatma Çakmak

Venom immunotherapy (VIT) is the only disease-modifying treatment for Hymenoptera venom allergy, but real-world data describing VIT-related reactions and individualized premedication practices remain limited. We retrospectively evaluated 16 adults undergoing VIT with a target maintenance dose of 100 µg administered every 4 weeks at a tertiary allergy center between January 2024 and December 2025, focusing on clinical characteristics, premedication, treatment-related reactions, and field-sting outcomes during follow-up of up to 24 months. All patients had severe index reactions (Müller grade III-IV); 13 (81.3%) received Vespula VIT and 3 (18.8%) honey bee VIT. Premedication was initiated before VIT in seven patients (43.8%), including omalizumab in five (31.3%). During the 7-week up-dosing phase, five patients (31.3%) developed local reactions, whereas no systemic reactions occurred (0/16; 95% CI: 0–20.6%). During maintenance, one systemic reaction occurred (6.3%), requiring adrenaline and one-night intensive care admission, after which VIT was discontinued. Two patients were re-stung during follow-up and developed only local reactions. In this small, selected real-world cohort, no systemic reactions were observed during the up-dosing phase, whereas one systemic reaction occurred during maintenance. Given the limited sample size, wide confidence intervals, and frequent use of individualized premedication, these findings should be interpreted as descriptive observations and not as evidence establishing the safety of VIT or the effectiveness of premedication.

Toxins doi: 10.3390/toxins18100450

Authors: Rui Xie Bei Qin Naila Khan Bangash Mingyao Xie Kunlong Yang Fengqin Song

Pyruvate carboxylase (PYC), a multifunctional biotin-dependent enzyme, is essential in diverse metabolic cascades including gluconeogenesis and the tricarboxylic acid (TCA) cycle. This study aimed to dissect the physiological roles of PYC and its succinylated site in Aspergillus flavus (A. flavus). In A. flavus, biological functions of PYC were characterized through gene deletion and site-directed mutagenesis. Phenotypic analysis and qRT-PCR demonstrated that PYC positively regulated sporulation and sclerotial formation, while negatively regulating aflatoxin B1 biosynthesis. The transcriptomics revealed that development-related DEGs of abaA, wetA, vosA, atfB, creA, crzA, and areA were downregulated, whereas veA and are2 were increased in Δpyc1. Meanwhile, aflatoxin-related DEGs of nor1, aflW, agiB, aflO, and aflM were upregulated, while cfoC and ucdC were lower in Δpyc1. In addition, a succinylated site on PYC (K1068) was characterized by noncovalent interactions. Combined with the transcriptomics analysis of this site, this study revealed that DEGs in pyc1K1068R (mimic desuccinylation) were enriched in fundamental metabolic pathways of glycolysis/gluconeogenesis, carbon metabolism, fructose and mannose metabolism, pentose phosphate pathway, etc., which mainly affected A. flavus development. This research provided insights into the regulatory mechanisms underlying A. flavus development and mycotoxin contamination.

Toxins doi: 10.3390/toxins18100446

Authors: Dario Lucchetti Daniela Delfino Marta Senofonte Laura Spinaci Rocco Baccelliere Alessandro Ubaldi Francesca Longo

The presence and accumulation of marine biotoxins, including paralytic shellfish toxins (PSTs) and tetrodotoxin (TTX) analogues, in marine organisms poses a significant food safety concern. This study describes the validation of a quantitative confirmatory HILIC–LC–MS/MS method for the simultaneous determination of a broad panel of marine toxins in four matrices: mussels, clams, sea snails, and sea urchins. The method covered 17 target compounds, including TTX analogues (TTX, 4-epiTTX, and 4.9-anhTTX), saxitoxin and analogues (STX, dcSTX, NEO, and dcNEO), gonyautoxins (GTX 1–4, GTX 2–3, GTX 5, GTX 6, and dcGTX 2–3), and sulfocarbamoyl toxins C1–C2. Particular attention was devoted to the evaluation of matrix effects (ME), a critical factor affecting LC–MS/MS quantification. The method was validated according to SANTE/11312/2021 guidelines, showing satisfactory selectivity, precision (RSD ≤ 20%), and accuracy (recoveries within 70–120%). Significant matrix-dependent signal suppression and enhancement were observed among the investigated species. Matrix-matched calibration enabled reliable quantification in mussels, clams, and sea snails, whereas sea urchin samples showed greater analytical complexity. The validated method provides a robust approach for the determination of PSTs and TTX analogues in different marine organisms and supports its application in official monitoring programs.

Toxins doi: 10.3390/toxins18100445

Authors: Yo Han Lee Ye Rin Bae Tae Hong Kang Jung Woong Yoon Seung Joon Lim Chae Hyun Lee Suhyeon Yun Sang In Lee

Fumonisin B1 (FB1) is a mycotoxin commonly found in animal feed, particularly in corn-based products. Exposure to FB1 can damage multiple organs, including the intestine, but its molecular effects on intestinal epithelial cells remain incompletely understood. This study investigated FB1-induced toxicity in porcine intestinal epithelial (IPEC-J2) cells and screened natural products for protective effects. Exploratory gene expression profiling identified genes showing predefined expression changes following FB1 exposure, and subsequent Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses indicated associations with apoptosis-related processes. Annexin V/propidium iodide staining showed increased apoptosis-associated staining, while quantitative real-time polymerase chain reaction (qRT-PCR) showed altered expression of BCL2, CASP3, and CASP9. To identify protective candidates, 332 natural products were evaluated using high-throughput screening. Among the screened extracts, C. communis showed the highest cell viability in the primary screening and was selected for further investigation. Co-treatment with C. communis was associated with reduced apoptosis-associated staining and partial counteraction of FB1-associated changes in apoptosis-related gene expression. These findings suggest that C. communis warrants further investigation as a natural product for mitigating FB1-associated intestinal epithelial toxicity.

Toxins doi: 10.3390/toxins18100444

Authors: Kyu-Ho Yi Kyu-Lim Lee

Botulinum neurotoxin type A (BoNT-A) dosing in facial aesthetics varies across anatomical targets, formulations, and treatment indications. This structured narrative review examined published dose ranges and the potential relevance of muscle thickness, target depth, functional anatomy, and injection localization to individualized treatment planning. English-language regulatory, clinical, anatomical, ultrasound, and consensus literature available through 13 April 2026 was considered. Evidence was interpreted according to source type, distinguishing approved product-specific regimens, controlled clinical studies, observational reports, and anatomy- or expert-derived recommendations. Muscle bulk may inform masseter contouring, but the literature does not establish a validated thickness-based dosing equation applicable across facial muscles. For facial expression muscles, contraction patterns, treatment objectives, and preservation of adjacent function are central considerations. Evidence for aesthetic dosing of the temporalis and salivary glands remains limited, and glandular indications require separate assessment from skeletal muscle treatment. Anatomically informed placement has a plausible mechanistic rationale, whereas evidence that motor-zone targeting consistently reduces toxin requirements is insufficient. Ultrasound may assist selected anatomically complex or previously treated cases but is not established as necessary for routine injections. Safe dose selection requires formulation-specific interpretation, indication-specific evidence, individualized anatomical assessment, and recognition of the limitations of existing studies.

Toxins doi: 10.3390/toxins18100443

Authors: João G. Brás Fokko P. Wieringa Babette H. Lentferink Simon Kootwijk Albert H. Westra Leonard J. van Schelven Mies van Steenbergen Joachim Jankowski Marianne C. Verhaar Karin G. F. Gerritsen Jeroen C. Vollenbroek

Current hemodialysis (HD) methods inefficiently remove protein-bound uremic toxins (PBUTs) due to their high binding affinity to plasma proteins, mainly albumin. This study aimed to investigate electromagnetic (EM) fields and temperature as potential extracorporeal, non-systemic methods to increase HD PBUT clearance. PBUT release was measured in infusible (caprylate-stabilized) albumin solutions spiked with indoxyl sulfate (IS) using an in vitro dialysis setup including a real-time, in-line IS-sensor. No measurable influence on PBUT release was observed during EM field-tests (f = 1–600 MHz, E <= 1 kV/m), but temperature tests (35–40 °C range) showed an absolute increase in IS free fraction of 0.7%/°C. Furthermore, static equilibrium dialysis tests, including IS, p-cresyl sulfate (pCS) and hippuric acid (HA), showed a free fraction increase for all three PBUTs upon a temperature increase from 37 °C to 43 °C (IS 1.30-fold, pCS 1.12-fold, HA 1.15-fold). Mathematical modeling showed a 1.14-fold decrease in plasma IS concentration after 5 weeks of HD, and exploratory hemocompatibility tests did not reveal pronounced changes over 15 heating cycles at 43 °C under static conditions. Extracorporeal temperature elevation is a promising technique to enhance PBUT removal during hemodialysis, which may be easily combined with current HD machines, does not break sterility, and avoids elevating free PBUTs inside the body.

Toxins doi: 10.3390/toxins18100442

Authors: Andrew D. Turner Karl J. Dean Monika Dhanji-Rapkova Alison O’Neill Benjamin H. Maskrey Patrycja Stubbs David M. Hartnell Shamina Page Naomi Baker Nadine Morrell John M. Turner Joan Turner Florence R. I. Turner Stephen D. Gregory Adam M. Lewis Katy Lewis

Cyanobacteria are microorganisms that can produce natural toxins with significant impacts on human, animal and ecosystem health. Portishead Marine Lake is a small freshwater lake in southwest England frequented by thousands of people each year for recreational activities but was associated periodically with prolific cyanobacterial blooms with potential risk to visitor health. Water samples were tested weekly between January and December 2016 for cyanobacteria and cyanotoxins. Citizen scientists undertook testing for nutrients, dissolved oxygen, pH, temperature and meteorological parameters. Four major blooms of Microcystis spp. were observed in the warmer months between May and September and were associated with high concentrations of microcystins. Complex toxin congener profiles were dominated by MC-LR and MC-RR, with lower contributions from MC-WR and MC-YR. Cyanobacterial cell densities and summed microcystin concentrations ranged from 1600 to 5.6 × 106 cells/mL and 0.1 to 18,069 µg/L, respectively, and correlated significantly. Toxin concentrations were positively correlated with nitrate levels and inversely correlated with pH, although mean pH levels did not fall below 7.0. There was little statistical evidence for influence of weather conditions overall, although significant intra-lake differences in cell densities and toxin concentrations were noted. Intervention measures employed by local authorities included the use of barley straw, hydrological manipulation, debris removal and ultrasound treatment. Whilst no statistical assessment of intervention effectiveness was possible without effective control samples, there was some limited evidence for the temporary reduction in toxin levels after hydrological intervention, but none following installation of ultrasound. Overall, the study utilized LC-MS/MS and microscopy to quantify microcystins and Microcystis spp., respectively, and evidenced risks to lake visitors and the importance of toxin testing in recreational waters and the use of appropriate risk mitigation approaches. Further work incorporating appropriate experimental controls would be needed to determine the effectiveness of hazard mitigation measures for reducing or eliminating human health threats.

