Microorganisms

13 pages, 2984 KiB

Open AccessArticle

The Influence of Light and Nutrient Starvation on Morphology, Biomass and Lipid Content in Seven Strains of Green Microalgae as a Source of Biodiesel

by Lorenza Rugnini, Catia Rossi, Simonetta Antonaroli, Arnold Rakaj and Laura Bruno

Microorganisms 2020, 8(8), 1254; https://doi.org/10.3390/microorganisms8081254 - 18 Aug 2020

Cited by 17 | Viewed by 3730

Abstract

The development of clean and renewable energy sources is currently one of the most important challenges facing the world. Although research interests in algae-based energy have been increasing in the last decade, only a small percentage of the bewildering diversity exhibited by microalgae [...] Read more.

The development of clean and renewable energy sources is currently one of the most important challenges facing the world. Although research interests in algae-based energy have been increasing in the last decade, only a small percentage of the bewildering diversity exhibited by microalgae has been investigated for biodiesel production. In this work, seven strains of green microalgae belonging to the genera Scenedesmus, Tetradesmus and Desmodesmus were grown in liquid medium with or without a nitrogen (N) source—at two different irradiances (120 ± 20 and 200 ± 20 μmol photons m⁻² s⁻¹)—to evaluate biomass production and FAME (fatty acid methyl esters) content for biodiesel production. The strains of Tetradesmus obliquus and Desmodesmus abundans grown in N-deprived medium showed the highest FAME content (22.0% and 34.6%, respectively); lipid profile characterization highlighted the abundance of saturated FAME (as C16:0 and C18:0) that favors better viscosity (flow properties) and applicability of biodiesel at low temperatures. Light microscopy and confocal laser scanning microscopy observations were employed as a fast method to monitor the vital status of cells and lipid droplet accumulation after Nile red staining in different culture conditions. Full article

(This article belongs to the Section Microbial Biotechnology)

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23 pages, 1956 KiB

Open AccessArticle

Host–Pathogen Interactions between Xanthomonas fragariae and Its Host Fragaria × ananassa Investigated with a Dual RNA-Seq Analysis

by Michael Gétaz, Joanna Puławska, Theo H.M. Smits and Joël F. Pothier

Microorganisms 2020, 8(8), 1253; https://doi.org/10.3390/microorganisms8081253 - 18 Aug 2020

Cited by 9 | Viewed by 4426

Abstract

Strawberry is economically important and widely grown, but susceptible to a large variety of phytopathogenic organisms. Among them, Xanthomonas fragariae is a quarantine bacterial pathogen threatening strawberry productions by causing angular leaf spots. Using whole transcriptome sequencing, the gene expression of both plant [...] Read more.

Strawberry is economically important and widely grown, but susceptible to a large variety of phytopathogenic organisms. Among them, Xanthomonas fragariae is a quarantine bacterial pathogen threatening strawberry productions by causing angular leaf spots. Using whole transcriptome sequencing, the gene expression of both plant and bacteria in planta was analyzed at two time points, 12 and 29 days post inoculation, in order to compare the pathogen and host response between the stages of early visible and of well-developed symptoms. Among 28,588 known genes in strawberry and 4046 known genes in X. fragariae expressed at both time points, a total of 361 plant and 144 bacterial genes were significantly differentially expressed, respectively. The identified higher expressed genes in the plants were pathogen-associated molecular pattern receptors and pathogenesis-related thaumatin encoding genes, whereas the more expressed early genes were related to chloroplast metabolism as well as photosynthesis related coding genes. Most X. fragariae genes involved in host interaction, recognition, and pathogenesis were lower expressed at late-phase infection. This study gives a first insight into the interaction of X. fragariae with its host. The strawberry plant changed gene expression in order to consistently adapt its metabolism with the progression of infection. Full article

(This article belongs to the Special Issue Integrating Science on Xanthomonas and Xylella for Integrated Plant Disease Management)

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18 pages, 2185 KiB

Open AccessArticle

The Absence of C-5 DNA Methylation in Leishmania donovani Allows DNA Enrichment from Complex Samples

by Bart Cuypers, Franck Dumetz, Pieter Meysman, Kris Laukens, Géraldine De Muylder, Jean-Claude Dujardin and Malgorzata Anna Domagalska

Microorganisms 2020, 8(8), 1252; https://doi.org/10.3390/microorganisms8081252 - 18 Aug 2020

Cited by 8 | Viewed by 3332

Abstract

Cytosine C5 methylation is an important epigenetic control mechanism in a wide array of eukaryotic organisms and generally carried out by proteins of the C-5 DNA methyltransferase family (DNMTs). In several protozoans, the status of this mechanism remains elusive, such as in Leishmania [...] Read more.

Cytosine C5 methylation is an important epigenetic control mechanism in a wide array of eukaryotic organisms and generally carried out by proteins of the C-5 DNA methyltransferase family (DNMTs). In several protozoans, the status of this mechanism remains elusive, such as in Leishmania, the causative agent of the disease leishmaniasis in humans and a wide array of vertebrate animals. In this work, we showed that the Leishmania donovani genome contains a C-5 DNA methyltransferase (DNMT) from the DNMT6 subfamily, whose function is still unclear, and verified its expression at the RNA level. We created viable overexpressor and knock-out lines of this enzyme and characterized their genome-wide methylation patterns using whole-genome bisulfite sequencing, together with promastigote and amastigote control lines. Interestingly, despite the DNMT6 presence, we found that methylation levels were equal to or lower than 0.0003% at CpG sites, 0.0005% at CHG sites, and 0.0126% at CHH sites at the genomic scale. As none of the methylated sites were retained after manual verification, we conclude that there is no evidence for DNA methylation in this species. We demonstrated that this difference in DNA methylation between the parasite (no detectable DNA methylation) and the vertebrate host (DNA methylation) allowed enrichment of parasite vs. host DNA using methyl-CpG-binding domain columns, readily available in commercial kits. As such, we depleted methylated DNA from mixes of Leishmania promastigote and amastigote DNA with human DNA, resulting in average Leishmania:human enrichments from 62× up to 263×. These results open a promising avenue for unmethylated DNA enrichment as a pre-enrichment step before sequencing Leishmania clinical samples. Full article

