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Keywords = polymerase chain reaction (PCR)

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12 pages, 585 KiB  
Article
Antimicrobial Resistance Profile and Biofilm Formation of Listeria monocytogenes Isolated from Meat
by Joana Paiva, Vanessa Silva, Patrícia Poeta and Cristina Saraiva
Antibiotics 2025, 14(5), 454; https://doi.org/10.3390/antibiotics14050454 - 30 Apr 2025
Viewed by 224
Abstract
Introduction: Listeria monocytogenes is the causative agent of listeriosis, a serious infectious disease with one of the highest case fatality rates among foodborne diseases affecting humans. Objectives: This study investigated the prevalence, antimicrobial resistance pattern and biofilm production capacity of L. monocytogenes isolated [...] Read more.
Introduction: Listeria monocytogenes is the causative agent of listeriosis, a serious infectious disease with one of the highest case fatality rates among foodborne diseases affecting humans. Objectives: This study investigated the prevalence, antimicrobial resistance pattern and biofilm production capacity of L. monocytogenes isolated in meats. Materials: A total of 75 samples were analyzed, including fresh meats and meat preparations, in Northern Portugal. Methods: The strains were identified using morphological and molecular methods. Antimicrobial resistance was determined using the Kirby–Bauer disk diffusion method, against a panel of 12 antibiotics and the presence of the respective antimicrobial resistance genes was investigated by polymerase chain reaction (PCR). The ability to form biofilms was evaluated by the microtiter biofilm assay. Results: The overall prevalence of L. monocytogenes among screened samples was 17.33%. The isolates were resistant to trimethoprim-sulfamethoxazole (85.71%), ciprofloxacin (38.10%), meropenem (33.33%), tetracycline and erythromycin (28.57%), rifampicin (23.81%), and kanamycin (14.29%). Six isolates (28.57%) exhibited a multidrug-resistance profile. All strains showed positive result for the virulence gene specific to listeriolysin O (hlyA). In the genotypic resistance analysis of the strains, the genes identified were tetK (23.81%), aadA, tetL, blaOXA-48 (14.29%), ermC, and msr(A/B) (4.76%). All isolates had the ability to form biofilms, with no significant differences in biofilm biomass production at 24 h and 48 h. Some of these strains showed a high capacity for biofilm production. Conclusions: These findings raise public health concerns due to resistance to first-line antibiotics and the biofilm-forming capacity of these isolates, which pose risks to the food industry. Enhanced monitoring and surveillance are essential to guide public health strategies in order to mitigate the threat posed by L. monocytogenes in food. Full article
(This article belongs to the Special Issue The Antimicrobial Resistance in the Food Chain)
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19 pages, 10572 KiB  
Article
Development and Application of a TaqMan-Based qPCR Assay for Detecting ENTV-2 in Goats
by Pengfei Li, Haike Yin, Xiaoan Cao, Xi Lan, Jinyan Wu, Jijun He, Ligang Yuan and Youjun Shang
Genes 2025, 16(5), 529; https://doi.org/10.3390/genes16050529 - 29 Apr 2025
Viewed by 124
Abstract
Background: In recent years, enzootic nasal tumor virus 2 (ENTV-2) has become prevalent in China, resulting in substantial economic losses for the goat industry. In order to enrich the availability of detection methods for ENTV-2, this study developed an expedited and accurate reverse-transcription [...] Read more.