Toxins doi: 10.3390/toxins18100441

Authors: Veronica Russo Sara Ragucci Miriam Russo Maria Antonietta Oliva Nicola Landi Antimo Di Maro Antonietta Arcella

Background: Glioblastoma (GBM) is the most aggressive primary brain tumor and remains difficult to treat because of its highly invasive behavior and resistance to conventional therapies despite surgery, radiotherapy, and chemotherapy. Sodin 5, a novel type-1 ribosome-inactivating protein (RIP), inhibits protein synthesis and displays cytotoxic activity against several tumor cell types, making it a promising candidate for GBM therapy. Methods: The antitumor activity of sodin 5 was evaluated in the U87MG glioblastoma cell line and patient-derived primary GBM cell lines (CG, ZAR, and NULU). Cell viability, migration, and molecular changes were assessed after treatment. Biological activity was also investigated under different pH conditions. Vimentin and phosphorylated ERK (p-ERK) expression were analyzed to investigate the mechanisms underlying sodin 5 activity. Results: Sodin 5 significantly reduced cell viability in all GBM cell lines. Treatment also inhibited cell migration, accompanied by decreased vimentin expression, suggesting cytoskeletal remodeling and reduced invasive potential. Furthermore, sodin 5 decreased p-ERK levels, indicating modulation of signaling pathways involved in proliferation and motility. The protein retained biological activity under different pH conditions, supporting its potential efficacy in the heterogeneous GBM microenvironment. Conclusions: These findings identify sodin 5 as a promising therapeutic candidate for GBM by targeting multiple processes involved in tumor growth and invasion.

Toxins doi: 10.3390/toxins18100440

Authors: Stephany Young Yusty Celestino Aguilar Alexa Prescilla-Ledezma Alexander A. Martínez Claudia González Hildaura Acosta de Patiño

Snakebite envenoming is a major neglected public health problem in Panama, yet culture-independent data on the oral microbiota of medically important viperid snakes remain limited. We characterized the oral microbiota of four captive species native to Panama (Bothrops asper, Bothriechis nigroviridis, Cerrophidion sasai, and Porthidium lansbergii) using forward-read analysis of 16S rRNA V3–V4 amplicons and compositional methods. Among 68 retained oral samples, communities were dominated by Pseudomonadota, Bacillota, Actinomycetota, and Bacteroidota. Observed ASV richness differed among species (Kruskal–Wallis H = 21.87, p < 0.001), whereas no significant differences were detected in Shannon diversity or Pielou’s evenness. Richness differences require caution because sequenced negative controls were unavailable and no independent depth-aware sensitivity analysis was performed. Community composition was significantly associated with host species (PERMANOVA R2 = 0.197, p = 0.001), although PERMDISP detected heterogeneous within-group dispersion (p = 0.005), requiring cautious interpretation. ANCOM-BC2 identified 21 differentially abundant genera, including Providencia, Morganella, Pseudomonas, Stenotrophomonas, and Staphylococcus, with high relative abundance of Mycoplasma in a subset of P. lansbergii samples. These findings provide a culture-independent taxonomic baseline for future culture-based, antimicrobial-susceptibility, and paired snake-oral/human-wound investigations in Panama.

Toxins doi: 10.3390/toxins18100439

Authors: Xiaoyu Xiao Junxuan Zeng Songheng Zheng Shifeng Liu Qi Deng Mei Qiu Zhijia Fang Gaoshuang Hu Yaling Wang Youxin Wang

Microcystins (MCs) produced by cyanobacteria can contaminate aquatic foods, but drinking water surveillance does not directly characterize dietary exposure. This critical narrative and framework review evaluates environmental transfer, tissue residues, analytical coverage, toxicity, exposure assessment, and management for fish, bivalves, crustaceans, and related products, with a separate assessment of algal dietary supplements. Evidence is organized by species, consumed tissue, concentration basis, analytical fraction, congener coverage, and consumption scenario. Reported concentrations are not directly comparable when studies analyze different organs or use different extraction procedures and mass bases. Paired analyses of tilapia muscle illustrate how measurements limited to free congeners can miss a substantial additional burden, while shellfish studies show why clearance requires verification in the edible tissue. Structural and toxicological evidence explains why MC-LR provides a reference for assessment but does not fully characterize every mixture. We propose a framework linking environmental warnings to product sampling, validated quantification, explicit exposure scenarios, and proportionate management. Priorities are comparable occurrence reporting, interlaboratory validation, characterization of the oral relevance of bound fractions, and congener-specific toxicity data. Neither a positive screening result nor an exceedance of a chronic intake benchmark alone establishes the probability of illness from an individual meal.

Toxins doi: 10.3390/toxins18100438

Authors: Ling Wang

Mycotoxins, toxic secondary metabolites produced by filamentous fungi, represent a persistent challenge to global food safety, animal health, and human well-being [...]

Toxins doi: 10.3390/toxins18100437

Authors: Christopher Hernandez Vincent Merlhiot Selma P. Snini Florence Mathieu Vanessa Durrieu Jean-Denis Bailly

Aflatoxins are carcinogenic mycotoxins that can contaminate many crops and whose distribution is increasing due to climate change. Given the mutagenic and carcinogenic properties of aflatoxins, protecting public health requires minimizing dietary exposure. In the European Union, maximum regulatory levels can be as low as 0.10 µg/kg for AFB1 in processed cereal-based foods and baby foods. Once produced, aflatoxins are stable, so the best way to protect consumers is to minimize aflatoxin production. Numerous studies have investigated sustainable and eco-friendly strategies to limit aflatoxin contamination in food and feed. These strategies rely on microorganisms or natural, plant-derived compounds that hinder the development of aflatoxigenic species and/or specifically inhibit toxin biosynthesis. However, a substantial translational gap remains between initial in vitro evidence of the effects on fungal growth and/or aflatoxin synthesis and the effective application of such biocontrol agents under field conditions. This review aims to clarify the current state of the art by presenting the diverse biocontrol strategies currently being investigated to combat aflatoxins. We discuss the advantages and disadvantages of such strategies and describe the level of technological readiness, which spans from in vitro tests to large-scale in-field applications.

Toxins doi: 10.3390/toxins18100436

Authors: Tamás Schieszl György Kövér Lucia Gambacorta Michele Solfrizzo Lajos Biró Györgyi Arató Andrea Zentai Brigitta Bóta András Szabó Melinda Kovács

Mycotoxins are common contaminants of the food and feed chain and have been associated with developmental disorders, hepatotoxicity, nephrotoxicity, and various types of cancer. This study aimed to estimate dietary mycotoxin exposure in the Hungarian population and to identify the major cereal-based food contributors. Urinary mycotoxin concentrations were measured by LC–MS/MS, and population exposure was assessed using estimated daily intake (EDI) values. Associations between food consumption and urinary mycotoxin levels were evaluated using Spearman correlation analysis on urine samples collected in year 2018 and 2020. The average EDI values were below the tolerable daily intake (TDI) for all mycotoxins in both years, except for ochratoxin A in 2018. Distinct exposure patterns were observed between the two sampling years. In 2018, consumption of oats and oat-based products showed a moderate positive correlation with urinary fumonisin B2 levels (r(s) = 0.497, adjusted p = 0.001962350574942). In 2020, wheat and wheat-based product consumption correlated with urinary deoxynivalenol levels (r(s) = 0.6868, adjusted p = 0.009509496), while corn and corn-based product consumption correlated with urinary zearalenone levels (r(s) = 0.6137, adjusted p = 0.04134226737). These findings highlight differences in dietary mycotoxin exposure between the studied populations and sampling periods and emphasize the importance of human biomonitoring to identify major dietary mycotoxin sources/carriers and support exposure and risk assessment.

Toxins doi: 10.3390/toxins18100435

Authors: David M. Simpson Marta Banach Jason Aldred Jack Lee William Ondo Fernando Pagan Anastasia Bagretsova Gudrun Klein Thorin L. Geister Irena Pulte Mandar Jog

This double-blind, placebo-controlled, phase II trial (NCT04766723) investigated the efficacy and safety of incobotulinumtoxinA (Xeomin®) in the treatment of essential tremor (ET) of the upper limb (UL). Adults with ET of the UL were randomized 2:1 (n = 78) and treated (n = 77) with intramuscular injections of incobotulinumtoxinA or placebo at semi-flexible doses into their motor-dominant arm. The primary endpoint was the change from baseline to week 6 in maximum tremor amplitude at wrist level. Baseline characteristics of treated patients were well matched across treatment arms. The primary endpoint was not met; maximum tremor amplitude at the wrist decreased from baseline to week 6 in both groups, with changes numerically favoring placebo (least squares mean treatment difference 0.21, p = 0.28). At week 6, compared with the placebo group, the incobotulinumtoxinA group had improvement in The Essential Tremor Rating Assessment Scale (TETRAS) performance dominant UL score (p = 0.006) and Quality of Life in ET (QUEST) total score (p = 0.073). However, these secondary outcomes must be interpreted descriptively as exploratory findings. Although the p-value and the 95% confidence interval including zero did not support statistical significance, the observed numerical treatment difference in change from baseline in QUEST total score suggests that the treatment effect may be consistent with the minimal clinically important difference in favor of incobotulinumtoxinA. Improvement in TETRAS Activities of Daily Living UL score was similar in both groups at week 6 (p = 0.933). IncobotulinumtoxinA was generally well tolerated. Treatment of UL ET with incobotulinumtoxinA did not show superiority over placebo for the primary endpoint. Secondary and other efficacy endpoints should therefore be viewed as exploratory, generating hypotheses suggesting potential improvements in TETRAS UL performance score and quality of life that require confirmation in further research. IncobotulinumtoxinA demonstrated a favorable safety profile without any new or unexpected safety concerns.

Toxins doi: 10.3390/toxins18100434

Authors: Hiromi Watari Koji Nakano Masaya Kitano Hiromu Kageyama Yoshikazu Tanaka Yuri Ishibashi Ryuichi Sakai

Soritesidine (SOR) is a potent proteinaceous toxin from the marine sponge Spongosorites sp. that is toxic to mammalian cells, brine shrimp larvae, sea hare embryos, and mice. Although SOR exhibits divalent metal ion-dependent DNase activity, its molecular identity and mechanism of action remained unknown. We determined the amino acid sequence of SOR and investigated its cellular mode of action. Degenerate PCR, 3′-RACE, inverse PCR, and nested PCR identified an open reading frame encoding a 947-amino-acid acidic protein. The deduced sequence showed no substantial similarity to known proteins except for approximately 30% sequence identity to a hypothetical protein from the marine α-proteobacterium Euryhalocaulis caribicus. Analysis of the AlphaFold 3-predicted N-terminal domain using Foldseek and structural superposition revealed a DNase I-like fold with a catalytic architecture similar to that of vertebrate DNase I. A recombinant N-terminal fragment (residues 1–310, N-SOR) exhibited DNA cleavage activity, supporting the assignment of this region as a DNase I-like endonuclease domain. The recombinant fragment lacked detectable cytotoxicity, suggesting that regions outside the N-terminal DNase I-like domain contribute to the cytotoxicity of SOR. Fluorescence imaging showed that SOR entered cells and accumulated in the nucleus, whereas its cellular uptake was attenuated at 4 °C, suggesting the involvement of an energy-dependent internalization pathway. Immunostaining revealed increased γH2AX signals in the nuclei of SOR-treated cells, indicating activation of a DNA damage response. Together, these findings identify SOR as an unusual multidomain protein toxin containing a functional DNase I-like domain and provide a molecular basis for further investigation of its cytotoxic mechanism.

Toxins doi: 10.3390/toxins18100433

Authors: Yucong Li Xiang Wan Fangqun Gan

Microcystin (MC) risk is often assessed within bloom-affected lakes and reservoirs, although hydrological connectivity can extend exposure into downstream river and estuarine networks. We synthesize evidence using a lake–river continuum framework that treats connected reaches as active processing environments. The framework separates the source pool, export flux, and downstream exposure, and distinguishes advective redistribution, reversible retention, irreversible transformation, and source or remobilization processes. Hydrology sets reaction opportunity, while phase partitioning, photochemical and microbial transformation, sediment exchange, and biological uptake determine how MC mass and bioavailability change during transport. Field evidence shows that downstream occurrence cannot be inferred reliably from visible source blooms: reaches may attenuate, retain, redistribute, or locally renew MC exposure depending on hydraulic state and process coupling. These dynamics shift exposure among water, particles, sediments, cells, and biota. Consequently, source-lake indicators or water concentrations alone can mischaracterize ecological exposure, and drinking-water guideline values are not ecological protection thresholds. Key priorities are flux-resolved monitoring, direct field estimates of process rates and transformation products, chronic ecotoxicology matched to realistic exposure routes, and hierarchical transport–biogeochemical models. This continuum perspective links source, process, pathway, receptor, and effect within a single source-to-risk framework.