(This article belongs to the Special Issue Leishmania and Leishmaniasis)

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11 pages, 2514 KiB

Open AccessCase Report

First Cases of Natural Infections with Borrelia hispanica in Two Dogs and a Cat from Europe

by Gabriele Margos, Nikola Pantchev, Majda Globokar, Javier Lopez, Jaume Rodon, Leticia Hernandez, Heike Herold, Noelia Salas, Anna Civit and Volker Fingerle

Microorganisms 2020, 8(8), 1251; https://doi.org/10.3390/microorganisms8081251 - 18 Aug 2020

Cited by 8 | Viewed by 3010

Abstract

Canine cases of relapsing fever (RF) borreliosis have been described in Israel and the USA, where two RF species, Borrelia turicatae and Borrelia hermsii, can cause similar clinical signs to the Borrelia persica in dogs and cats reported from Israel, including fever, [...] Read more.

Canine cases of relapsing fever (RF) borreliosis have been described in Israel and the USA, where two RF species, Borrelia turicatae and Borrelia hermsii, can cause similar clinical signs to the Borrelia persica in dogs and cats reported from Israel, including fever, lethargy, anorexia, thrombocytopenia, and spirochetemia. In this report, we describe the first clinical cases of two dogs and a cat from Spain (Cordoba, Valencia, and Seville) caused by the RF species Borrelia hispanica. Spirochetes were present in the blood smears of all three animals, and clinical signs included lethargy, pale mucosa, anorexia, cachexia, or mild abdominal respiration. Laboratory findings, like thrombocytopenia in both dogs, may have been caused by co-infecting pathogens (i.e., Babesia vogeli, confirmed in one dog). Anemia was noticed in one of the dogs and in the cat. Borrelia hispanica was confirmed as an infecting agent by molecular analysis of the 16S rRNA locus. Molecular analysis of housekeeping genes and phylogenetic analyses, as well as successful in vitro culture of the feline isolate confirmed the causative agent as B. hispanica. Full article

(This article belongs to the Special Issue Advance in Tick-Borne Diseases Research)

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18 pages, 1736 KiB

Open AccessReview

Progress in Developing Inhibitors of SARS-CoV-2 3C-Like Protease

by Qingxin Li and CongBao Kang

Microorganisms 2020, 8(8), 1250; https://doi.org/10.3390/microorganisms8081250 - 18 Aug 2020

Cited by 95 | Viewed by 10254

Abstract

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The viral outbreak started in late 2019 and rapidly became a serious health threat to the global population. COVID-19 was declared a pandemic by the World Health Organization in [...] Read more.

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The viral outbreak started in late 2019 and rapidly became a serious health threat to the global population. COVID-19 was declared a pandemic by the World Health Organization in March 2020. Several therapeutic options have been adopted to prevent the spread of the virus. Although vaccines have been developed, antivirals are still needed to combat the infection of this virus. SARS-CoV-2 is an enveloped virus, and its genome encodes polyproteins that can be processed into structural and nonstructural proteins. Maturation of viral proteins requires cleavages by proteases. Therefore, the main protease (3 chymotrypsin-like protease (3CL^pro) or M^pro) encoded by the viral genome is an attractive drug target because it plays an important role in cleaving viral polyproteins into functional proteins. Inhibiting this enzyme is an efficient strategy to block viral replication. Structural studies provide valuable insight into the function of this protease and structural basis for rational inhibitor design. In this review, we describe structural studies on the main protease of SARS-CoV-2. The strategies applied in developing inhibitors of the main protease of SARS-CoV-2 and currently available protein inhibitors are summarized. Due to the availability of high-resolution structures, structure-guided drug design will play an important role in developing antivirals. The availability of high-resolution structures, potent peptidic inhibitors, and diverse compound scaffolds indicate the feasibility of developing potent protease inhibitors as antivirals for COVID-19. Full article

(This article belongs to the Special Issue Antiviral Drug Discovery and Development in the Twenty-First Century)

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20 pages, 2253 KiB

Open AccessFeature PaperReview

Mycobacteriosis in Aquatic Invertebrates: A Review of Its Emergence

by Nadav Davidovich, Danny Morick and Francesca Carella

Microorganisms 2020, 8(8), 1249; https://doi.org/10.3390/microorganisms8081249 - 17 Aug 2020

Cited by 16 | Viewed by 15142

Abstract

Mycobacteriosis is a chronic bacterial disease reported in aquatic and terrestrial animals, including humans. The disease affects a wide range of cultured and wild organisms worldwide. Mycobacteriosis is well-known in aquatic vertebrates (e.g., finfish, marine mammals), while in the last few years, reports [...] Read more.

Mycobacteriosis is a chronic bacterial disease reported in aquatic and terrestrial animals, including humans. The disease affects a wide range of cultured and wild organisms worldwide. Mycobacteriosis is well-known in aquatic vertebrates (e.g., finfish, marine mammals), while in the last few years, reports of its presence in aquatic invertebrates have been on the rise, for both freshwater and marine species. The number of cases is likely to increase as a result of increased awareness, surveillance and availability of diagnostic methods. Domestication of wild aquatic species and the intensification of modern aquaculture are also leading to an increase in the number of reported cases. Moreover, climate changes are affecting fresh and marine aquatic ecosystems. The increasing reports of mycobacteriosis in aquatic invertebrates may also be influenced by global climate warming, which could contribute to the microbes’ development and survival rates, pathogen transmission and host susceptibility. Several species of the genus Mycobacterium have been diagnosed in aquatic invertebrates; a few of them are significant due to their wide host spectrum, economic impact in aquaculture, and zoonotic potential. The impact of mycobacteriosis in aquatic invertebrates is probably underestimated, and there is currently no effective treatment other than facility disinfection. In this review, we provide an overview of the diversity of mycobacterial infections reported in molluscs, crustaceans, cnidarians, echinoderms and sponges. We highlight important issues relating to its pathological manifestation, diagnosis and zoonotic considerations. Full article