Background: In recent years, enzootic nasal tumor virus 2 (ENTV-2) has become prevalent in China, resulting in substantial economic losses for the goat industry. In order to enrich the availability of detection methods for ENTV-2, this study developed an expedited and accurate reverse-transcription quantitative real-time polymerase chain reaction (RT-qPCR) assay to facilitate the detection and quantification of ENTV-2. Methods: Specifically, a pair of primers and a TaqMan probe targeting conserved regions of the pro gene were designed to allow the specific amplification and detection of viral RNA in clinical samples. Moreover, modifying the method for use in a quantitative real-time PCR (qPCR) assay enables the detection of proviral DNA in tumor specimens. Results: Both methods exhibited a detection limit for the ENTV-2 standard plasmid at 100 copies/µL. The detection methods we established exhibited high specificity and sensitivity to ENTV-2, without cross-reactivity with other pathogens causing respiratory diseases or endogenous retroviruses (EBRVs). We performed an ENTV-2 analysis of clinical samples in goats via RT-qPCR using nasal swab samples (n = 558) collected from three geographically distinct flocks in Lingyou County, Baoji City, Shaanxi Province, China, and 58 positive samples were detected for a positivity rate of 10.4%. After euthanasia, the autopsy report showed nasal cavity masses. Histopathological analysis demonstrated an epithelial neoplasm, in compliance with the features of enzootic nasal adenocarcinoma (ENA). Three full-length genomes were sequenced to assess genomic sequence conservation and variation. Multiple-sequence alignment demonstrated the existence of sequence variations among strains. Phylogenetic analysis of the nucleotide sequences revealed that the ENTV-2 SX1~3 isolates were phylogenetically related to the Chinese ENTV-2 isolates, especially the JY strain. Furthermore, recombination analysis suggested that both ENTV-2 SX1 and ENTV-2 SX2 might be recombinant variants. Conclusions: In conclusion, both methods are highly specific for the pro gene of ENTV-2, and the development of this assay has been deemed crucial to the early identification and subsequent control of this viral infection. Our results provide valuable information for further research on the genetic variation and evolution of ENTV-2 in China. Full article
(This article belongs to the Section Animal Genetics and Genomics)
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15 pages, 4253 KiB  
Article
Whole-Genome DNA Methylation Analysis in Age-Related Hearing Loss
by Marie Valerie Roche, Denise Yan, Yan Guo, Naser Hamad, Juan I. Young, Susan H. Blanton, Feng Gong and Xue Zhong Liu
Genes 2025, 16(5), 526; https://doi.org/10.3390/genes16050526 - 29 Apr 2025
Viewed by 177
Abstract
Background: Presbycusis, also known as age-related hearing loss (ARHL), is the most frequent sensory disability affecting elderly adults worldwide. ARHL is characterized by bilateral, progressive, sensorineural hearing loss that is more pronounced at a high frequency. Conventional factors associated with ARHL include diabetes, [...] Read more.
Background: Presbycusis, also known as age-related hearing loss (ARHL), is the most frequent sensory disability affecting elderly adults worldwide. ARHL is characterized by bilateral, progressive, sensorineural hearing loss that is more pronounced at a high frequency. Conventional factors associated with ARHL include diabetes, hypertension, and a family history of hearing loss. The severity of hearing impairment varies between individuals. The defined causative molecular pathogenesis for ARHL is unknown, thus the identification of underlying pathogenic mechanisms involved in ARHL is imperative for the development of effective therapeutic approaches. Epigenetics is the study of phenotypic changes caused by the modification of gene expression rather than the alteration of a DNA sequence. While it is hypothesized that ARHL could result from undiscovered epigenetic susceptibility, there is a shortage of information on the role that epigenetic modification plays in ARHL. Here we present an investigation on the involvement of DNA methylation in ARHL. Results: Clinical, audiometric and DNA testing, and high-throughput methylation pattern screening were undertaken for ARHL patients and matched control subjects. Our results demonstrate a strong correlation between patients’ hearing measurements and methylation at CpG sites cg1140494 (ESPN) and cg27224823 (TNFRSF25). We identified 136 differentially methylated CpGs that were shared between a high and low audiometric frequency in the patient’s cohort. CpG cites in hearing loss candidate genes, KCNQ1, TMEM43, GSTM1, TCF25, and GSR, were found to be highly methylated in presbycusis patients as compared to the controls. A methylation polymerase chain reaction (PCR) assay was used to confirm methylation levels at a specific gene locus in ARHL patients and controls. Conclusions: Altered DNA methylation and its impact on gene expression has been implicated in many biological processes. By interrogating the methylation status across the genome of both hearing loss patients and those with normal hearing, our study can help to establish an association between the audiometric patterns and methylation status in ARHL, yielding new avenues for the identification of potential candidate genes for hearing loss. Full article
(This article belongs to the Section Human Genomics and Genetic Diseases)
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15 pages, 1090 KiB  
Article
Exploring Life Detection on Mars: Understanding Challenges in DNA Amplification in Martian Regolith Analogue After Fe Ion Irradiation
by Alessia Cassaro, Claudia Pacelli and Silvano Onofri
Life 2025, 15(5), 716; https://doi.org/10.3390/life15050716 - 29 Apr 2025
Viewed by 248
Abstract
The search for life beyond Earth currently hinges on the detection of biosignatures that are indicative of current or past life, with terrestrial life being the sole known example. Deoxyribonucleic acid (DNA), which acts as the long-term storage of genetic information in all [...] Read more.