Toxins doi: 10.3390/toxins18100432

Authors: Safy M. Sh. M. Abaza Jiaqi Zhou Ziqin Wang Zanyu Pan Haoran Ma Yingdi Sun Ruben K. Dagda Edward S. Gasanoff

Bee venom melittin is a small protein with significant pharmacological potential. In this study, AutoDock simulations identified Arg22 as a key residue of melittin responsible for targeting the phosphate groups of cardiolipin (CL). Recombinant melittin with the native amino acid sequence, as well as a modified version with Arg22 substituted by Gly22, were cloned, expressed, and purified. The effects of both melittins on proton adsorption in CL-containing membranes, mitochondrial ATP production, and the formation of non-bilayer lipid structures were examined. Based on the array of computational and experimental data obtained in this work, together with published findings, we propose that Arg22, in tandem with Lys23, targets and neutralizes CL bound to conserved Lys7 and Lys43 of the ATP synthase rotor, thereby detaching CL from the rotor. These events promote a conical shape of CL, which triggers the formation of inverted micelles that entrap protons from the Fo subunit and transfer them to the proton pump of a respiratory complex. This study advances both the conceptual framework of mitochondrial energetics and the translational application of natural toxins. Specifically, the findings support the development of new pharmaceuticals aimed at counteracting the loss of mitochondrial energy production that occurs with aging and pathological conditions.

Toxins doi: 10.3390/toxins18100431

Authors: Yuval Levy Ayalon Reis Galit Blecher Oksana Daron Sara Dadon Raz Zarivach Noam Zilberberg

Kv4 channels generate transient A-type potassium currents, but the contribution of Kv4.1 remains difficult to define because few pharmacological tools discriminate among Kv4 isoforms. We investigated the molecular basis of Kv4.1-selective inhibition by BlTx2, an α-KTx15 scorpion toxin from Buthacus leptochelys. Using electrophysiology, mutagenesis, tandem-channel analysis, structural modeling, and evolutionary analysis, we mapped the determinants governing BlTx2 activity and selectivity. Substitution of conserved α-KTx residues Lys27 and Tyr36 abolished detectable inhibition, whereas N30A reduced potency, supporting a canonical pore-blocking mechanism. A Kv4.1/Kv4.3 tandem chimera reduced BlTx2 potency, consistent with a contribution of adjacent Kv4.1 subunits to the high-affinity toxin-binding surface. Channel mutagenesis identified the Kv4.2/Kv4.3 lysine corresponding to Kv4.1 Asn355 as a key barrier to BlTx2 activity, since introducing this lysine into Kv4.1 abolished detectable inhibition. Mutational mapping of BlTx2 revealed that pore-facing residues required for inhibition are separated from peripheral residues that tune potency and cross-reactivity. In particular, Q9K and I18K conferred Kv1.2 activity, partly through interaction with Kv1.2 Glu355. Evolutionary analysis linked conserved pore-facing residues to purifying selection, and peripheral determinants to diversification. These findings identify distinct conserved and peripheral determinants that together shape BlTx2 activity and selectivity, thereby establishing BlTx2 as a useful probe for Kv4.1 function.

Toxins doi: 10.3390/toxins18100430

Authors: Tomé Azevedo Lucas Ribeiro Joana Azevedo Rui Barbosa Rita Mendes Joana Martins Marisa Freitas Vitor Vasconcelos Alexandre Campos

Cyanobacterial blooms and the associated production of cyanotoxins represent a growing concern for freshwater ecosystems, particularly in regions increasingly affected by eutrophication and climate change. In Portugal, data on the occurrence and distribution of cyanotoxins remain limited, especially in southern multipurpose reservoirs that support the water supply and irrigation in a region severely affected by water scarcity. This study investigates the presence of cyanobacteria and their major cyanotoxin groups—microcystins (MCs), cylindrospermopsin (CYN), and anatoxin-a (ANTX)—in three reservoirs of the Alqueva water storage system (EFEMA), combining chemical analyses with molecular tools to assess toxin-producing cyanobacteria. Liquid chromatography–mass spectrometry (LC–MS) detected MC-LR, MC-RR, and MC-YR exclusively in the particulate fraction of water samples, while no toxins were detected in the dissolved fraction. MC concentrations were generally low (<0.1 µg/L), with the highest levels observed during summer, particularly in July. Among the variants, MC-LR was the most common and abundant. Molecular analyses (16S rRNA) confirmed the widespread presence of cyanobacteria and identified the genus Microcystis in all reservoirs. Several genes from the microcystin biosynthesis cluster (mcyA, mcyB, mcyD, mcyE, and mcyG) were amplified, and sequencing of the mcyA gene revealed high homology with Microcystis aeruginosa. Although the detected MC concentrations remained below World Health Organization (WHO) guideline values for drinking water, their recurrent detection and the confirmed presence of toxin-producing genotypes highlight the need for continued monitoring. This study underscores the value of integrating chemical and molecular approaches for assessing cyanotoxin risks in large multipurpose reservoir systems under increasing environmental pressure.

Toxins doi: 10.3390/toxins18100429

Authors: Stephen A. Ashford Mark Weatherall Klemens Fheodoroff Jorge Jacinto Richard J. Siegert Christian Hannes Lynne Turner-Stokes

Improvement in active function (making it easier to use the limb) is a common goal following treatment of upper limb spasticity with botulinum toxin (BoNT-A). ULIS-III was a large international, observational, longitudinal study (N = 953) conducted in the context of usual clinical practice over two years. In this secondary post hoc analysis, we examine whether goals for improvement in active function were met over repeated injection cycles. Patients were selected if improved active function (AF) was a primary/secondary goal for at least one cycle of treatment (n = 286/953). They were assessed at the start and end of each cycle using the Goal Attainment T-score alongside the Arm Activity measure (ArmA). Serial changes and predictors of improvement were explored. Across eight cycles, there were 1167 injections for active function, 875/1998 (44%) AF goals were set and 493 (57%) of those goals were achieved. Patients continued to show significant improvement in AF (p < 0.001) for each injection at least up to three cycles, which is likely to be clinically significant, especially in patients with milder spasticity who were naïve to BoNT-A. No statistically significant predictors were confirmed on mixed effects modeling. Conclusions: Repeated BoNT-A continued to produce significant improvement in active function for at least three cycles, regardless of patient- or treatment-related factors in this cohort, but type II errors due to small numbers cannot be excluded.

Toxins doi: 10.3390/toxins18100428

Authors: Kazuya Yoshida

Botulinum neurotoxin (BoNT) therapy has expanded from focal chemodenervation to a versatile therapeutic modality used across multiple medical specialties. The stomatognathic system is particularly suited to BoNT therapy because motor, sensory, articular, secretory, and aesthetic functions converge within a compact anatomical region. Current and emerging applications include oromandibular dystonia, dyskinesia, bruxism, tremor, hemimasticatory spasm, embouchure dystonia, masticatory muscle and temporomandibular disorder-related pain, trigeminal neuralgia, recurrent temporomandibular joint dislocation, sialorrhea, Frey syndrome, masseter prominence, gummy smile, and perioral wrinkles. Evidence and regulatory status vary substantially among indications, and many applications remain off-label. Dentistry and oral and maxillofacial surgery have historically played a limited role in BoNT therapy despite their detailed anatomical and functional expertise in this region. However, interest is increasing, and the recent approval of onabotulinumtoxinA in Japan for masseter prominence in adults younger than 65 years represents an important milestone. This review synthesizes current applications, limitations, safety considerations, and future directions, and proposes the stomatognathic system as a major expanding frontier for BoNT therapy.

Toxins doi: 10.3390/toxins18100427

Authors: Christine Rasetti-Escargueil Stefano Palea

The present review is focused on a range of BoNT/A molecular targets and the subsequent modulation of cellular signaling pathways focused on gene regulation, skin and nerve regeneration, vascular effects, and contributions to pain relief. This brings fresh insights to the diversity of BoNT/A mechanisms of action, opening new avenues of research into therapeutic options for patients. A summary of the mechanisms of action of BoNT/A formulations will be presented in parallel with their specific indications, showing the versatility of the BoNTs’ efficiency through different pathways well beyond the canonical SNARE cleavage mechanism. The current research progress is summarized, ranging from the molecular mechanism of BoNT/A involved in the treatment of inflammation, pathological scars, pain, and a range of neurological and vascular disorders via clinical investigations and experimental studies.

Toxins doi: 10.3390/toxins18100426

Authors: Imene Ghorbal Sara Redouane-Salah Safa Smail Amandine Nachtergael Mohammed Aufy Mohamed F. Abdallah

Mycotoxigenic fungi, particularly species of Aspergillus, Fusarium, and Penicillium, contribute substantially to food contamination through the production of chemically diverse mycotoxins. Plant-derived essential oils (EOs) and extracts have therefore attracted increasing interest as potential alternatives or complements to conventional antifungal strategies. The genus Artemisia, which comprises several aromatic species with extensive biological properties, may constitute a promising source of such natural agents. This review critically examines the available data on the use of EOs and extracts from four Artemisia species (A. absinthium, A. campestris, A. herba-alba, and A. judaica) against mycotoxigenic fungi, with an emphasis on chemical composition, antifungal efficacy, mechanisms of action, and safety. Across the available studies, evidence suggests that Artemisia preparations frequently inhibit fungal growth, but their efficacy varies substantially according to species, geographical origin, plant part, harvesting conditions, extraction method, chemical composition, fungal species, and experimental assay. Importantly, evidence for direct inhibition of mycotoxin biosynthesis remains considerably more limited than evidence for fungal growth inhibition. Proposed antifungal mechanisms include disruption of the fungal cell wall and plasma membrane, altered mitochondrial function, and interference with cellular transport processes, although mechanistic evidence specifically for Artemisia preparations remains limited. Available toxicological data further indicate that efficacy and safety must be evaluated together, particularly for chemotypes containing potentially hazardous constituents such as thujone. Overall, Artemisia EOs and extracts may represent promising but not yet sufficiently validated candidates for food-associated mycotoxin control. Future studies could prioritize chemical standardization, mechanistic characterization, mycotoxin-specific endpoints, food-matrix validation, and integrated efficacy–safety assessment.

Toxins doi: 10.3390/toxins18100425

Authors: Tanno Van Herpen Radja Millenio Kosasih Jeroen Kool Julien Slagboom

Between 80,000 and 140,000 fatalities occur yearly because of snakebite envenoming, which causes it to be classified as a Neglected Tropical Disease (NTD), stressing the importance of snake venom toxin characterisation to aid the development of next-generation antivenoms. Snake venom is made up of a mixture of bioactive proteins that are also known as venom toxins. These venom toxins, being complex and structurally diverse, present a significant challenge for analytical separation and functional characterisation. Traditional methods, such as reversed-phase liquid chromatography (RPLC), often involve denaturing conditions that can alter the native state of these proteins, potentially reducing their bioactivity and limiting subsequent bioassays. Therefore, this study developed and optimised an analytical methodology using non-denaturing Hydrophobic Interaction Chromatography (HIC) for venom separation in conjunction with post-column nanofractionation to perform post-column toxin identification using high-throughput venomics with parallel coagulation bioassays. The results demonstrate that the combination of HIC and nanofractionation allows separation and qualitative profiling of venom toxins with detectable post-column activity, although several toxins co-elute. The post-column bioassays demonstrated detectable coagulation-modulating activity, and high-throughput venomics provided rapid toxin identification capabilities and qualitative information on candidate venom proteins. This approach offers a robust platform for the detailed study of venom toxins, facilitating the discovery of novel bioactive compounds and enhancing our understanding of venom biology.