(This article belongs to the Section Public Health Microbiology)

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15 pages, 1755 KiB

Open AccessArticle

Niche Differentiation of Active Methane-Oxidizing Bacteria in Estuarine Mangrove Forest Soils in Taiwan

by Yo-Jin Shiau, Chiao-Wen Lin, Yuanfeng Cai, Zhongjun Jia, Yu-Te Lin and Chih-Yu Chiu

Microorganisms 2020, 8(8), 1248; https://doi.org/10.3390/microorganisms8081248 - 17 Aug 2020

Cited by 14 | Viewed by 3928

Abstract

Mangrove forests are one of the important ecosystems in tropical coasts because of their high primary production, which they sustain by sequestering a substantial amount of CO₂ into plant biomass. These forests often experience various levels of inundation and play an important [...] Read more.

Mangrove forests are one of the important ecosystems in tropical coasts because of their high primary production, which they sustain by sequestering a substantial amount of CO₂ into plant biomass. These forests often experience various levels of inundation and play an important role in CH₄ emissions, but the taxonomy of methanotrophs in these systems remains poorly understood. In this study, DNA-based stable isotope probing showed significant niche differentiation in active aerobic methanotrophs in response to niche differentiation in upstream and downstream mangrove soils of the Tamsui estuary in northwestern Taiwan, in which salinity levels differ between winter and summer. Methylobacter and Methylomicrobium-like Type I methanotrophs dominated methane-oxidizing communities in the field conditions and were significantly ¹³C-labeled in both upstream and downstream sites, while Methylobacter were well adapted to high salinity and low temperature. The Type II methanotroph Methylocystis comprised only 10–15% of all the methane oxidizers in the upstream site but less than 5% at the downstream site under field conditions. ¹³C-DNA levels in Methylocystis were significantly lower than those in Type I methanotrophs, while phylogenetic analysis further revealed the presence of novel methane oxidizers that are phylogenetically distantly related to Type Ia in fresh and incubated soils at a downstream site. These results suggest that Type I methanotrophs display niche differentiation associated with environmental differences between upstream and downstream mangrove soils. Full article

(This article belongs to the Special Issue Microbial Cycling of Atmospheric Trace Gases)

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13 pages, 316 KiB

Open AccessReview

Antimicrobial Prosthetic Surfaces in the Oral Cavity—A Perspective on Creative Approaches

by Jorge L. Garaicoa, Amber M. Bates, Gustavo Avila-Ortiz and Kim A. Brogden

Microorganisms 2020, 8(8), 1247; https://doi.org/10.3390/microorganisms8081247 - 17 Aug 2020

Cited by 17 | Viewed by 3284

Abstract

Replacement of missing teeth is an essential component of comprehensive dental care for patients suffering of edentulism. A popular option is implant-supported restorations. However, implant surfaces can become colonized with polymicrobial biofilms containing Candida species that may compromise peri-implant health. To prevent this, [...] Read more.

Replacement of missing teeth is an essential component of comprehensive dental care for patients suffering of edentulism. A popular option is implant-supported restorations. However, implant surfaces can become colonized with polymicrobial biofilms containing Candida species that may compromise peri-implant health. To prevent this, implant components may be treated with a variety of coatings to create surfaces that either repel the attachment of viable microorganisms or kill microorganisms on contact. These coatings may consist of nanoparticles of pure elements (more commonly silver, copper, and zinc), sanitizing agents and disinfectants (quaternary ammonium ions and chlorhexidine), antibiotics (cefalotin, vancomycin, and gentamicin), or antimicrobial peptides (AMPs). AMPs in bioactive coatings have a number of advantages. They elicit a protective action against pathogens, inhibit the formation of biofilms, are less toxic to host tissues, and do not prompt inflammatory responses. Furthermore, many of these coatings may involve unique delivery systems to direct their antimicrobial capacity against pathogens, but not commensals. Coatings may also contain multiple antimicrobial substances to widen antimicrobial activity across multiple microbial species. Here, we compiled relevant information about a variety of creative approaches used to generate antimicrobial prosthetic surfaces in the oral cavity with the purpose of facilitating implant integration and peri-implant tissue health. Full article

(This article belongs to the Special Issue The Role of the Oral Mucosa and Salivary Gland in Anti-Fungal Immunity)

20 pages, 935 KiB

Open AccessArticle

Association of the Gut Microbiota with Weight-Loss Response within a Retail Weight-Management Program

by Samitinjaya Dhakal, Lacey McCormack and Moul Dey

Microorganisms 2020, 8(8), 1246; https://doi.org/10.3390/microorganisms8081246 - 16 Aug 2020

Cited by 22 | Viewed by 4954

Abstract

Retail programs offer popular weight-loss options amid the ongoing obesity crisis. However, research on weight-loss outcomes within such programs is limited. This prospective-cohort observational study enrolled 58 men and women between ages 20 and 72 years from a retail program to assess the [...] Read more.

Retail programs offer popular weight-loss options amid the ongoing obesity crisis. However, research on weight-loss outcomes within such programs is limited. This prospective-cohort observational study enrolled 58 men and women between ages 20 and 72 years from a retail program to assess the influence of client features on energy-restriction induced weight-loss response. DESeq2 in R-studio, a linear regression model adjusting for significantly correlating covariates, and Wilcoxon signed-rank and Kruskal–Wallis for within- and between-group differences, respectively, were used for data analyses. An average 10% (~10 kg) reduction in baseline-weight along with lower total-, android-, gynoid-, and android:gynoid-fat were observed at Week 12 (all, p < 0.05). Fifty percent of participants experienced a higher response, losing an average of 14.5 kg compared to 5.9 kg in the remaining low-response group (p < 0.0001). Hemoglobin-A1C (p = 0.005) and heart rate (p = 0.079) reduced in the high-response group only. Fat mass and A1C correlated when individuals had high android:gynoid fat (r = 0.55, p = 0.008). Gut-microbial β-diversity was associated with BMI, body fat%, and android-fat (all, p < 0.05). Microbiota of the high-response group had a higher baseline OTU-richness (p = 0.02) as well as differential abundance and/or associations with B. eggerthi, A. muciniphila, Turicibacter, Prevotella, and Christensenella (all, p/p_adj < 0.005). These results show that intestinal microbiota as well as sex and body composition differences may contribute to variable weight-loss response. This highlights the importance of various client features in the context of real-world weight control efforts. Full article