The search for life beyond Earth currently hinges on the detection of biosignatures that are indicative of current or past life, with terrestrial life being the sole known example. Deoxyribonucleic acid (DNA), which acts as the long-term storage of genetic information in all known organisms, is considered a biosignature of life. Techniques like the Polymerase Chain Reaction (PCR) are particularly useful as they allow for the amplification of DNA fragments, allowing the detection of even trace amounts of genetic material. This study aimed to detect DNA extracted from colonies of an Antarctic black fungus both when (i) alone and (ii) mixed with a Sulfatic Mars Regolith Simulant (S-MRS), after exposure to increasing doses of Fe ions (up to 1 kGy). PCR-based amplification methods were used for detection. The findings of this study revealed no DNA amplification in samples mixed with Sulfatic Mars Regolith Simulant, providing important insights into the potential application of these techniques for in situ DNA detection during future space exploration missions or for their application on the Mars sample return program; it also gives input in the planetary protection discussions. Full article
(This article belongs to the Section Astrobiology)
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20 pages, 3689 KiB  
Article
Molecular Characterization, Antibiotic Resistance, and Biofilm Formation of Escherichia coli Isolated from Commercial Broilers from Four Chinese Provinces
by Saqib Nawaz, Muhammad Shoaib, Cuiqin Huang, Wei Jiang, Yinli Bao, Xiuyi Wu, Lianhua Nie, Wenyan Fan, Zhihao Wang, Zhaoguo Chen, Huifang Yin and Xiangan Han
Microorganisms 2025, 13(5), 1017; https://doi.org/10.3390/microorganisms13051017 - 28 Apr 2025
Viewed by 240
Abstract
Escherichia coli (E. coli) represents a significant etiological agent of colibacillosis in poultry, resulting in considerable economic losses for the global poultry sector. The present study aimed to determine molecular characterization, antibiotic resistance, and biofilm formation of E. coli strains isolated [...] Read more.
Escherichia coli (E. coli) represents a significant etiological agent of colibacillosis in poultry, resulting in considerable economic losses for the global poultry sector. The present study aimed to determine molecular characterization, antibiotic resistance, and biofilm formation of E. coli strains isolated from diseased broilers from four provinces of China. A total of 200 tissue samples were collected from the intestine, liver, crop, heart, and spleen and processed for microbiological examination. Molecular detection of E. coli strains, virulence genes, and serotypes was performed using polymerase chain reaction (PCR). Antibiotic susceptibility testing and biofilm formation were assessed using disk diffusion and 96-well microtiter plate assays. The study retrieved 68% (136/200) of E. coli strains from collected samples. Most of the E. coli strains were resistant to enrofloxacin (56%), followed by cefepime (54%), amoxicillin/clavulanate (52%), streptomycin (50%), ampicillin (48%), clindamycin (47%), kanamycin (41%), polymyxin B (37%), tetracycline (35%), sulfamethoxazole/trimethoprim (33%), ceftazidime (31%), meropenem (4.7%), and florfenicol (2.9%). Similarly, the E. coli strains tested positive for at least one virulence gene and specific serotypes. Among these, O145 was the most prevalent serotype, identified in 22 isolates (16.2%), followed by O8 (12.5%), O102 (11.8%), and O9 (11.0%). The tsh gene (10.2%) was the most prevalent virulence gene. This study found that 47.1% of E. coli strains were biofilm-producing, with 62.5% exhibiting weak biofilm production, 29.7% mild biofilm production, and 7.8% strong biofilm production. Similarly, 24.2% of the E. coli strains were avian pathogenic E. coli strains due to the presence of five or more virulence genes, specifically tsh, ompA, fimC, iss, fyuA, and astA, in a single strain by multiplex PCR. The present study recommends continuous surveillance and effective control measures to reduce the burden of avian pathogenic E. coli-related infections in poultry. Full article
(This article belongs to the Special Issue Poultry Pathogens and Poultry Diseases, 2nd Edition)
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12 pages, 3228 KiB  
Article
Common Diagnoses from Surgical Biopsies and Investigation of Leporipoxvirus in Pet Rabbits (Oryctolagus cuniculi) in Taiwan
by Ya-Mei Chen, Yang-Chun Wu, Ching-Liang Kuo, Wei-Hao Lin and Kuo-Ping Shen
Animals 2025, 15(9), 1234; https://doi.org/10.3390/ani15091234 - 27 Apr 2025
Viewed by 122
Abstract
This study investigated common diagnoses from surgical biopsies of domestic rabbits (Oryctolagus cuniculi) in Taiwan and examined the role of leporipoxvirus in tumor pathogenesis. Biopsy specimens from 70 rabbits collected between 2014 and 2023 were retrospectively analyzed, yielding 85 diagnoses. Polymerase [...] Read more.