Toxins doi: 10.3390/toxins18100424

Authors: Yue Lin Yiseul Choi Xiaoqing Liu Hee-Dae Jeon Hyun Jin Park

Regulatory approval may occur before or after a research topic becomes prominent in the scientific literature. We examined this temporal relationship using botulinum neurotoxin (BoNT) as a longitudinal case study. We analyzed 2065 anatomy-related BoNT publications indexed in Web of Science and Scopus from 1983 to 2024. The literature was divided into 16 periods based on U.S. FDA approval dates for onabotulinumtoxinA, and indication-related keyword rankings and thematic clusters were compared with the corresponding approval dates. Among the 10 indication groups examined, 7 showed prominent indication-related keywords before FDA approval, 2 showed signals around the time of approval, and 1 showed a post-approval signal. We describe these patterns as “proactive exploration” when scientific attention became prominent before approval and “reactive catalysis” when it became more prominent around or after approval. Differences in the observed lag may be related to target anatomical complexity and market orientation, but this remains conceptual. Overall, the findings show that time-resolved scientometrics can retrospectively identify scientific patterns associated with regulatory milestones. The approach is hypothesis-generating and requires prospective validation before it can be used for regulatory forecasting.

Toxins doi: 10.3390/toxins18100423

Authors: Kathleen D. Cusick Jackie Palau Malcolm N. McFarland Edith A. Widder

Pyrodinium bahamense is a saxitoxin-producing bioluminescent dinoflagellate that forms annual recurring blooms in the Indian River Lagoon (IRL), FL. Little information exists as to the extent of bloom toxicity. This study examined toxin dynamics in P. bahamense IRL field populations from two sites in the northern IRL (NIRL). Additionally, genomic diversity and transcription were assessed using quantitative molecular assays targeting key functional genes in P. bahamense ecology: sxtA4 (toxin biosynthesis), rbcL (photosynthesis), and lcf (bioluminescence). A positive correlation was found between sxtA4 transcript copy number and toxicity, mirroring the trend recorded in previous seasons and indicating that sxtA4 transcripts can serve as a molecular proxy for bloom toxicity. Lab experiments with 32 strains isolated from the NIRL showed a large variation in copy number for both sxtA4 (42–318 copies cell−1, dependent on strain) and rbcL (1385 to 4911 copies cell−1). An lcf qPCR assay developed here showed substantial variation among isolates as well. These results are indicative of the underlying diversity in field populations, as genome copy number variation is a major driver of genome evolution and diversity. A negative correlation was found between cell abundance and toxin quota in field populations. Growth and toxin quota were also found to have a negative correlation in the 32 isolates. Based on these collective lab and field results, a model is presented in which P. bahamense populations utilize both their toxicity and bioluminescence in bloom development and persistence.

Toxins doi: 10.3390/toxins18100422

Authors: Suguru Yamamoto Takahiro Tanaka Nobutaka Kitamura Miho Kawachi Akio Yokoseki Minako Wakasugi Osamu Onodera Naoto Endo Masakazu Wada Kenji Sato Kaori Kikuchi Ayano Konagai Shin Goto Ichiei Narita

Cognitive dysfunction is increasingly recognized as a significant complication of chronic kidney disease (CKD), but the clinical relevance of protein-bound uremic toxins (PBUTs) to cognition remains insufficiently characterized. This cross-sectional analysis included 1217 community-dwelling individuals. Cognitive function was assessed using the Mini-Mental State Examination (MMSE), with scores < 28 indicating possible mild cognitive impairment (MCI). Serum concentrations of five PBUTs—indoxyl sulfate, p-cresyl sulfate, phenyl sulfate, indoleacetic acid, and hippuric acid—were measured, and a composite PBUT score was derived using principal component analysis. Multivariable logistic regression was used to examine the associations of MCI with the PBUT score and with each PBUT. Serum PBUT concentrations increased with advancing CKD stage. The mean MMSE score was 26.7 ± 3.4, and scores progressively declined with advancing CKD stage. A higher PBUT score was independently associated with MCI (coefficient, 0.290 ± 0.077; odds ratio, 1.337; 95% confidence interval, 1.151–1.554; p < 0.001). In conclusion, a greater PBUT burden was independently associated with cognitive impairment. These findings identify PBUT burden as a potential correlate of cognitive impairment in CKD. Prospective studies are needed to clarify its temporal relationship with cognitive decline and to assess whether it provides predictive value beyond established measures of kidney function.

Toxins doi: 10.3390/toxins18100421

Authors: Ibraimo Teleha Chabite Jamal Bakari Kussaga Frida Nyamete Quenton Kritzinger Shivan Maharaj Rashid Suleiman

Aflatoxin contamination in maize remains a major food safety and public health concern in Nampula Province, Mozambique. Maize samples obtained from farmers, traders, and milling units across the districts of Malema, Mossuril, Mecuburi, Monapo, and Nampula in 2024 and 2025 were analysed using UPLC-ESI-MS/MS. In 2024, aflatoxins were detected in samples from Mecuburi (43%), Mossuril (17%), Monapo (17%), and traders (13%), with some exceeding the regulatory limit of 15 µg/kg. AFB1 was the predominant aflatoxin, followed by AFB2. At the farmer level, Mecuburi recorded the highest contamination, with AFB1 and AFB2 detected in 43% of samples and AFG1 and AFG2 in 29%, while Mossuril had 20% contamination with AFB1 and AFG1. The maximum concentration was 4221.33 µg/kg of AFB1 in Mecuburi, whereas the minimum detected concentration was 2.82 µg/kg of AFB2. Aflatoxins were not detected above the limit of detection in milling samples or in samples collected in 2025. Despite the relatively low prevalence, the occurrence of extremely high aflatoxin concentrations highlights persistent contamination risks and the need for integrated mitigation strategies throughout the maize supply chain.

Toxins doi: 10.3390/toxins18100420

Authors: Guoping Li Lingli Meng Siyuan Feng Pengjuan Cui Gemei Liang Kongming Wu Qiuju Qin Hongqiang Feng

Mythimna (=Leucania) loreyi (Lepidoptera: Noctuidae) is an important pest of summer maize (Zea mays) in China’s Huang–Huai–Hai region. The approved transformation events DBN9936 (Cry1Ab) and Ruifeng 125 (Cry1Ab/Cry2Aj) were primarily developed to target Ostrinia furnacalis, and their efficacy against other lepidoptera pests, such as M. loreyi, remains largely unknown. Using age-stage two-sex life tables under controlled conditions, we quantified lethal and sublethal effects on these transformation events on first- to third-instar M. loreyi larvae fed V6-V8 leaves, and further evaluated how host plant movement scenarios affected larval development. Within a 7-day exposure period, both transformation events exerted significant lethal effects on M. loreyi. Corrected mortalities for first- to third-instar larvae were 93.43%, 17.33%, and 8.61% for DBN9936, and 44.33%, 8.53%, and 3.34% for Ruifeng 125, respectively. Individuals that survived on Bt maize exhibited significant sublethal effects, including prolonged larval development (ΔT = 8.12~9.93 d), reduced pupation rates (decreased by 69.41~100.00%), lower pupal mass (reduced by 31.14~33.64%), and decreased fecundity (by 177.86~262.96 eggs/female). Life table analysis revealed that both transformation events suppressed M. loreyi population growth in an instar-dependent manner. First-instar larvae failed to survive on either Bt transformation event. For second-instar larvae, DBN9936 remained completely lethal, while Ruifeng 125 allowed minimal survival (R0 = 7.51, relative fitness = 0.03). For third-instar larvae, partial survival occurred on both Bt events (DBN9936: R0 = 21.68, relative fitness = 0.09; Ruifeng 125: R0 = 32.34, relative fitness = 0.14), with rm values (0.06–0.07), lower than those on conventional controls (0.14–0.16). Host plant movement scenarios also affected M. loreyi survival and mass gain in an age-dependent manner. Brief early exposure to Bt maize did not reduce survival, but significantly suppressed body mass, whereas continuous Bt exposure caused drastic mortality in young larvae (2.08–8.33% survival). Older larvae tolerated Bt maize without survival loss, yet still exhibited significantly reduced body mass (p < 0.05). These findings demonstrate that DBN9936 and Ruifeng 125 exert substantial significant lethal and sublethal effects on M. loreyi, but the ability of older larvae to survive on Bt maize, combined with potential larval movement between Bt and non-Bt plants may facilitate resistance evolution.

Toxins doi: 10.3390/toxins18100419

Authors: Martina Ferrillo Claudio Curci Dario Calafiore Lorenzo Lippi Yasemin Yumuşakhuylu Michele Vecchio Vincenzo Quinzi Alfredo De Rosa Marco Invernizzi Alessandro de Sire

Temporomandibular disorders (TMDs) are heterogeneous musculoskeletal conditions frequently characterized by pain, impaired mandibular function, and reduced quality of life. Botulinum neurotoxin type A (BoNT-A) has emerged as a potential therapeutic option, particularly for persistent myogenous TMDs associated with masticatory muscle hyperactivity. This narrative review critically examines the mechanisms of action, current clinical evidence, therapeutic applications, and safety profile of BoNT-A in TMD management. Available evidence suggests that BoNT-A can reliably reduce masticatory muscle activity and may provide pain relief in selected patients with refractory myogenous TMDs; however, its superiority over placebo or established conservative treatments remains inconsistent. Considerable heterogeneity in patient selection, injected muscles, dosing regimens, injection techniques, and outcome assessment currently prevents the definition of standardized treatment protocols. Although short-term adverse events are generally mild and transient, repeated injections may induce muscle atrophy, reduced bite force, and potentially mandibular skeletal changes, whose long-term clinical relevance remains uncertain. Therefore, BoNT-A should not be considered a first-line treatment but rather a carefully selected adjunct within a multimodal rehabilitation strategy after appropriate conservative management has failed. Future phenotype-stratified comparative trials with standardized protocols and long-term structural safety outcomes are warranted.

Toxins doi: 10.3390/toxins18100418

Authors: Roko Gladović João G. Brás Jeroen C. Vollenbroek Karin G. F. Gerritsen

Human serum albumin (HSA) is the most abundant plasma protein and plays essential roles in regulating oncotic pressure, transporting endogenous and exogenous compounds, and maintaining redox balance. Because of its biochemical properties, HSA is widely used in clinical medicine, biotechnology, and pharmaceutical development. However, albumin is not a static molecule. It undergoes numerous post-translational modifications (PTMs), including oxidation, glycation, carbonylation, carbamylation, and aggregation, which can significantly alter its structure, ligand-binding properties, antioxidant capacity, and pharmacokinetics. Furthermore, commercial albumin products exhibit substantial heterogeneity arising from differences in source material, manufacturing processes, formulation conditions, and storage practices. This review summarizes the current understanding of PTMs affecting human serum albumin, with particular emphasis on their molecular origins and functional implications. We discuss how physiological, pathological, and manufacturing-induced modifications influence albumin activity and contribute to variability among commercial albumin products. In addition, we examine established and emerging applications of PTM-modified albumin in diagnostics, critical care, extracorporeal detoxification, drug delivery, nanomedicine, and regenerative medicine. Finally, we highlight current challenges related to product heterogeneity, quality control, regulatory assessment, and discuss future opportunities in albumin-based technologies. A deeper understanding of albumin PTMs is essential for improving critical care and developing next-generation albumin-based biomedical products.