(This article belongs to the Special Issue The Human Gut Microbiome, Diets and Health)

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25 pages, 2469 KiB

Open AccessArticle

Ibuprofen Degradation and Associated Bacterial Communities in Hyporheic Zone Sediments

by Cyrus Rutere, Kirsten Knoop, Malte Posselt, Adrian Ho and Marcus A. Horn

Microorganisms 2020, 8(8), 1245; https://doi.org/10.3390/microorganisms8081245 - 16 Aug 2020

Cited by 29 | Viewed by 5031

Abstract

Ibuprofen, a non-steroidal anti-inflammatory pain reliever, is among pharmaceutical residues of environmental concern ubiquitously detected in wastewater effluents and receiving rivers. Thus, ibuprofen removal potentials and associated bacteria in the hyporheic zone sediments of an impacted river were investigated. Microbially mediated ibuprofen degradation [...] Read more.

Ibuprofen, a non-steroidal anti-inflammatory pain reliever, is among pharmaceutical residues of environmental concern ubiquitously detected in wastewater effluents and receiving rivers. Thus, ibuprofen removal potentials and associated bacteria in the hyporheic zone sediments of an impacted river were investigated. Microbially mediated ibuprofen degradation was determined in oxic sediment microcosms amended with ibuprofen (5, 40, 200, and 400 µM), or ibuprofen and acetate, relative to an un-amended control. Ibuprofen was removed by the original sediment microbial community as well as in ibuprofen-enrichments obtained by re-feeding of ibuprofen. Here, 1-, 2-, 3-hydroxy- and carboxy-ibuprofen were the primary transformation products. Quantitative real-time PCR analysis revealed a significantly higher 16S rRNA abundance in ibuprofen-amended relative to un-amended incubations. Time-resolved microbial community dynamics evaluated by 16S rRNA gene and 16S rRNA analyses revealed many new ibuprofen responsive taxa of the Acidobacteria, Actinobacteria, Bacteroidetes, Gemmatimonadetes, Latescibacteria, and Proteobacteria. Two ibuprofen-degrading strains belonging to the genera Novosphingobium and Pseudomonas were isolated from the ibuprofen-enriched sediments, consuming 400 and 300 µM ibuprofen within three and eight days, respectively. The collective results indicated that the hyporheic zone sediments sustain an efficient biotic (micro-)pollutant degradation potential, and hitherto unknown microbial diversity associated with such (micro)pollutant removal. Full article

(This article belongs to the Special Issue Terrestrial Ecotoxicology- How Biocides of Building Materials Impact Soil Microbial Communities)

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19 pages, 7243 KiB

Open AccessArticle

Assessing the Pyloric Caeca and Distal Gut Microbiota Correlation with Flesh Color in Atlantic Salmon (Salmo salar L., 1758)

by Chan D. H. Nguyen, Gianluca Amoroso, Tomer Ventura and Abigail Elizur

Microorganisms 2020, 8(8), 1244; https://doi.org/10.3390/microorganisms8081244 - 16 Aug 2020

Cited by 19 | Viewed by 9429

Abstract

The Atlantic salmon (Salmo salar L., 1758) is a temperate fish species native to the northern Atlantic Ocean. The distinctive pink–red flesh color (i.e., pigmentation) significantly affects the market price. Flesh paleness leads to customer dissatisfaction, a loss of competitiveness, a drop [...] Read more.

The Atlantic salmon (Salmo salar L., 1758) is a temperate fish species native to the northern Atlantic Ocean. The distinctive pink–red flesh color (i.e., pigmentation) significantly affects the market price. Flesh paleness leads to customer dissatisfaction, a loss of competitiveness, a drop in product value and, consequently, severe economic losses. This work extends our knowledge on salmonid carotenoid dynamics to include the interaction between the gut microbiota and flesh color. A significant association between the flesh color and abundance of specific bacterial communities in the gut microbiota suggests that color may be affected either by seeding resilient beneficial bacteria or by inhibiting the negative effect of pathogenic bacteria. We sampled 96 fish, which covered all phenotypes of flesh color, including the average color and the evenness of color of different areas of the fillet, at both the distal intestine and the pyloric caeca of each individual, followed by 16S rRNA sequencing at the V3-V4 region. The microbiota profiles of these two gut regions were significantly different; however, there was a consistency in the microbiota, which correlated with the flesh color. Moreover, the pyloric caeca microbiota also showed high correlation with the evenness of the flesh color (beta diversity index, PERMANOVA, p = 0.002). The results from the pyloric caeca indicate that Carnobacterium, a group belonging to the lactic acid bacteria, is strongly related to the flesh color and the evenness of the color between the flesh areas. Full article

(This article belongs to the Section Gut Microbiota)

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16 pages, 2695 KiB

Open AccessArticle

Improving Biodegradation of Clofibric Acid by Trametes pubescens through the Design of Experimental Tools

by Claudia Veronica Ungureanu, Lidia Favier and Gabriela Elena Bahrim

Microorganisms 2020, 8(8), 1243; https://doi.org/10.3390/microorganisms8081243 - 15 Aug 2020

Cited by 12 | Viewed by 2407

Abstract

Clofibric acid (CLF) is the main pharmacologically active metabolite in composition of the pharmaceutical products used for controlling blood lipid content. This xenobiotic compound is highly persistent in the aquatic environment and passes unchanged or poorly transformed in wastewater treatment plants. A white-rot [...] Read more.