This study investigated common diagnoses from surgical biopsies of domestic rabbits (Oryctolagus cuniculi) in Taiwan and examined the role of leporipoxvirus in tumor pathogenesis. Biopsy specimens from 70 rabbits collected between 2014 and 2023 were retrospectively analyzed, yielding 85 diagnoses. Polymerase chain reaction (PCR) was performed to detect leporipoxvirus in formalin-fixed, paraffin-embedded tissues diagnosed with fibroma, fibrosarcoma, or myxosarcoma. The most commonly affected systems were the integumentary (n = 41) and reproductive (n = 36) systems. Common integumentary tumors included fibrosarcomas (n = 12), trichoblastomas (n = 8), mammary gland tumors (n = 5), and fibromas (n = 4). In the reproductive system, the most common lesions were uterine adenocarcinomas (n = 16), uterine endometrial cystic hyperplasia and hypertrophy (n = 5), and uterine adenomyosis (n = 4). The 15 cases of fibroma, fibrosarcoma, and myxosarcoma were tested for leporipoxvirus using PCR. No viral sequences were detected in these tumors. This study identified the common diagnoses from rabbit biopsy specimens and found no leporipoxvirus infection in samples of fibrosarcoma and fibroma. This is the first study on tumors in pet rabbit biopsies and the first in Taiwan to investigate leporipoxvirus infection, providing valuable insights for future diagnosis and management. Full article
(This article belongs to the Section Veterinary Clinical Studies)
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14 pages, 5530 KiB  
Article
Intranasal Application of Foxp3 Introduced with Poly(d,l-lactic-co-glycolic acid) (PLGA) Nanoparticles (Foxp3 NPs) Attenuates Allergic Inflammation in a Mouse Model of Allergic Rhinitis
by Seung Cheol Han, Sunhee Yeon, Hyejeen Kim and Sookyoung Park
Pharmaceutics 2025, 17(5), 575; https://doi.org/10.3390/pharmaceutics17050575 - 27 Apr 2025
Viewed by 190
Abstract
Background: Allergic rhinitis (AR) is a common disease that requires more convenient, safe, and effective therapy. This study aimed to investigate the therapeutic effect of Forkhead box protein3 (Foxp3) introduced with poly(d,l-lactic-co-glycolic acid) (PLGA) nanoparticles (Foxp3 NPs) in [...] Read more.
Background: Allergic rhinitis (AR) is a common disease that requires more convenient, safe, and effective therapy. This study aimed to investigate the therapeutic effect of Forkhead box protein3 (Foxp3) introduced with poly(d,l-lactic-co-glycolic acid) (PLGA) nanoparticles (Foxp3 NPs) in an AR mouse model. Methods: A murine model of allergic rhinitis was established using BALB/c mice through initial sensitization by intraperitoneal administration of ovalbumin (OVA), followed by repeated intranasal OVA challenges. Foxp3 plasmid-loaded PLGA nanoparticles were subsequently administered via either the intranasal or intraperitoneal route to evaluate therapeutic efficacy. Episodes of sneezing and nose rubbing were counted. The serum total IgE, OVA-specific IgE, and cytokine levels in nasal lavage fluid (NALF) were determined by ELISA (Enzyme-Linked ImmunoSorbent Assay). Nasal mucosa from each group were analyzed using protein, reverse transcriptase–polymerase chain reaction (RT-PCR), and histological analyses. Result: Rubbing and sneezing symptoms improved in the Foxp3 NPs intranasal administration group. Foxp3 NPs intranasal administration markedly ameliorated OVA-induced nasal allergic inflammation. The total IgE and OVA-specific IgE serum level and IL-4, IL-13 expression levels of NALF were significantly decreased in the treated Foxp3 NPs group. The histopathological results of nasal mucosa were also normal, with no cellular infiltration and no inflammation in the Foxp3 NPs group. Conclusions: These results suggest that Foxp3 NPs alleviate nasal allergic inflammation and may have therapeutic value in the treatment of AR. Full article
(This article belongs to the Section Nanomedicine and Nanotechnology)
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17 pages, 5333 KiB  
Article
Comprehensive Identification of HD-Zip Family Genes in Coix lacryma-jobi L. and Their Potential Roles in Response to Abiotic Stress
by Yongle Wang, Hongjuan Wang, Xianyong Lu, Chun Yu, Benli Jiang, Jiabao Zhu and Yujiao Wang
Plants 2025, 14(9), 1318; https://doi.org/10.3390/plants14091318 - 26 Apr 2025
Viewed by 223
Abstract
HD-Zip (homeodomain-leucine zipper) transcription factors play a crucial role in plant growth, development, and stress response; however, the HD-Zip gene family of Coix lacryma-jobi L. has not been identified. In this study, a total of 40 HD-Zip gene family members were identified in [...] Read more.