Toxins doi: 10.3390/toxins18100417

Authors: Gianpaolo Ronconi Rossella Calciano Alberto Cutaia Rosa Cioffi Eslam Abdelkhalek Farag Anna Sanfilippo Martina Ferrillo Alessandro de Sire Paola Emilia Ferrara

Temporomandibular disorders (TMDs) represent a heterogeneous group of conditions, including myogenous and arthrogenous, with a significant impact on quality of life. Botulinum toxin type A (BoNT-A) has been proposed as a therapeutic option, particularly for patients with refractory myogenous TMD and bruxism. This systematic review aimed to evaluate the clinical effectiveness of BoNT-A in the management of TMDs overall and bruxism, with particular focus on myogenous TMDs compared to placebo and non-surgical treatments. RCTs published between 2020 and 2025 were identified through PubMed, Scopus, and Google Scholar. Overall, 15 RCTs involving 797 participants were included. The available evidence suggests potential benefits of BoNT-A, particularly in patients with localized myofascial pain, myogenous TMD, or marked masticatory muscle hyperactivity. Interpretation of the findings is limited by substantial heterogeneity in treatment protocols, toxin dosage, diagnostic criteria, outcome measures, and follow-up duration. Current evidence does not support the routine use of BoNT-A as a first-line treatment for TMDs overall. Instead, BoNT-A may represent an adjunctive or second-line option for selected patients with myogenous TMD, localized myofascial pain, or marked masticatory muscle hyperactivity who do not adequately respond to conventional noninvasive treatment. Larger and methodologically rigorous RCTs are needed to establish appropriate indications, duration of treatment effects, and long-term safety across different TMD subtypes.

Toxins doi: 10.3390/toxins18100416

Authors: Marta Mira Jørs Geoffrey Masuyer René Jørgensen

Diphthamide-specific mono-ADP-ribosylating toxins form a structurally conserved but phylogenetically diverse family of AB-type bacterial toxins, with Pseudomonas exotoxin A (PE) serving as a prototypical member. Here, we expand the known diversity of PE-like toxins (PLTs) through comprehensive genome mining and phylogenetic, structural and sequence analyses of PE, Chx, and their Aeromonas and Vibrio cholerae homologues. Aeromonas toxins separate into two distinct groups: a PE-like cluster (AEI) and a cholix-like cluster (AEII), while a fourth cholix subgroup (ChxIV) is identified within V. cholerae, expanding the previously described ChxI–III classification. Although all newly identified toxins retain the core catalytic architecture and hallmark residues essential for ADP-ribosylation of eEF2, they differ in key functional motifs, including receptor-binding residues, furin cleavage loops, and C-terminal ER-retrieval signals, suggesting variation in host-cell uptake routes and intracellular trafficking. Signal peptides were present in all toxins but varied modestly between clusters, consistent with normal species-level Sec-pathway variation rather than divergent secretion mechanisms. Together, our findings provide a significantly expanded overview of PLT diversity and indicate that Aeromonas-derived toxins comprise at least two evolutionarily and mechanistically distinct groups. These insights offer a framework for future functional studies and highlight PLTs as emerging virulence factors in environmentally and clinically relevant bacteria.

Toxins doi: 10.3390/toxins18100415

Authors: Kalina Zhivkova

Ochratoxin A (OTA), a mycotoxin produced by Aspergillus and Penicillium species, is a widespread feed contaminant with nephrotoxic, hepatotoxic, immunotoxic, reproductive, and teratogenic effects. Despite the particular susceptibility of rabbits to OTA, information on ochratoxicosis in this species remains limited. This review summarizes current evidence on OTA occurrence, toxicokinetics, mechanisms of toxicity, clinicopathological effects, immunotoxicity, and preventive strategies in rabbits. Following gastrointestinal absorption, OTA has a high affinity for plasma proteins and preferentially accumulates in the kidneys and liver. Its toxicity is mediated by the inhibition of protein synthesis, mitochondrial dysfunction, oxidative stress, genotoxicity, apoptosis, and immune dysregulation, ultimately leading to renal and hepatic injury, hematological and biochemical alterations, impaired growth, and reproductive toxicity. OTA-induced immunosuppression may also increase susceptibility to secondary infections, including Pasteurella multocida. Importantly, pathological and biochemical alterations may occur following relatively low-level exposure, while residual OTA levels may persist in tissues after exposure is discontinued. Effective prevention of OTA contamination in feed is therefore crucial for controlling ochratoxicosis. Microbial and enzymatic detoxification, probiotics, and phytogenic additives represent promising approaches for the prevention and mitigation of OTA toxicity; however, further studies in rabbits are needed to establish their efficacy and optimal application and to evaluate the effects of different OTA doses, early biomarkers, and tissue residue kinetics.

Toxins doi: 10.3390/toxins18100414

Authors: Qiaomin Duan Wanjun Zhang Ying Liu Sunlin Luo Wenjun He Wenjun Yang Yiqiang Chen

Aflatoxin B1 (AFB1) toxicity in poultry is often investigated using naturally contaminated ingredients, which can confound its effects with co-occurring mycotoxins and changes in feed quality. Male Arbor Acres broilers received a common basal diet supplemented with purified AFB1 at nominal concentrations of 0, 5, 10, 20, 50, 100, or 200 μg/kg for 42 d. The reported analyses detected no significant treatment differences in growth performance or major organ indices. Hepatic antioxidant enzyme activities and total antioxidant capacity decreased at selected exposures, while serum antioxidant enzymes, cytokines, and hematological indices showed selective responses. Jejunal villus height was lower in the nominal 50–200 μg/kg groups. Renal tubular lesions were most evident at 50–100 μg/kg. Bile-acid profiling in four selected groups showed compartment-specific differences. These findings suggest that selected biological and morphological responses may occur without detectable growth impairment under the conditions tested. Isolated biomarker differences remain exploratory, and the data do not establish a formal ranking of endpoint sensitivity or a dietary safety threshold.

Toxins doi: 10.3390/toxins18100413

Authors: Enrico Pambianco Octavian Augustin Mihalache Chiara Dall’Asta Antonia Garrido Frenich Laura Carbonell-Rozas

The aim of this study was to evaluate the co-occurrence and to assess the dietary health risk of multiple mycotoxins in fruit-based purées intended for infant consumption, combining targeted quantification and dietary exposure assessment with high-resolution suspect screening. Forty samples with varying apple content were randomly collected from Italian (n = 20) and Spanish markets (n = 20). A fit-for-purpose ultra-high-performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS) method was developed and validated for the simultaneous determination of patulin (PAT) and three Alternaria toxins, including alternariol (AOH), alternariol monomethyl ether (AME), and tentoxin (TEN), providing satisfactory sensitivity, accuracy and precision. Targeted analysis revealed frequent co-occurrence of PAT, AOH, and AME, whereas TEN was not detected. Dietary exposure assessment indicated that PAT posed negligible health risks, whereas AOH and AME exceeded their respective Thresholds of Toxicological Concern (TTCs) across all evaluated population groups, indicating Alternaria toxins as the main contributors to dietary risk. High-resolution mass spectrometry (HRMS) suspect screening expanded the mycotoxin profile through the identification of tenuazonic acid, the tentative identification of altenusin and aflatoxin G2, and the annotation of 2,3-dihydroxybenzoic acid, a potential interferent in PAT determination. Overall, the proposed workflow provides a comprehensive strategy for mycotoxin monitoring in infant fruit purées and supports enhanced surveillance of emerging Alternaria toxins.

Toxins doi: 10.3390/toxins18100412

Authors: Marilena Caporale Giulia Marsico Patrizia Scafato Stefano Superchi

An efficient synthesis of phytotoxin (±)-3-deoxyradicinin was developed from commercially available 4-hydroxy-6-methyl-2H-pyran-2-one. The propenyl side chain was introduced through regioselective bromination followed by Horner–Wadsworth–Emmons olefination, while the second pyranone ring was constructed via a Lewis acid-catalyzed reaction with crotonic acid. The synthetic route was designed according to the principles of operational simplicity, with the aim of minimizing the use of hazardous reagents and solvents, as well as demanding purification procedures and cryogenic conditions. These objectives were successfully achieved for most of the synthetic sequence, with the use of CCl4 in the bromination step representing the main remaining limitation. Four of the six steps were successfully performed on a gram scale, demonstrating the practical potential of the route for a possible scale-up. In addition, a practical synthesis of the natural product 5,6-dehydrokawain was established. Overall, 319 mg of (±)-3-deoxyradicinin were obtained in 23% overall yield, providing a practical approach with reduced reliance on hazardous reagents and low-temperature conditions. This synthetic strategy therefore represents a useful platform for further scale-up of 3-deoxyradicinin preparation and for the synthesis of structurally related analogues.

Toxins doi: 10.3390/toxins18100411

Authors: Euikyung Kim

Animal venoms are among nature’s most sophisticated biochemical systems [...]

Toxins doi: 10.3390/toxins18100410

Authors: Marta Spochacz-Santoro Szymon Chowański Karolina Walkowiak-Nowicka Monika Szymczak-Cendlak Zbigniew Adamski

Glycoalkaloids are plant secondary metabolites known to affect multiple physiological processes in insects; however, their effects on carbohydrate and polyol metabolism remain poorly understood. This study investigated the effects of solasonine on hemolymph metabolites and aldose reductase activity in the fat body of Galleria mellonella larvae. Larvae were fed a diet supplemented with five nominal dietary concentrations of solasonine (10−4–1 mg/kg diet) for three days. Hemolymph metabolites were analyzed by RP-HPLC, and aldose reductase activity was determined in fat body lysates. Inositol levels differed significantly among experimental groups (one-way ANOVA, p < 0.0001), with the 10−4 mg/kg diet treatment differing significantly from the vehicle control (p = 0.0015). Aldose reductase activity was significantly lower at 10−2, 10−3, and 10−4 mg/kg diet compared with the untreated control (p = 0.0439, p = 0.027, and p < 0.0001, respectively), while only the 10−4 mg/kg treatment differed significantly from the vehicle control (p = 0.0029). Formic acid also significantly affected several metabolic parameters. Overall, solasonine exposure was associated with altered aldose reductase activity and changes in polyol-related metabolites, although some metabolic effects may be attributable to the vehicle.

Toxins doi: 10.3390/toxins18100409

Authors: Bryan G. Fry

Debate over the origin of squamate venom is frequently distorted by treating molecular availability, toxin recruitment and secretion, functional venom, and delivery-system elaboration as a single evolutionary event. This review separates these transitions and reassesses early venom evolution in anguimorph lizards and snakes using anatomical, molecular, proteomic, functional, ecological, and phylogenetic evidence. The species-level ancestral-state reconstructions of a recent Wüster, Modahl, and Arbuckle manuscript are internally robust to the four coding schemes tested and accommodate uncertain tips and reversible venom loss. Their inference nevertheless remains conditional on a binary definition of observed functional venom, a single hidden precursor category, a homogeneous transition process, and non-caenophidian snake states that were not varied in the relevant sensitivity analyses. While they coded two origins of venom within Anguimorpha, gland anatomy and derived oral products make an early functional origin followed by retention, reduction, loss, and parallel elaboration a strongly supported alternate hypothesis. However, as critical bridge taxa lack direct evidence of natural delivery, realistic dose, or secretion-specific fitness benefit, resolution of the history remains unresolved. For Heloderma, field studies of H. suspectum and H. horridum, together with painful, prolonged defensive bites, support a defensive role, although its primacy (to the exclusion of an additional predatory function) as advocated by them is sensitive to observational bias. Neurotoxic, cardiovascular, and coagulotoxic activities make a predatory contribution supported, leaving the original selective driver unresolved. Within Serpentes, their placement of venom at the base of the Caenophidia is challenged by distributed maxillary, mandibular, and rictal glands and derived oral products in non-caenophidian taxa, making an origin before Caenophidia a viable alternative hypothesis. Derived mucous systems in boas and pythons and specialised Scolecophidia cannot alone polarise the ancestral snake state. Current evidence is therefore insufficient to distinguish one shared functional origin from two functionalisations of a homologous ancestral oral secretory system. No alternative comparative reconstruction is performed; the review shows only that earlier origins remain viable and categorical absence coding is insufficiently secure for several poorly investigated taxa. Resolution requires matched gland cell biology, orthology- and synteny-resolved genomics, quantitative secretion proteomics, realistic functional assays, secretion-specific ecological tests, broader Iguania and non-caenophidian snake sampling, and ancestral-state models that represent graded evidence, both false-negative and false-positive classification, reduction, loss, and phylogenetic uncertainty.