Clofibric acid (CLF) is the main pharmacologically active metabolite in composition of the pharmaceutical products used for controlling blood lipid content. This xenobiotic compound is highly persistent in the aquatic environment and passes unchanged or poorly transformed in wastewater treatment plants. A white-rot fungal strain of Trametes pubescens was previously selected, for its ability for clofibric acid biodegradation (up to 30%) during cultivation in submerged system under aerobic conditions at an initial CLF concentration of 15 mg L⁻¹. Plackett-Burman design (PBD) and response surface methodology (RSM) were used for experimental planning, mathematical modelling and statistical analysis of data of the biotechnological process of CLF biotransformation by Trametes pubescens fungal strain. After optimization, the capacity of the selected Trametes pubescens strain to degrade CLF was increased by cultivation in a liquid medium containing 3 g·L⁻¹ yeast extract, 15 g·L⁻¹ peptone, 5 g·L⁻¹ glucose and mineral salts, inoculated at 2% (v/v) vegetative inoculum and cultivated at pH 5.5, during 14 days at 25 °C and 135 rpm. In these optimized biotechnological conditions, the CLF biotransformation yield was 60%. Full article

(This article belongs to the Section Environmental Microbiology)

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18 pages, 2033 KiB

Open AccessArticle

Do Metabolomics and Taxonomic Barcode Markers Tell the Same Story about the Evolution of Saccharomyces sensu stricto Complex in Fermentative Environments?

by Luca Roscini, Angela Conti, Debora Casagrande Pierantoni, Vincent Robert, Laura Corte and Gianluigi Cardinali

Microorganisms 2020, 8(8), 1242; https://doi.org/10.3390/microorganisms8081242 - 15 Aug 2020

Cited by 3 | Viewed by 2724

Abstract

Yeast taxonomy was introduced based on the idea that physiological properties would help discriminate species, thus assuming a strong link between physiology and taxonomy. However, the instability of physiological characteristics within species configured them as not ideal markers for species delimitation, shading the [...] Read more.

Yeast taxonomy was introduced based on the idea that physiological properties would help discriminate species, thus assuming a strong link between physiology and taxonomy. However, the instability of physiological characteristics within species configured them as not ideal markers for species delimitation, shading the importance of physiology and paving the way to the DNA-based taxonomy. The hypothesis of reconnecting taxonomy with specific traits from phylogenies has been successfully explored for Bacteria and Archaea, suggesting that a similar route can be traveled for yeasts. In this framework, thirteen single copy loci were used to investigate the predictability of complex Fourier Transform InfaRed spectroscopy (FTIR) and High-performance Liquid Chromatography–Mass Spectrometry (LC-MS) profiles of the four historical species of the Saccharomyces sensu stricto group, both on resting cells and under short-term ethanol stress. Our data show a significant connection between the taxonomy and physiology of these strains. Eight markers out of the thirteen tested displayed high correlation values with LC-MS profiles of cells in resting condition, confirming the low efficacy of FTIR in the identification of strains of closely related species. Conversely, most genetic markers displayed increasing trends of correlation with FTIR profiles as the ethanol concentration increased, according to their role in the cellular response to different type of stress. Full article

(This article belongs to the Special Issue Yeast in Winemaking)

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18 pages, 3944 KiB

Open AccessArticle

Microbial Diversity of Fermented Greek Table Olives of Halkidiki and Konservolia Varieties from Different Regions as Revealed by Metagenomic Analysis

by Konstantina Argyri, Agapi I. Doulgeraki, Evanthia Manthou, Athena Grounta, Anthoula A. Argyri, George-John E. Nychas and Chrysoula C. Tassou

Microorganisms 2020, 8(8), 1241; https://doi.org/10.3390/microorganisms8081241 - 14 Aug 2020

Cited by 34 | Viewed by 5775

Abstract

Current information from conventional microbiological methods on the microbial diversity of table olives is insufficient. Next-generation sequencing (NGS) technologies allow comprehensive analysis of their microbial community, providing microbial identity of table olive varieties and their designation of origin. The purpose of this study [...] Read more.

Current information from conventional microbiological methods on the microbial diversity of table olives is insufficient. Next-generation sequencing (NGS) technologies allow comprehensive analysis of their microbial community, providing microbial identity of table olive varieties and their designation of origin. The purpose of this study was to evaluate the bacterial and yeast diversity of fermented olives of two main Greek varieties collected from different regions—green olives, cv. Halkidiki, from Kavala and Halkidiki and black olives, cv. Konservolia, from Magnesia and Fthiotida—via conventional microbiological methods and NGS. Total viable counts (TVC), lactic acid bacteria (LAB), yeast and molds, and Enterobacteriaceae were enumerated. Microbial genomic DNA was directly extracted from the olives’ surface and subjected to NGS for the identification of bacteria and yeast communities. Lactobacillaceae was the most abundant family in all samples. In relation to yeast diversity, Phaffomycetaceae was the most abundant yeast family in Konservolia olives from the Magnesia region, while Pichiaceae dominated the yeast microbiota in Konservolia olives from Fthiotida and in Halkidiki olives from both regions. Further analysis of the data employing multivariate analysis allowed for the first time the discrimination of cv. Konservolia and cv. Halkidiki table olives according to their geographical origin. Full article

(This article belongs to the Special Issue Food Microbial Diversity)

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16 pages, 1018 KiB

Open AccessArticle

Drug Resistance Determinants in Clinical Isolates of Enterococcus faecalis in Bangladesh: Identification of Oxazolidinone Resistance Gene optrA in ST59 and ST902 Lineages

by Sangjukta Roy, Meiji Soe Aung, Shyamal Kumar Paul, Salma Ahmed, Nazia Haque, Emily Rahman Khan, Tridip Kanti Barman, Arup Islam, Sahida Abedin, Chand Sultana, Anindita Paul, Muhammad Akram Hossain, Noriko Urushibara, Mitsuyo Kawaguchiya, Ayako Sumi and Nobumichi Kobayashi

Microorganisms 2020, 8(8), 1240; https://doi.org/10.3390/microorganisms8081240 - 14 Aug 2020

Cited by 14 | Viewed by 3263

Abstract

Enterococcus faecalis is one of the major causes of urinary tract infection, showing acquired resistance to various classes of antimicrobials. The objective of this study was to determine the prevalence of drug resistance and its genetic determinants for E. faecalis clinical isolates in [...] Read more.