HD-Zip (homeodomain-leucine zipper) transcription factors play a crucial role in plant growth, development, and stress response; however, the HD-Zip gene family of Coix lacryma-jobi L. has not been identified. In this study, a total of 40 HD-Zip gene family members were identified in the genome of Coix. According to phylogenetic analysis, the Coix HD-Zip gene was divided into four subfamilies (I–IV), of which the HD-Zip I subfamily can be further divided into five branches. Moreover, HD-Zip members of the same subfamily usually share similar gene structures and conserved motifs. The transcription factor binding site enrichment analysis showed that there are many motifs for binding with transcription factors such as ERF (Ethylene responsive factor), MYB (v-myb avian myeloblastosis viral oncogene homolog), and ARF (Auxin Response Factor) in the promoter region of the ClHDZ genes. The results of qPCR (Quantitative Polymerase Chain Reaction) and expression profile analysis showed that ClHD-Zip I genes showed different levels of expression under different stress treatments. Among them, ClHDZ4 was located in the nucleus, and its expression pattern was significantly upregulated under salt, drought, and high-temperature stress. In addition, ectopic expression of ClHDZ4 enhanced the growth of yeast strains under drought, salt, or high-temperature treatment. In summary, these results laid a foundation for further research on the resistance function of the Coix HD-Zip gene. Full article
(This article belongs to the Special Issue Physiological and Genetic Responses of Crops to Abiotic Stress)
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17 pages, 3293 KiB  
Article
Epicatechin Decreases UCP2 Gene Expression in MDA-MB-231 Breast Cancer Cells by the Presence of a Regulatory Element in the Promoter
by Fernando Pereyra-Vergara, Ivonne María Olivares-Corichi, Juan Pedro Luna-Arias, David Méndez-Luna and José Rubén García-Sánchez
Int. J. Mol. Sci. 2025, 26(9), 4102; https://doi.org/10.3390/ijms26094102 - 25 Apr 2025
Viewed by 234
Abstract
Uncoupling protein 2 (UCP2) plays an important role in normal cells because it mitigates the cytotoxic effect of reactive oxygen species (ROS). However, its overexpression in cancer cells is related to drug resistance and increased cell proliferation due to a decrease in ROS [...] Read more.
Uncoupling protein 2 (UCP2) plays an important role in normal cells because it mitigates the cytotoxic effect of reactive oxygen species (ROS). However, its overexpression in cancer cells is related to drug resistance and increased cell proliferation due to a decrease in ROS production. In this context, molecules that regulate or block UCP2 have potential as anticancer agents. (-)-Epicatechin, a flavonoid that inhibits cell proliferation, increases ROS, and induces apoptosis in cancerous cells, was evaluated for its effects on UCP2 gene expression. For this purpose, the real-time quantitative polymerase chain reaction (qRT–PCR) and Western blotting were performed in MDA-MB-231 and MCF-10A cells to determine the effects of (-)-epicatechin on UCP2 expression. Furthermore, the impact of (-)-epicatechin on cell viability was also determined. To analyze the transcriptional regulation of the UCP2 gene by (-)-epicatechin, a 5′-region of the human UCP2 gene (−2093/+297) was amplified, sequenced, cloned, and inserted into a reporter plasmid. To analyze the promoter activity and regulatory motif involved in the effects of (-)-epicatechin, several deletions of the UCP2 promoter were generated and transfected into MDA-MB-231 and MCF-10A cells. An electrophoretic mobility shift assay (EMSA) was carried out to detect the interaction between DNA and proteins involved in the effect of (-)-epicatechin. The increased expression of the UCP2 gene in MDA-MB-231 cells was decreased by (-)-epicatechin, and the opposite effect was observed in MCF-10A cells. The promoter region of the human UCP2 gene (−2093/+297) showed activity, which was decreased by (-)-epicatechin. A sequence of 117 bp located at position −109 b to +8 b has a fragment of 90 bp that is related to the (-)-epicatechin effect. Bioinformatics analysis and EMSA of this sequence revealed the presence of a regulatory site for a protein with zinc fingers. The presence of a response element to (-)-epicatechin in the human UCP2 promoter revealed that the inhibition of this gene in MDA-MB-231 breast cancer cells occurred at the transcriptional level. In this study, we propose the mechanism of action of (-)-epicatechin that could aid in cancer treatment. Full article
(This article belongs to the Special Issue Molecular Research in Triple-Negative Breast Cancer)
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19 pages, 6335 KiB  
Article
Response of Soil Microbial Diversity to Triple-Cropping System in Paddy Fields in Middle Reaches of Yangtze River
by Haiying Tang, Junlin Zhou, Ning Liu, Yao Huang, Qin Liu, Faizah Amer Altihani and Binjuan Yang
Plants 2025, 14(9), 1292; https://doi.org/10.3390/plants14091292 - 24 Apr 2025
Viewed by 285
Abstract
To explore the characteristics of soil microbial community structure diversity for different planting patterns in paddy fields, and to screen out the planting patterns suitable for the promotion of double-cropping rice areas in the middle reaches of the Yangtze River, five typical planting [...] Read more.