Toxins doi: 10.3390/toxins18100408

Authors: Maxine Bennek Martina Kerscher

Botulinum neurotoxin type A (BoNTA) is widely used for the treatment of dynamic facial wrinkles, where precise control of toxin diffusion is essential to achieve predictable aesthetic outcomes while preserving natural facial expression. However, the relative influence of injection pattern, dose, and dilution on diffusion characteristics remains incompletely understood. In this exploratory case series, treatment-naïve subjects undergoing aesthetic treatment of horizontal forehead lines with LetibotulinumtoxinA (Leti-BoNTA) were assigned to one of four injection paradigms according to their wrinkle pattern: a conventional five-point injection pattern, the previously published LEBO injection pattern, dose variation (2–5 U per injection point), or dilution variation (reconstitution volumes 0.675–2.5 mL). Diffusion was assessed using the Minor iodine–starch test, and anhidrotic areas were quantified in cm2. Increasing dose was associated with progressively larger anhidrotic areas in this exploratory case series. Greater reconstitution volumes were associated with broader toxin distribution, whereas lower dilution volumes produced more confined diffusion areas. The conventional five-point injection pattern resulted in relatively extensive diffusion, whereas the LEBO protocol demonstrated a more localized and consistent distribution profile. Despite differences in diffusion characteristics, natural facial dynamics were preserved across all treatment paradigms. These findings suggest that injection pattern, dose, and dilution collectively influence Leti-BoNTA diffusion and should be considered when optimizing individualized treatment strategies.

Toxins doi: 10.3390/toxins18100407

Authors: Yuan Guo Yuxin An Jiaojiao Xu Zengxuan Cai Meijia Xu Xiaomin Xu

Bongkrekic acid (BA) is a metabolite that is lethal to humans and is produced by Burkholderia gladioli pathovar cocovenenans (BGC) containing the bon gene cluster. An analytical method for BA and its stereoisomer, iso-bongkrekic acid (IBA), was established for deeply fermented rice products, such as red yeast rice and its related products, by liquid chromatography–tandem mass spectrometry (LC-MS/MS). BA and IBA in samples were extracted with 80% methanol/water containing 1% ammonia by ultrasonic assistance, cleaned by solid-phase extraction (SPE) with an anion-exchange cartridge, separated with an XBridgeTM BEH C18 chromatographic column (150 mm × 3.0 mm, 2.5 μm), measured by MS/MS, and quantified using an isotope-labeled internal standard. The influencing factors such as extraction solvents, ultrasonic extraction time and loading volume during SPE clean-up were optimized to improve the recoveries of BA and IBA. Also, the extraction efficiency between the simulated quality control and directly spiked samples, the matrix effects with the use of different chromatographic columns, the impact of co-eluting interferences under different mobile phase compositions and the conversion of BA to IBA are discussed. The linear range of BA and IBA was 0.1–50 ng/mL (equivalent to 1–500 μg/kg in the sample), with the correlation coefficient exceeding 0.999. The limits of detection and quantification were 0.3–0.5 μg/kg and 1.0–1.5 μg/kg, respectively. The average recoveries of the analytes were 88.4–107% with relative standard deviations (RSDs, n = 6) between 2.9% and 11.8% in red yeast rice and its related products under three spiking levels. This optimized method was applied to study the contamination of BA and IBA in commercially available samples. This study can provide technical support for the supervision and risk assessment of BA and IBA in red yeast rice and its related products.

Toxins doi: 10.3390/toxins18100406

Authors: Rozenn Souillard Sandra Rouxel Virginie Dorenlor Grégoire Kuntz Félix Mahé Typhaine Poezevara Florent Eono Amandine Avouac Stéphane Kerphérique Léa Jambou Eloïse Cailton Mathys Gouet Sébastien Solanas Loïc Balaine Adeline Huneau-Salaün Marianne Chemaly Sophie Le Bouquin Alain Hartmann Caroline Le Maréchal

Animal botulism mainly affects poultry and cattle, with risks of cross-contamination. This study aimed to evaluate Clostridium botulinum contamination in manure and soil following manure spreading on 13 clinically affected farms (10 cattle and three poultry) and 39 clinically unaffected farms (20 cattle and five poultry farms without any previous botulism outbreak, six poultry farms with a history of botulism and eight mixed poultry/cattle farms with an ongoing bovine botulism outbreak). Twelve manure samples, two swabs on a manure pile, 12 soil samples, and two boot swabs from fields were collected before manure spreading. Twelve soil samples and two boot swabs from fields were obtained after manure spreading. C. botulinum types C, D, C/D, and D/C were detected using real-time PCR after an enrichment step. On clinically unaffected poultry and cattle farms with no history of botulism, C. botulinum was detected in 1.4% and 1.8% of samples, respectively. Detection rates reached 8% on poultry farms with a history of botulism and 42.8% on poultry farms with ongoing bovine outbreaks. On clinically affected poultry and cattle farms, 34% and 18.9% of samples were positive, respectively. This study highlights the presence of C. botulinum in manure and soil and emphasizes the importance of biosecurity measures to prevent cross-contamination and disease recurrence.

Toxins doi: 10.3390/toxins18090405

Authors: Tulio Custódio Reis Ana Clara Lunardi Yagi Eloise T. Filardi Marcela Romanazzi Michele Ferreira da Silva Mela Rui S. Ferreira Júnior Felipe A. Cerni Márcia A. S. Graminha Manuela B. Pucca

Leishmaniasis and Chagas disease are neglected tropical diseases that continue to pose significant public health challenges, highlighting the need for new therapeutic strategies. Brazilian snake venoms represent a valuable and underexplored source of bioactive molecules with diverse pharmacological activities and promising therapeutic potential. In this context, the present study aimed to investigate the in vitro antiprotozoal potential of venoms from Bothrops and Crotalus species, including Bothrops jararacussu, Bothrops pauloensis, Bothrops alternatus, Bothrops leucurus, Bothrops atrox, Bothrops jararaca, Crotalus durissus terrificus, and Crotalus durissus ruruima, metalloprotease (MTL)- and crotoxin (CTX)-enriched, against Leishmania spp. and Trypanosoma cruzi. Crude venom protein profiles were analyzed by SDS-PAGE, and cytotoxicity was assessed in differentiated C2C12 myotubes and murine peritoneal macrophages. Antiprotozoal activity was evaluated against extracellular parasite forms, including promastigotes of Leishmania amazonensis and Leishmania infantum and epimastigotes of T. cruzi. Venoms exhibiting the highest selectivity indices (SIs), calculated using murine macrophage cytotoxicity data, were further tested against intracellular amastigotes of L. amazonensis. Among the tested samples, B. jararacussu venom showed activity against promastigotes of both L. amazonensis (IC50 = 2.9 µg mL−1) and L. infantum (IC50 = 2.8 µg mL−1), as well as in epimastigotes of T. cruzi (IC50 = 15.5 µg mL−1). The MTLand CTX pools from C. d. ruruima venom exhibited the highest activity against promastigotes of L. amazonensis, with IC50 values of 1.6 and 3.8 µg mL−1, respectively. Against intracellular amastigotes, MTL and CTX showed IC50 values of 4.1 and 2.5 µg mL−1, respectively, with selectivity indices (SIs) of 73.1 and 120, respectively. Overall, these findings provide preliminary evidence for the antiparasitic potential of snake venom-derived molecules, supporting further characterization and in vivo evaluation.

Toxins doi: 10.3390/toxins18090404

Authors: Kazuya Yoshida

Botulinum neurotoxin (BoNT) therapy has developed from a treatment primarily used for neurological disorders into a versatile therapeutic modality applied across a broad range of medical and dental conditions [...]

Toxins doi: 10.3390/toxins18090403

Authors: Andrea Felice Armenti Francesco Armenti

Pharmacodynamic models of botulinum toxin type A map a mean concentration onto a smooth dose–response curve, yet the outcome is binary and arises from a finite population of near-threshold motor units. We analyse a discrete model in which N units are silenced once local SNAP-25 cleavage exceeds a unit-specific threshold, and a functional block of neuromuscular transmission follows once the silenced fraction exceeds a collective threshold. Whether that collective threshold is a fraction of the population or a fixed count changes the location of the dose–response curve and not its steepness: under a fraction, potency is invariant to unit number at fixed dose–concentration gain, and the transition contracts in absolute dose where fixed-count architectures require it to widen. The width contracts as N−1/2, an exponent that is prior and not claimed here. One consequence needs no measurement: a quantal terminal response caps the silenced fraction below unity, so any fixed count fails above a target size it determines, whereas the toxin block targets differ by orders of magnitude. A second reinterprets existing data, flat cohort dose–response curves being what a near-step individual response predicts once convolved with between-subject dispersion. Outcome is accordingly less repeatable in the smallest targets. No parameter is fitted.

Toxins doi: 10.3390/toxins18090402

Authors: Bart Eeckhaut Steven Truijen Parham Haghshenas Antriana Roussou Petros Taflampas Wim Saeys

Background: Botulinum toxin type A (BoNT-A) is globally recognized as a standard treatment for post-stroke spasticity (PSS), although its secondary mechanisms remain underinvestigated. The objective of this study was to investigate the neurophysiological mechanisms underlying BoNT-A treatment in PSS and explore their implications for mechanism-based rehabilitation. Methods: A systematic review and meta-analysis (PROSPERO registration number ID: CRD420261352230) of 39 studies involving 760 patients was conducted on 18 August 2026, following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. PubMed, Web of Science, Scopus, and Embase were systematically searched using the PICO framework. Where possible, quantitative data were pooled according to predefined neurophysiological constructs, including passive stretch-evoked activity, voluntary activation of the injected muscle, compound muscle action potential (CMAP), reciprocal motor control, normalized Hmax/Mmax, and specific spinal inhibitory mechanisms. Results: Quantitative synthesis demonstrated construct-specific neurophysiological changes following BoNT-A treatment. Passive stretch-evoked activity showed the largest pooled effect (Hedges’ g = 0.80, 95% CI 0.35–1.24), followed by CMAP amplitude (g = 0.69, 95% CI 0.23–1.14) and reciprocal motor control (g = 0.52, 95% CI 0.23–0.81). Effects on voluntary activation of the injected muscle (g = 0.31, 95% CI −0.38 to 1.00) and normalized Hmax/Mmax (g = 0.22, 95% CI −0.74 to 1.17) were smaller and more heterogeneous. Reciprocal and recurrent inhibition were each represented by single-study estimates, while supraspinal outcomes were synthesized narratively due to methodological heterogeneity. Conclusions: The findings support a predominantly peripheral mechanism of BoNT-A while suggesting possible secondary changes within spinal and supraspinal motor control pathways. However, the certainty of evidence was low to very low. These neurophysiological effects provide a rationale for individualized, mechanism-based rehabilitation during the post-injection therapeutic window, although higher-quality evidence is required to confirm this framework.