Enterococcus faecalis is one of the major causes of urinary tract infection, showing acquired resistance to various classes of antimicrobials. The objective of this study was to determine the prevalence of drug resistance and its genetic determinants for E. faecalis clinical isolates in north-central Bangladesh. Among a total of 210 E. faecalis isolates, isolated from urine, the resistance rates to erythromycin, levofloxacin, and gentamicin (high level) were 85.2, 45.7, and 11.4%, respectively, while no isolates were resistant to ampicillin, vancomycin and teicoplanin. The most prevalent resistance gene was erm(B) (97%), and any of the four genes encoding aminoglycoside modifying enzyme (AME) were detected in 99 isolates (47%). The AME gene aac(6′)-Ie-aph(2”)-Ia was detected in 46 isolates (21.9%) and was diverse in terms of IS256-flanking patterns, which were associated with resistance level to gentamicin. Tetracycline resistance was ascribable to tet(M) (61%) and tet(L) (38%), and mutations in the quinolone resistance-determining region of both GyrA and ParC were identified in 44% of isolates. Five isolates (2.4%) exhibited non-susceptibility to linezolide (MIC, 4 μg/mL), and harbored the oxazolidinone resistance gene optrA, which was located in a novel genetic cluster containing the phenicol exporter gene fexA. The optrA-positive isolates belonged to ST59, ST902, and ST917 (CC59), while common lineages of other multiple drug-resistant isolates were ST6, ST28, CC16, and CC116. The present study first revealed the prevalence of drug resistance determinants of E. faecalis and their genetic profiles in Bangladesh. Full article

(This article belongs to the Section Antimicrobial Agents and Resistance)

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13 pages, 1417 KiB

Open AccessArticle

EMS-Induced Mutagenesis of Clostridium carboxidivorans for Increased Atmospheric CO₂ Reduction Efficiency and Solvent Production

by Naoufal Lakhssassi, Azam Baharlouei, Jonas Meksem, Scott D. Hamilton-Brehm, David A. Lightfoot, Khalid Meksem and Yanna Liang

Microorganisms 2020, 8(8), 1239; https://doi.org/10.3390/microorganisms8081239 - 14 Aug 2020

Cited by 9 | Viewed by 3264

Abstract

Clostridium carboxidivorans (P7) is one of the most important solvent-producing bacteria capable of fermenting syngas (CO, CO₂, and H₂) to produce chemical commodities when grown as an autotroph. This study aimed to develop ethyl methanesulfonate (EMS)-induced P7 mutants that [...] Read more.

Clostridium carboxidivorans (P7) is one of the most important solvent-producing bacteria capable of fermenting syngas (CO, CO₂, and H₂) to produce chemical commodities when grown as an autotroph. This study aimed to develop ethyl methanesulfonate (EMS)-induced P7 mutants that were capable of growing in the presence of CO₂ as a unique source of carbon with increased solvent formation and atmospheric CO₂ reduction to limit global warming. Phenotypic analysis including growth and end product characterization of the P7 wild type (WT) demonstrated that this strain grew better at 25 °C than 37 °C when CO₂ served as the only source of carbon. In the current study, 55 mutagenized P7-EMS mutants were developed by using 100 mM and 120 mM EMS. Interestingly, using a forward genetic approach, three out of the 55 P7-EMS mutants showed a significant increase in ethanol, butyrate, and butanol production. The three P7-EMS mutants presented on average a 4.68-fold increase in concentrations of ethanol when compared to the P7-WT. Butyric acid production from 3 P7-EMS mutants contained an average of a 3.85 fold increase over the levels observed in the P7-WT cultures under the same conditions (CO₂ only). In addition, one P7-EMS mutant presented butanol production (0.23 ± 0.02 g/L), which was absent from the P7-WT under CO₂ conditions. Most of the P7-EMS mutants showed stability of the obtained end product traits after three transfers. Most importantly, the amount of reduced atmospheric CO₂ increased up to 8.72 times (0.21 g/Abs) for ethanol production and up to 8.73 times higher (0.16 g/Abs) for butyrate than the levels contained in the P7-WT. Additionally, to produce butanol, the P7-EMS_III-J mutant presented 0.082 g/Abs of CO₂ reduction. This study demonstrated the feasibility and effectiveness of employing EMS mutagenesis in generating solvent-producing anaerobic bacteria mutants with improved and novel product formation and increased atmospheric CO₂ reduction efficiency. Full article

(This article belongs to the Special Issue Anaerobes in Biogeochemical Cycles)

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15 pages, 4761 KiB

Open AccessArticle

Impact of Cyanidin-3-Glucoside on Gut Microbiota and Relationship with Metabolism and Inflammation in High Fat-High Sucrose Diet-Induced Insulin Resistant Mice

by Fei Huang, Ruozhi Zhao, Min Xia and Garry X. Shen

Microorganisms 2020, 8(8), 1238; https://doi.org/10.3390/microorganisms8081238 - 14 Aug 2020

Cited by 41 | Viewed by 3548

Abstract

The present study assessed the effects of freeze-dried cyanidin-3-glucoside (C3G), an anthocyanin enriched in dark-red berries, compared to Saskatoon berry powder (SBp) on metabolism, inflammatory markers and gut microbiota in high fat-high sucrose (HFHS) diet-induced insulin-resistant mice. Male C57 BL/6J mice received control, [...] Read more.