To explore the characteristics of soil microbial community structure diversity for different planting patterns in paddy fields, and to screen out the planting patterns suitable for the promotion of double-cropping rice areas in the middle reaches of the Yangtze River, five typical planting patterns were set up in this study. The five patterns are Chinese milk vetch–early rice–late rice (CRR, CK), Chinese milk vetch–early rice–sweet potato || late soybean (CRI), rapeseed–early rice–late rice (RRR), rapeseed–early rice–sweet potato || late soybean (RRI) and potato–early rice–late rice (PRR). The variation characteristics of soil microbial community structure diversity and the correlation between soil environmental factors and soil microbial community structure diversity under the triple-cropping system in the double-cropping rice area of the middle reaches of the Yangtze River were studied by 16S rRNA high-throughput sequencing and real-time fluorescence quantitative polymerase chain reaction (PCR). The results showed that after two years of experiment, the pH values of each treatment increased, and the rapeseed–early rice–late rice (RRR) model performed better. The soil organic matter and total nitrogen content of the milk vetch–early rice–sweet potato || late soybean (CRI) model was the highest, which increased by 7.89~35.02% and 6.59~26.80% compared with other treatments. The content of soil available phosphorus and available potassium in the potato–early rice–late rice (PRR) model was higher than that in other treatments, which was increased by 29.48% and 126.49% compared with the control. The Chinese milk vetch–early rice–sweet potato || late soybean (CRI) and rapeseed–early rice–sweet potato || late soybean (RRI) models were beneficial to increasing soil nitrate nitrogen and ammonium nitrogen content. Chinese milk vetch–early rice–sweet potato || late soybean (CRI) and rapeseed–early rice–late rice (RRR) patterns were beneficial for improving the microbial diversity index. Proteobacteria, Chloroflexi, and Actinobacteria are the top three dominant phyla in terms of the relative abundance of soil bacteria, and the top three dominant fungi are Ascomycota, Basidiomycota, and Mucor. The Chinese milk vetch–early rice–sweet potato || late soybean (CRI) and rapeseed–early rice–sweet potato || late soybean (RRI) patterns increased the relative abundance of soil Actinobacteria and Ascomycota. The contents of ammonium nitrogen, total organic carbon, nitrate nitrogen, and available phosphorus were the main environmental factors affecting soil microbial community structure. The findings can provide references for screening out the planting patterns suitable for the promotion of double-cropping rice areas in the middle reaches of the Yangtze River. Full article
(This article belongs to the Section Plant–Soil Interactions)
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13 pages, 239 KiB  
Article
Genetic Variants of the ATIC Gene and Therapeutic Response to Methotrexate in Patients with Rheumatoid Arthritis
by Sergio Gabriel Gallardo-Moya, Laura Gonzalez-Lopez, Betsabe Contreras-Haro, Mario Alberto Mireles-Ramirez, Alejandra Villagomez-Vega, María Cristina Morán-Moguel, Miriam Méndez-Del Villar, María Luisa Vazquez-Villegas, Jorge Ivan Gamez-Nava and Ana Miriam Saldaña-Cruz
Int. J. Mol. Sci. 2025, 26(9), 4013; https://doi.org/10.3390/ijms26094013 - 24 Apr 2025
Viewed by 149
Abstract
Methotrexate (MTX) is the conventional synthetic disease-modifying anti-rheumatic drug (csDMARD) recommended as the first-choice anti-rheumatic drug for rheumatoid arthritis (RA). However, responses to MTX may be influenced by genetic variants. We aim to evaluate the association of the rs2372536, rs4673990, and rs4673993 genetic [...] Read more.