Toxins doi: 10.3390/toxins18090401

Authors: Corinne M. Gibble Melissa A. Miller Raphael M. Kudela Angelina Reed Kevin Keel Rebecca S. Duerr

In 2017, an unusually large number of live stranded and dead marine birds were found in California, coincident with a large bloom of the diatom Pseudo-nitzschia that produced the toxin domoic acid (DA). The most abundant bird species affected by this large die-off and stranding event were Pacific Loons (Gavia pacifica) and Red-throated Loons (Gavia stellata). International Bird Rescue (IBR; San Pedro, CA) received 117 live loons with suspected DA toxicosis. Many birds that survived the first few days in rehabilitation quickly re-attained suitable health metrics; however, 68 loons (58.1%) died or were considered unreleasable due to ongoing abnormal behavior or injuries that necessitated humane euthanasia. In this descriptive study, both live stranded euthanized birds and dead stranded birds were examined to characterize and better understand the effects of DA toxicosis in the avian brain and body. Utilizing a suite of diagnostics, our analysis revealed novel pathological patterns that divided these birds into three groups: those suspected of having acute, subacute, and chronic DA toxicosis. Domoic Acid was found in all groups, and the severity of the microscopic lesions appeared to correspond with the amount of time each bird survived post-stranding. Our investigation is the first known account identifying the pathological progression from acute to chronic DA toxicosis in birds.

Toxins doi: 10.3390/toxins18090400

Authors: Riccardo Marvulli Serena Caforio Montesardo Chiara Moccia Marisa Megna Maurizio Ranieri

Upper limb spasticity is a common complication of stroke that negatively affects motor function, range of motion, and quality of life. Although abobotulinumtoxinA is widely used for the treatment of focal post-stroke spasticity, evidence regarding its early onset of action remains limited. This prospective observational pilot study investigated the early clinical, functional, and neurophysiological effects of abobotulinumtoxinA in 20 adults with post-stroke upper limb spasticity. Patients received ultrasound-guided injections into the biceps brachii and flexor digitorum superficialis muscles and were evaluated at baseline, 24 h, 5 days, 7 days, 14 days, and 30 days after treatment. Outcome measures included the Modified Ashworth Scale (MAS), passive range of motion (ROM), caregiver-reported hand function assessed using a visual analog scale (VAS), and compound muscle action potential (cMAP) recordings. No significant changes were observed at 24 h. A significant reduction in spasticity was detected at the day-5 assessment, accompanied by significant improvements in ROM and functional outcomes. Neurophysiological assessment demonstrated significant reductions in cMAP amplitudes at the day-5 assessment, indicating early neuromuscular transmission blockade. Clinical improvements were also first detected at day-5 assessment and were maintained at subsequent evaluations through day 30. These findings indicate that clinically meaningful effects of abobotulinumtoxinA were detectable by the first scheduled assessment at day 5 after injection. Given the absence of intermediate assessments between 24 h and day 5, the exact timing of treatment onset cannot be determined. A multimodal assessment approach may help characterize early treatment-related changes and provide useful information for future studies investigating the timing of adjunctive rehabilitation interventions.

Toxins doi: 10.3390/toxins18090399

Authors: Audra Janovskienė Justina Stučinskaitė-Maračinskienė Žygimantas Petronis Jan Pavel Rokicki Dainius Razukevičius Loreta Pilipaitytė

Background: Botulinum toxin type A (BoNT-A) has been proposed as an adjunct to orthognathic surgery because its temporary neuromuscular effects may reduce muscular loading during postoperative healing and potentially influence fixation stability, skeletal relapse, and postoperative recovery. This systematic review aimed to evaluate the available clinical evidence regarding the adjunctive use of BoNT-A in orthognathic surgery. Methods: A systematic literature search was conducted in PubMed, the Cochrane Library, and Google Scholar in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Clinical studies directly comparing orthognathic surgery with and without adjunctive BoNT-A administration and reporting outcomes related to mechanical stability, skeletal stability, or postoperative recovery were included. Risk of bias was assessed using the Cochrane Risk of Bias 2 (RoB 2) tool for randomized controlled trials and ROBINS-I for the non-randomized study. Results: Three studies met the eligibility criteria, comprising two randomized controlled trials and one retrospective comparative study. Considerable heterogeneity was observed in surgical procedures, targeted muscles, BoNT-A doses, timing of administration, and evaluated outcomes. Individual studies reported associations between BoNT-A administration and a lower incidence of fixation plate fracture, reduced anteroposterior skeletal relapse at the Pogonion, or lower preanalgesic pain scores and postoperative opioid consumption. However, these findings were derived from different studies evaluating distinct interventions and outcomes and were not consistently demonstrated across comparable outcome measures. Conclusions: BoNT-A may represent a promising adjunct to orthognathic surgery, with potential benefits for fixation-related mechanical stability, selected parameters of skeletal relapse, and postoperative pain management. However, the limited number of studies, methodological limitations, and substantial clinical heterogeneity preclude definitive conclusions. Further well-designed randomized controlled trials with standardized injection protocols and longer follow-up are required before routine clinical use can be recommended.

Toxins doi: 10.3390/toxins18090398

Authors: Aideen Steed Heather Williams Jessie Alfonso Denise Hancock Stephen A. Ashford

Botulinum toxin type A (BoNT-A) is an established treatment for spasticity. Goal Attainment Scaling (GAS), alongside standardized measures such as Arm Activity Measure (ArmA) and Leg Activity Measure (LegA), enables evaluation of patient-centered outcomes. This real-world cohort study evaluated goal attainment and standardized measure functional changes following BoNT-A for arm and/or leg spasticity, evaluated using GAS as well as passive function subscales of ArmA (ArmA-PF) and LegA (LegA-PF), and active function subscales (ArmA-AF; LegA-AF). Inclusion in the analysis was based on documented baseline and post-intervention GAS scores following BoNT-A therapy (n = 390). Across 855 treatment cycles, 1258 goals were set and categorized, of which 150 (11.9%) were related to active function and 705 (56.0%) to passive function. The mean GAS T-score improved from 36.2 to 49.4 (change 13.1, p < 0.001), with significant gains observed for both passive (change 13.7, p < 0.001) and active goals (change 12.6, p < 0.001). Passive function improved significantly on ArmA-PF (change 2.5) and LegA-PF (change 2.1), while significant changes in ArmA-AF and LegA-AF were not seen. In conclusion, goal-directed spasticity management incorporating BoNT-A resulted in improvements in goal attainment, which corresponded to a clinically meaningful change in passive function measured by ArmA-PF and LegA-PF, but not for active function. Passive function was the most common treatment priority. Findings provide real-world evidence to support community-based spasticity services and the use of structured goal setting, ArmA and LegA application, in routine practice.

Toxins doi: 10.3390/toxins18090397

Authors: Noah M. Souza Alexander O. Hale Richard S. Beard Juliette K. Tinker

AB5 toxins are important virulence factors produced by Gram-negative bacteria and have been identified in many non-typhoidal Salmonella (NTS) serovars; however, their effects on host cell signaling and epithelial barrier function remain poorly understood. This study characterized the molecular and functional effects of purified ArtAB toxin and its binding subunit, ArtB, from multidrug-resistant Salmonella Typhimurium definitive type 104 (DT104). Equimolar concentrations of ArtAB (10 µg/mL) and ArtB (7.25 µg/mL) were evaluated in Chinese hamster ovary (CHO-K1) cells using cytotoxicity assays, quantitative proteomics, and mRNA sequencing, while barrier function was assessed in C2BBe1 human intestinal epithelial cells using electric cell-substrate impedance sensing (ECIS). In CHO-K1 cells, both proteins exhibited minimal cell death. ArtB elicited pronounced proteomic changes associated with endoplasmic reticulum (ER) homeostasis, vesicular trafficking, and lysosomal processing, but limited transcriptional responses. ArtAB instead altered host transcriptional programs associated with cell growth, inflammatory signaling, and cytoskeletal organization. Unlike ArtB, ArtAB significantly reduced barrier function in C2BBe1 cells. Together, these findings show that ArtAB remodels host cell signaling without overt cytotoxicity in CHO-K1 cells and produces a distinct functional effect in C2BBe1 cells. Although obtained in different cellular models, these results provide insight into the potential role of ArtAB in NTS pathogenesis.

Toxins doi: 10.3390/toxins18090396

Authors: Guangyao Ying Maofa Zeng Liang Hong Zhaokui Li Jun Li Huili Xia

Mycotoxin contamination represents one of the most pressing food safety challenges worldwide, with millions of tons of agricultural commodities affected annually. Conventional detection methods, while accurate, are labor-intensive, destructive, and unsuitable for large-scale screening. Hyperspectral imaging (HSI) has emerged as a transformative non-destructive analytical technique capable of simultaneously capturing spatial and spectral information across hundreds of contiguous wavelengths. This review critically evaluates the current state of HSI technology for mycotoxin detection in food products, covering the physical principles underlying spectral–mycotoxin interactions, systematic applications across major mycotoxin classes (aflatoxins, deoxynivalenol, ochratoxin A, fumonisins, and zearalenone), and the integration of machine learning and deep learning algorithms for spectral data analysis. The review reveals that while Vis-NIR (400–1000 nm) and SWIR (1000–2500 nm) HSI systems have achieved classification accuracies exceeding 90% for several mycotoxin–matrix combinations, fundamental challenges persist in model transferability, direct quantification at regulatory thresholds, and scalability for industrial deployment. Recent advances in transformer architectures, transfer learning, interpretable deep learning, and portable multispectral systems demonstrate encouraging progress toward practical implementation. This review concludes by identifying critical research gaps and proposing strategic directions for translating HSI-based mycotoxin detection from laboratory proof-of-concept to routine industrial application.

Toxins doi: 10.3390/toxins18090395

Authors: Galina Nugumanova

Cyanobacterial harmful algal blooms (cyanoHABs) typically involve the release of a wide spectrum of toxic metabolites produced by different cyanobacteria, which can cause adverse health effects in animals and humans. At the same time, many cyanobacteria are capable of producing more than one toxin simultaneously. As a result, the multiclass toxins released into aquatic environments can exert complex and combined effects on various ecosystem components. This review aims to provide an overview of the occurrence and effects of different cyanotoxins in aquatic systems and current analytical methods of their simultaneous detection. The main classes of cyanotoxins have been considered in this review. The occurrences, properties, and environmental and health impacts of these toxins are described. The main methods for cyanobacterial toxin analysis are covered. A comprehensive review and analysis of liquid chromatography-tandem mass spectrometry (LC-MS/MS) detection and sample preparation approaches used to identify various classes of cyanotoxins is provided, together with a practical decision tree for analytical workflow selection according to toxin class and sample matrix, highlighting key trends and outlining directions for further research in this area.

Toxins doi: 10.3390/toxins18090394

Authors: Yong-Hua Wu Zheng-Xu Zhong Yan Li Jing-An Wang Zheng-Wen Ou Hou-Jin Li Wen-Jian Lan

Vespa basalis Smith venom (VBsV) causes painful and potentially severe systemic reactions, yet the relationships among its composition, bioactivity, and cytotoxicity remain poorly understood. Proteomic and metabolomic profiling was combined with ultrafiltration and gel filtration to characterize VBsV and obtain >3 kDa, operationally defined 700–3000 Da, and <700 Da fractions. The effect on LPS-induced NO production was evaluated by nitric oxide (NO) release in LPS-stimulated RAW264.7 cells, and cytotoxicity was assessed in tumor and THLE-2 liver cells. Five venom allergen proteins were identified, including two phospholipase A1 isoforms, hyaluronidase A, antigen 5, and venom dipeptidyl peptidase IV. Targeted profiling also detected several neurotransmitters and energy metabolites, including 5-hydroxytryptamine, L-glutamate, histamine, γ-aminobutyric acid, inosine, and adenosine. The 700–3000 Da fraction exhibited the strongest NO-suppressive and tumor-cell-viability effects among the tested fractions. The 700–3000 Da fraction suppressed NO release (IC50, approximately 23 μg/mL) and reduced tumor-cell viability, while minimally affecting THLE-2 cells at 10–80 μg/mL; by contrast, the >3 kDa fraction showed greater THLE-2 cytotoxicity. These findings identify the 700–3000 Da fraction as a priority for direct compositional characterization and isolation of the responsible molecules.