The present study assessed the effects of freeze-dried cyanidin-3-glucoside (C3G), an anthocyanin enriched in dark-red berries, compared to Saskatoon berry powder (SBp) on metabolism, inflammatory markers and gut microbiota in high fat-high sucrose (HFHS) diet-induced insulin-resistant mice. Male C57 BL/6J mice received control, HFHS, HFHS + SBp (8.0 g/kg/day) or HFHS + C3G (7.2 mg/kg/day, equivalent C3G in SBp) diet for 11 weeks. The HFHS diet significantly increased plasma levels of glucose, cholesterol, triglycerides, insulin resistance and inflammatory markers. The HFHS + SBp diet increased the Bacteroidetes/Firmicutes (B/F) ratio and relative abundance of Muriculaceae family bacteria in mouse feces detected using 16S rRNA gene sequencing. The HFHS + SBp or HFHS + C3G diet attenuated glucose, lipids, insulin resistance and inflammatory markers, and increased the B/F ratio and Muriculaceae relative abundance compared to the HFHS diet alone. The relative abundances of Muriculaceae negatively correlated with body weight, glucose, lipids, insulin resistance and inflammatory mediators. Functional predication analysis suggested that the HFHS diet upregulated gut bacteria genes involved in inflammation, and downregulated bacteria involved in metabolism. C3G and SBp partially neutralized HFHS diet-induced alterations of gut bacteria. The results suggest that C3G is a potential prebiotic, mitigating HFHS diet-induced disorders in metabolism, inflammation and gut dysbiosis, and that C3G contributes to the metabolic beneficial effects of SBp. Full article

(This article belongs to the Section Gut Microbiota)

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26 pages, 1299 KiB

Open AccessReview

Patents on Endophytic Fungi for Agriculture and Bio- and Phytoremediation Applications

by Humberto E. Ortega, Daniel Torres-Mendoza and Luis Cubilla-Rios

Microorganisms 2020, 8(8), 1237; https://doi.org/10.3390/microorganisms8081237 - 14 Aug 2020

Cited by 43 | Viewed by 5653

Abstract

Plant endophytic fungi spend all or part of their lives inside host tissues without causing disease symptoms. They can colonize the plant to protect against predators, pathogens and abiotic stresses generated by drought, salinity, high concentrations of heavy metals, UV radiation and temperature [...] Read more.

Plant endophytic fungi spend all or part of their lives inside host tissues without causing disease symptoms. They can colonize the plant to protect against predators, pathogens and abiotic stresses generated by drought, salinity, high concentrations of heavy metals, UV radiation and temperature fluctuations. They can also promote plant growth through the biosynthesis of phytohormones and nutrient acquisition. In recent years, the study of endophytic fungi for biological control of plant diseases and pests has been intensified to try to reduce the ecological and public health impacts due the use of chemicals and the emergence of fungicide resistance. In this review, we examine 185 patents related to endophytic fungi (from January 1988 to December 2019) and discuss their applicability for abiotic stress tolerance and growth promotion of plants, as agents for biocontrol of herbivores and plant pathogens and bio- and phytoremediation applications. Full article

(This article belongs to the Section Microbial Biotechnology)

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5 pages, 240 KiB

Open AccessCase Report

Compassionate Use of Cefiderocol to Treat a Case of Prosthetic Joint Infection Due to Extensively Drug-Resistant Enterobacter hormaechei

by Soline Siméon, Laurent Dortet, Frédérique Bouchand, Anne-Laure Roux, Rémy A. Bonnin, Clara Duran, Jean-Winoc Decousser, Simon Bessis, Benjamin Davido, Grégory Sorriaux and Aurélien Dinh

Microorganisms 2020, 8(8), 1236; https://doi.org/10.3390/microorganisms8081236 - 13 Aug 2020

Cited by 23 | Viewed by 3063

Abstract

We report the case of a 67-year old man with a right knee prosthetic joint infection due to extensively drug-resistant Enterobacter hormaechei. The resistance phenotype was due to the overproduction of the intrinsic cephalosporinase (ACT-5) associated with the production of three acquired [...] Read more.

We report the case of a 67-year old man with a right knee prosthetic joint infection due to extensively drug-resistant Enterobacter hormaechei. The resistance phenotype was due to the overproduction of the intrinsic cephalosporinase (ACT-5) associated with the production of three acquired β-lactamases (CTX-M-15, TEM-1B and OXA-1), and a putative membrane decreased permeability. He was first treated with colistin-tigecyclin due to adverse drug reactions; treatment was switched to cefiderocol for a 12-week antibiotic duration, with a favorable outcome. Full article

(This article belongs to the Special Issue Molecular Epidemiology of Antimicrobial Resistance)

19 pages, 969 KiB

Open AccessArticle

Evaluation of the Diagnostic Potential of Recombinant Coxiella burnetii Com1 in an ELISA for the Diagnosis of Q Fever in Sheep, Goats and Cattle

by Mareike Stellfeld, Claudia Gerlach, Ina-Gabriele Richter, Peter Miethe, Dominika Fahlbusch, Birgitta Polley, Reinhard Sting, Martin Pfeffer, Heinrich Neubauer and Katja Mertens-Scholz

Microorganisms 2020, 8(8), 1235; https://doi.org/10.3390/microorganisms8081235 - 13 Aug 2020

Cited by 13 | Viewed by 3895

Abstract

Coxiella burnetii is the causative agent of Q fever, a zoonosis infecting domestic ruminants and humans. Currently used routine diagnostic tools offer limited sensitivity and specificity and symptomless infected animals may be missed. Therefore, diagnostic tools of higher sensitivity and specificity must be [...] Read more.