Methotrexate (MTX) is the conventional synthetic disease-modifying anti-rheumatic drug (csDMARD) recommended as the first-choice anti-rheumatic drug for rheumatoid arthritis (RA). However, responses to MTX may be influenced by genetic variants. We aim to evaluate the association of the rs2372536, rs4673990, and rs4673993 genetic variants of the ATIC gene with therapeutic failure of MTX in patients with RA. A case–control study was performed. Disease activity was measured using the disease activity score based on erythrocyte sedimentation rate (DAS28-ESR). RA patients were classified into two groups: (a) responders (DAS28-ESR ≤ 3.2), which is the group of patients who did respond to methotrexate, and (b) non-responders (DAS28-ESR > 3.2), which is the group of patients who did not respond to methotrexate. Serum levels of the 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) enzyme and Interleukin-6 (IL-6) were quantified using an enzyme-linked immunosorbent assay (ELISA). Genotyping of ATIC genetic variants was performed with quantitative polymerase chain reaction (qPCR) using TaqMan probes. A total of 260 patients with RA were included. In total, 142 (54.6%) were non-responders to MTX. IL-6 levels were increased in the non-responder group (p = 0.002), while no statistical differences were observed in the AICAR levels. The variables associated with non-response were higher HAQ-Di, weekly MTX dose, glucocorticoid use, erythrocyte sedimentation rate, and carriers of the polymorphic homozygous variant of rs4673993 (OR = 4.5, 95% CI: 1.04–19.34; p = 0.04). The use of sulfazaline offered protective effects. Our findings indicate that the polymorphism rs4673993 gene variant of the AICAR protein may significantly influence MTX resistance. Therefore, these results support the importance of the pathway generating extracellular adenosine and its effects on promoting the immune regulation for the mechanism of MTX therapy of RA. Full article
(This article belongs to the Special Issue Rheumatoid Arthritis: From Molecular Basis to Therapies)
14 pages, 5249 KiB  
Article
Selection of Bactrocera tau (Walker) Reference Genes for Quantitative Real-Time PCR
by Yutong Zhai, Yonghao Yu, Pengfei Xu, Xianru Zeng, Xiuzhen Long, Dewei Wei, Zhan He and Xuyuan Gao
Insects 2025, 16(5), 445; https://doi.org/10.3390/insects16050445 - 24 Apr 2025
Viewed by 184
Abstract
The selection of appropriate reference genes is critical for standardizing quantitative real-time reverse transcription polymerase chain reaction (RT-qPCR) data, thereby ensuring accurate and reliable results of gene expression analysis. In this study, we identified 10 candidate reference genes (encoding α-tubulin, G6PDH, [...] Read more.
The selection of appropriate reference genes is critical for standardizing quantitative real-time reverse transcription polymerase chain reaction (RT-qPCR) data, thereby ensuring accurate and reliable results of gene expression analysis. In this study, we identified 10 candidate reference genes (encoding α-tubulin, G6PDH, Rab1, RT, RPS13, β-tubulin, DPH1, HSP90, GAPDH, and CP) and evaluated their suitability for use as reference genes in the pest insect, Bactrocera tau. Analysis was conducted using three software-based methods—Delta CT, NormFinder, and BestKeeper—alongside the online tool RefFinder. Expression levels of these genes were analyzed across various B. tau developmental stages and body parts. The overall ranking of reference gene stability scores was as follows: α-tubulin > G6PDH > CP > β-tubulin > RT > HSP90 > GAPDH > DPH1 > RPS13 > Rab1. Ultimately, α-tubulin and G6PDH were identified as the most stable reference genes for B. tau. Full article
(This article belongs to the Section Insect Molecular Biology and Genomics)
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13 pages, 869 KiB  
Article
Detection of SARS-CoV-2 Using the Abbott™ PANBIO™ COVID-19 SELF-TEST Rapid Test in Patients Seen at INER
by Kenny Alonso Cantón Cruz, Martha Angella Durán Barrón, Israel Agustín Morales Lozada, Mario Alberto Mujica Sánchez, Grecia Gabriela Deloya Brito, María del Carmen García Colín, Hansel Hugo Chávez Morales, José Nicolás Aguirre Pineda, Cinthya Karen Cid del Prado Rojas, Stephanie Jara Valdés and Eduardo Becerril Vargas
Biomedicines 2025, 13(5), 1012; https://doi.org/10.3390/biomedicines13051012 - 22 Apr 2025
Viewed by 279
Abstract
The COVID-19 pandemic has highlighted the need for rapid and accurate tests to detect SARS-CoV-2. Objectives: This study evaluates the effectiveness of the Panbio™ COVID-19 Antigen Self-Test rapid test compared to reverse transcriptase polymerase chain reaction (RT-PCR). Methods: A prospective diagnostic testing [...] Read more.