Toxins doi: 10.3390/toxins18090393

Authors: Bazbek Davletov Alexandr Ignatchenko Aisha Zhantleuova Rashid A. Giniatullin

Alpha-latrotoxin (α-LTX) is the principal vertebrate-specific neurotoxin in widow spider (Latrodectus) venom and a powerful presynaptic secretagogue. Although its receptors, pore-forming activity and stimulation of neurotransmitter release have been studied extensively in central and motor neurons, its relationship to the severe, persistent pain of latrodectism remains poorly understood. This focused review re-examines α-LTX from a nociceptive perspective. The available evidence supports a model in which α-LTX binds adhesion G protein-coupled receptor L1 (ADGRL1; latrophilin-1) and neurexin-1α on susceptible sensory neurons; inserts a large cation-permeable pore; and promotes membrane depolarisation, calcium entry and release of pain-associated neuropeptides. The transcript-level gene expression of α-LTX receptor genes in dorsal root ganglion neurons and evidence of toxin-evoked neuropeptide release provide a molecular basis for direct nociceptor activation, although functional validation in defined nociceptor subclasses remains necessary. A comparison with the nociceptive ion channels transient receptor potential vanilloid 1 (TRPV1) and transient receptor potential ankyrin 1 (TRPA1) highlights their convergence on calcium-dependent sensory excitation, while distinguishing α-LTX from toxins that modulate endogenous channels. We propose that the pain of latrodectism is a composite state in which direct nociceptor activation complements muscle spasm and tissue-derived signalling, positioning α-LTX as a distinctive probe of pain pathways.

Toxins doi: 10.3390/toxins18090392

Authors: Padmini Vaka Laharika Kappari Gokul V. Selvaraj Ramesh K. Selvaraj Todd J. Applegate Revathi Shanmugasundaram

Mycotoxins are widespread contaminants in animal feeds and continue to pose serious threats to animal welfare, production efficiency, food security, and sustainable economic growth worldwide. Chickens are highly susceptible to multiple mycotoxins, such as aflatoxins (AFLs), deoxynivalenol (DON), fumonisins (FBs), zearalenone (ZEA), ochratoxin A (OTA), and T-2 toxin. Exposure to these toxins can damage intestinal integrity, compromise immune functions, impair nutrient absorption, and ultimately reduce production efficiency even at subclinical concentrations. Conventional mycotoxin detection methods, such as enzyme-linked immunosorbent assays (ELISA) and chromatographic techniques, provide accurate quantification but remain expensive, labor-intensive, time-consuming, and inefficient for large-scale screening, particularly when masked mycotoxins are present. The frequent co-occurrence of multiple mycotoxins further complicates risk assessment and effective management. Emerging analytical technologies, including hyperspectral imaging, biosensors, Internet of Things (IoT)-based platforms, and machine-learning algorithms, offer promising advancements for rapid detection and predictive risk forecasting. This review summarizes the toxicological impacts of major mycotoxins on poultry, outlines critical challenges in detection and prevention, and evaluates current artificial intelligence (AI) and machine learning (ML) approaches for mycotoxin identification, prediction, and management. A systematic literature search conducted across PubMed, ScienceDirect and Google Scholar identified 176 unique studies published between 2010 and 2026. Key knowledge gaps include limited availability of high-quality datasets and source code, inconsistent model interpretability, and poor reproducibility across production environments. By integrating conventional toxicology with data-driven approaches, this review highlights how predictive modeling can strengthen mycotoxin surveillance and support proactive mitigation strategies in modern poultry production systems.

Toxins doi: 10.3390/toxins18090391

Authors: Farshad Mazda Shirazi Srikar R. Adhikari Vance G. Nielsen Geoffrey Thomas Smelski

Neurotoxic envenomation by rattlesnakes has been of great concern throughout the Americas and in Arizona. In addition to clinical weakness, patients sometimes display myokymia, a disordered fasciculation of skeletal muscle, which is mediated by Mojave toxin. As neurotoxic venoms have been associated with increased release of creatine kinase (CK) activity and concordant increases in length of stay (LOS) in the hospital, it was retrospectively determined if point-of-care ultrasound detected myokymia at the bite site was associated with increased CK activity, LOS, and healthcare cost compared to rattlesnake envenomations not associated with myokymia. The Arizona Poison and Drug Information Center patient database was queried for patients with point-of-care ultrasound exam results at or near the bite site from 2017 to 2021. Using point-of-care ultrasound to detect subclinical myokymia, thirty-seven patients that met inclusion criteria were determined to have myokymia (n = 16) and/or not display it (n = 21). Patients with myokymia had twice the CK activities, double the LOS, and two-fold the healthcare costs of patients without myokymia detected. These findings serve as a rationale for future investigations to link ultrasound detected myokymia with patient outcomes.

Toxins doi: 10.3390/toxins18090390

Authors: Oana Mîrza Denisia Pașca Lara Manyes Anamaria Cozma-Petruț Roxana Banc Simona Codruța Hegheș Lorena Filip Béla Kiss

Mycotoxins are secondary fungal metabolites that have continuously attracted research interest, both for their potential negative effects on animal and human health and for the economic losses they entail. Studies assessing exposure to mycotoxins have become increasingly widespread. The aim of this study was to provide a comprehensive description of mycotoxins and to evaluate the evolution of research on monitoring mycotoxin exposure using human urine samples, concerning analytical techniques and occurrence. Studies published in English over the last 25 years from four databases (Web of Science, PubMed, Science Direct, Scopus) were analyzed. Evaluating 73 original articles, a constant interest in the analysis of ochratoxin A in human urine samples could be observed (maximum level 148 ng/mL), a fact also justified by its demonstrated toxicity. The most commonly used analytical technique to evaluate mycotoxin presence in human urine was liquid chromatography coupled with high-performance detectors. The limitations of the review include the large period covered and the lack of mycotoxin concentrations expressed in ng biomarker/mL urine in some articles. Finally, an overview of how mycotoxin levels in human urine can be applied to mycotoxin exposure risk assessment was included.

Toxins doi: 10.3390/toxins18090389

Authors: Ren-Chieh Wu Kuang-Ting Yeh

Acute kidney injury (AKI) is a severe complication of Eastern Russell’s viper (Daboia siamensis) envenoming, but species-specific mechanisms, treatment effects, and long-term outcomes remain uncertain. This review addressed four questions: the human renal evidence specific to D. siamensis; venom and host pathways contributing to AKI; conclusions supported for antivenom timing and thrombotic microangiopathy (TMA); and quantifiable long-term kidney risk. PubMed and OpenAlex were searched from inception through 3 August 2026. Species-authenticated human evidence was separated from mixed-species or D. russelii studies and from venom-analytical and experimental evidence. Referral cohorts show AKI and dialysis burdens, but heterogeneous ascertainment, definitions, and care preclude pooled incidence. Proteomic and experimental studies support interacting coagulopathic, endothelial, hemodynamic, pigment, inflammatory, and direct renal pathways; no human study quantifies their relative contribution, and no D. siamensis study localizes specific venom antigens within renal tissue. Observational data support prompt use of indicated, regionally appropriate antivenom but do not validate a universal two-hour renal-protection threshold. TMA and long-term outcome evidence remain largely indirect. Serial renal and coagulation surveillance is warranted irrespective of whether injury is direct or indirect. Prospective, species-authenticated studies linking venom kinetics and toxin profiles to standardized renal phenotypes and follow-up are required.

Toxins doi: 10.3390/toxins18090388

Authors: Kuo-Hao Lo Kuo-Cheng Lu Yen-Hung Liu Yi-Chou Hou

Streptococcal toxic shock syndrome (STSS) is a fulminant manifestation of invasive group A Streptococcus infection characterized by superantigen-driven hyperinflammation, refractory shock, and rapidly progressive multiorgan dysfunction. Despite antimicrobial therapy, source control, intravenous immunoglobulin, and intensive organ support, some patients deteriorate rapidly, prompting interest in extracorporeal immunomodulation. This narrative review examines the biological rationale and clinical experience of hemoadsorption in STSS, supported by an illustrative case and comparison with published reports. Our patient developed vasopressor-dependent shock, acute kidney injury, thrombocytopenia, hepatic and myocardial injury, hyperlactatemia, and metabolic acidosis within 48 h. CytoSorb was integrated with continuous venovenous hemofiltration during ongoing deterioration, followed by hemodynamic and organ recovery; however, concurrent therapies preclude attribution of benefit to hemoadsorption. Published cases show a similar pattern, with extracorporeal escalation initiated during refractory shock and accumulating organ dysfunction rather than at a predefined biomarker threshold. Although experimental and observational evidence supports the biological activity of hemoadsorption, clinical evidence remains insufficient to establish a survival benefit. Current evidence does not support routine hemoadsorption for STSS. A rapidly progressive, multidimensional hyperinflammatory trajectory may instead provide a framework for future studies of patient selection and treatment timing.

Toxins doi: 10.3390/toxins18090387

Authors: Suguru Yamamoto

The number of patients with chronic kidney disease (CKD) has increased worldwide [...]

Toxins doi: 10.3390/toxins18090386

Authors: Ivonne Salazar-Rodríguez Iván López Patricia Glorio-Paulet Isabel Cristina Molina-Botero Arturo Gerardo Valdivia-Flores Carlos Gómez-Bravo

The strategy of adding mycotoxin binders (MTBs) to feed lactating cows to reduce the absorption of aflatoxins (AFs) in the gastrointestinal tract and reduce milk contamination by hydroxylated AF compounds, especially AFM1, to below the maximum contaminant level (MCL) has become widespread. However, the indiscriminate use of MTBs is very common. The objective of the study was to determine, under field conditions, the efficacy of an MTB in reducing AFM1 contamination in cow’s milk. Feed and raw milk were evaluated on-farm using lateral flow immunochromatography devices. In a 2 × 2 Latin square design, in two separate test periods, two diets were administered consecutively to each of twelve Holstein cows (multiparous, 220 ± 20 lactation days): 1: Basal diet naturally contaminated (120 AFB1 µg/d); 2: Basal diet plus an MTB (calcium carbonate and yeast fragments; 150 g/d). MTB intake effectively reduced the mean concentration, daily excretion, and percentage transfer rate of AFM1 in milk (0.724 ± 0.045 to 0.479 ± 0.055 µg/kg, p < 0.01; 15.6 ± 1.7 to 9.7 ± 1.5 µg/d, p < 0.05; and 13.0 ± 1.4 to 8.1 ± 1.3%, p < 0.05, respectively). These results highlight the need to evaluate, under field conditions, the effectiveness of MTBs as part of a mycotoxin contamination control program on dairy farms.

Toxins doi: 10.3390/toxins18090385

Authors: Francesca Bosco Chiara Mollea Gabriele Baldissone Micaela Demichela Davide Fissore

In the present work, the correlation between the respirometric activity and the aflatoxin synthesis of the hazelnut microbiota has been investigated. Microcosms containing a monolayer of hazelnuts were maintained at controlled conditions in terms of temperature and humidity. To monitor microbial growth, CO2 was evaluated along with the specific O2 uptake rate, RO2, measured by a respirometric sensor, usually applied in the evaluation of soil microbial activity. The respiratory quotient (RQ) was calculated and a linear relationship between RQ and produced aflatoxins was obtained. Depending on incubation time, four main clusters of RQ values were identified: for higher incubation time (15–40 days) RQ was lower than 2 and an aflatoxin amount of 0.31–0.35 μg/kg was observed. CO2, O2, and aflatoxin data have been used to develop an autoregressive model and excellent agreement between measurements and estimations was obtained. In most cases, an error lower than 1% on the estimated value of the mycotoxin amount revealed the accuracy of the evaluation of the RQ. The present work represents the first attempt to develop an effective method, fast and low-cost, for the in-line monitoring of mycotoxin contamination.