The COVID-19 pandemic has highlighted the need for rapid and accurate tests to detect SARS-CoV-2. Objectives: This study evaluates the effectiveness of the Panbio™ COVID-19 Antigen Self-Test rapid test compared to reverse transcriptase polymerase chain reaction (RT-PCR). Methods: A prospective diagnostic testing study was performed. Patients with respiratory symptoms who provided informed consent were included. Results: We included 205 patients with COVID-19 symptoms who underwent both tests. The mean age was 35.55 ± 12.62 years, and 64% of the participants were female. Sensitivity and specificity were 71.2% (95% CI: 62.5–79.9%) and 100% (95% CI: 96.4–100%), respectively. Conclusions: If a test is positive within the first 72 h after the onset of symptoms, we can be sure that it is a case of COVID-19; however, when the antigen test is negative, confirmation with RT-PCR is necessary. Its ease of use and results with moderate precision make it a valuable tool for early detection. Full article
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8 pages, 2999 KiB  
Communication
Diagnosis of Community-Acquired Pneumonia Due to Influenza or Respiratory Syncytial Virus: Evaluation of RT-PCR Sensitivity in Nasopharyngeal, Saliva, and Sputum Samples
by Julio Ramirez, Stephen Furmanek, Thomas Chandler, Ruth Carrico, Ashley Wilde, Alan Junkins and Anupama Raghuram
Pathogens 2025, 14(5), 400; https://doi.org/10.3390/pathogens14050400 - 22 Apr 2025
Viewed by 249
Abstract
Detection of viral RNA in nasopharyngeal (NP) samples by reverse transcription polymerase chain reaction (RT-PCR) is the standard diagnostic test for influenza or respiratory syncytial virus (RSV) in hospitalized patients with community-acquired pneumonia (CAP). This study compared the sensitivity of RT-PCR using NP [...] Read more.
Detection of viral RNA in nasopharyngeal (NP) samples by reverse transcription polymerase chain reaction (RT-PCR) is the standard diagnostic test for influenza or respiratory syncytial virus (RSV) in hospitalized patients with community-acquired pneumonia (CAP). This study compared the sensitivity of RT-PCR using NP swab, saliva, and sputum samples for the diagnosis of CAP due to influenza or RSV. A total of 60 patients were evaluated, of which 40 (67%) had influenza CAP, 19 (32%) had RSV CAP, and one patient (1%) had both RSV and influenza CAP. RT-PCR on NP swab, saliva, and sputum samples was performed using the Luminex ARIES platform. In patients with influenza CAP, the sensitivity was 34% for NP swabs, 68% for saliva, and 71% for sputum. In patients with RSV CAP, the sensitivity was 60% for NP swabs, 75% for saliva, and 85% for sputum. RT-PCR of nasopharyngeal swab samples was associated with a significant number of false negative results. A negative NP swab RT-PCR test should not be used to rule out CAP due to influenza or RSV. Saliva and sputum samples should be considered when performing a microbiological work-up in patients with suspected influenza or RSV CAP. Full article
(This article belongs to the Section Viral Pathogens)
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25 pages, 3049 KiB  
Article
HCM-Associated MuRF1 Variants Compromise Ubiquitylation and Are Predicted to Alter Protein Structure
by Jitpisute Chunthorng-Orn, Maya Noureddine, Peter W. J. Dawson, Samuel O. Lord, Jimi Ng, Luke Boyton, Katja Gehmlich, Fiyaz Mohammed and Yu-Chiang Lai
Int. J. Mol. Sci. 2025, 26(8), 3921; https://doi.org/10.3390/ijms26083921 - 21 Apr 2025
Viewed by 544
Abstract
MuRF1 [muscle RING (Really Interesting New Gene)-finger protein-1] is an ubiquitin-protein ligase (E3), which encode by TRIM63 (tripartite motif containing 63) gene, playing a crucial role in regulating cardiac muscle size and function through ubiquitylation. Among hypertrophic cardiomyopathy (HCM) patients, 24 [...] Read more.
MuRF1 [muscle RING (Really Interesting New Gene)-finger protein-1] is an ubiquitin-protein ligase (E3), which encode by TRIM63 (tripartite motif containing 63) gene, playing a crucial role in regulating cardiac muscle size and function through ubiquitylation. Among hypertrophic cardiomyopathy (HCM) patients, 24 TRIM63 variants have been identified, with 1 additional variant linked to restrictive cardiomyopathy. However, only three variants have been previously investigated for their functional effects. The structural impacts of the 25 variants remain unexplored. This study investigated the effects of 25 MuRF1 variants on ubiquitylation activity using in vitro ubiquitylation assays and structural predictions using computational approaches. The variants were generated using site-directed PCR (Polymerase Chain Reaction) mutagenesis and subsequently purified with amylose affinity chromatography. In vitro ubiquitylation assays demonstrated that all 25 variants compromised the ability of MuRF1 to monoubiquitylate a titin fragment (A168-A170), while 17 variants significantly impaired or completely abolished auto-monoubiquitylation. Structural modelling predicted that 10 MuRF1 variants disrupted zinc binding or key stabilising interactions, compromising structural integrity. In contrast, three variants were predicted to enhance the structural stability of MuRF1, while six others were predicted to have no discernible impact on the structure. This study underscores the importance of functional assays and structural predictions in evaluating MuRF1 variant pathogenicity and provides novel insights into mechanisms by which these variants contribute to HCM and related cardiomyopathies. Full article
(This article belongs to the Special Issue Advanced Research on Protein Structure and Protein Dynamics)
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