Coxiella burnetii is the causative agent of Q fever, a zoonosis infecting domestic ruminants and humans. Currently used routine diagnostic tools offer limited sensitivity and specificity and symptomless infected animals may be missed. Therefore, diagnostic tools of higher sensitivity and specificity must be developed. For this purpose, the C. burnetii outer membrane protein Com1 was cloned and expressed in Escherichia coli. The His-tagged recombinant protein was purified and used in an indirect enzyme-linked immunosorbent assay (ELISA). Assay performance was tested with more than 400 positive and negative sera from sheep, goats and cattle from 36 locations. Calculation of sensitivity and specificity was undertaken using receiver operating characteristic (ROC) curves. The sensitivities and specificities for sheep were 85% and 68% (optical density at 450nm, OD₄₅₀ cut-off value 0.32), for goats 94% and 77% (OD₄₅₀ cut-off value 0.23) and for cattle 71% and 70% (OD₄₅₀ cut-off value 0.18), respectively. These results correspond to excellent, outstanding and acceptable discrimination of positive and negative sera. In summary, recombinant Com1 can provide a basis for more sensitive and specific diagnostic tools in veterinary medicine. Full article

(This article belongs to the Section Public Health Microbiology)

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18 pages, 3826 KiB

Open AccessArticle

Four New Genes of Cyanobacterium Synechococcus elongatus PCC 7942 Are Responsible for Sensitivity to 2-Nonanone

by Olga A. Koksharova, Alexandra A. Popova, Vladimir A. Plyuta and Inessa A. Khmel

Microorganisms 2020, 8(8), 1234; https://doi.org/10.3390/microorganisms8081234 - 13 Aug 2020

Cited by 3 | Viewed by 3840

Abstract

Microbial volatile organic compounds (VOCs) are cell metabolites that affect many physiological functions of prokaryotic and eukaryotic organisms. Earlier we have demonstrated the inhibitory effects of soil bacteria volatiles, including ketones, on cyanobacteria. Cyanobacteria are very sensitive to ketone action. To investigate the [...] Read more.

Microbial volatile organic compounds (VOCs) are cell metabolites that affect many physiological functions of prokaryotic and eukaryotic organisms. Earlier we have demonstrated the inhibitory effects of soil bacteria volatiles, including ketones, on cyanobacteria. Cyanobacteria are very sensitive to ketone action. To investigate the possible molecular mechanisms of the ketone 2-nonanone influence on cyanobacterium Synechococcus elongatus PCC 7942, we applied a genetic approach. After Tn5-692 transposon mutagenesis, several 2-nonanone resistant mutants have been selected. Four different mutant strains were used for identification of the impaired genes (Synpcc7942_1362, Synpcc7942_0351, Synpcc7942_0732, Synpcc7942_0726) that encode correspondingly: 1) a murein-peptide ligase Mpl that is involved in the biogenesis of cyanobacteria cell wall; 2) a putative ABC transport system substrate-binding proteins MlaD, which participates in ABC transport system that maintains lipid asymmetry in the gram-negative outer membrane by aberrantly localized phospholipids transport from outer to inner membranes of bacterial cells; 3) a conserved hypothetical protein that is encoding by gene belonging to phage gene cluster in Synechococcus elongatus PCC 7942 genome; 4) a protein containing the VRR-NUC (virus-type replication-repair nuclease) domain present in restriction-modification enzymes involved in replication and DNA repair. The obtained results demonstrated that 2-nonanone may have different targets in Synechococcus elongatus PCC 7942 cells. Among them are proteins involved in the biogenesis and functioning of the cyanobacteria cell wall (Synpcc7942_1362, Synpcc7942_0351, Synpcc7942_0732) and protein participating in stress response at DNA restriction-modification level (Synpcc7942_0726). This paper is the first report about the genes that encode protein products, which can be affected by 2-nonanone. Full article

(This article belongs to the Section Environmental Microbiology)

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13 pages, 1336 KiB

Open AccessArticle

Rapid Detection and Antibiotic Susceptibility of Uropathogenic Escherichia coli by Flow Cytometry

by Alexandra Mihaela Velican, Luminiţa Măruţescu, Crina Kamerzan, Violeta Corina Cristea, Otilia Banu, Elvira Borcan and Mariana-Carmen Chifiriuc

Microorganisms 2020, 8(8), 1233; https://doi.org/10.3390/microorganisms8081233 - 13 Aug 2020

Cited by 10 | Viewed by 3390

Abstract

Background: Early preliminary data on antibiotic resistance patterns available before starting the empiric therapy of urinary tract infections (UTIs) in patients with risk factors for acquiring antibiotic resistance could improve both clinical and epidemiological outcomes. The aim of the present study was two-fold: [...] Read more.

Background: Early preliminary data on antibiotic resistance patterns available before starting the empiric therapy of urinary tract infections (UTIs) in patients with risk factors for acquiring antibiotic resistance could improve both clinical and epidemiological outcomes. The aim of the present study was two-fold: (i) to assess the antibiotic susceptibility of uropathogenic Escherichia coli isolates, exhibiting different antibiotic resistance phenotypes, directly in artificially contaminated urine samples using a flow cytometry (FC) based protocol; (ii) to optimize the protocol on urine samples deliberately contaminated with bacterial suspensions prepared from uropathogenic E. coli strains. Results: The results of the FC based antimicrobial susceptibility testing (AST) protocol were compared with the reference AST methods results (disk diffusion and broth microdilution) for establishing the sensitivity and specificity. The proposed FC protocol allowed the detection and quantification of uropathogenic E. coli strains susceptibility to nitrofurantoin, trimethoprim–sulfamethoxazole, ciprofloxacin, and ceftriaxone within 4 h after the inoculation of urine specimens. The early availability of preliminary antibiotic susceptibility results provided by direct analysis of clinical specimens could essentially contribute to a more targeted emergency therapy of UTIs in the anticipation of AST results obtained by reference methodology. Conclusions: This method will increase the therapeutic success rate and help to prevent the emergence and dissemination of drug resistant pathogens. Full article

(This article belongs to the Special Issue Rapid Diagnosis of Microbial Pathogens)

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9 pages, 1090 KiB

Open AccessCommunication

Genomic Insight of VIM-harboring IncA Plasmid from a Clinical ST69 Escherichia coli Strain in Italy

by Vittoria Mattioni Marchetti, Ibrahim Bitar, Aurora Piazza, Alessandra Mercato, Elena Fogato, Jaroslav Hrabak and Roberta Migliavacca

Microorganisms 2020, 8(8), 1232; https://doi.org/10.3390/microorganisms8081232 - 12 Aug 2020

Cited by 5 | Viewed by 3279

Abstract

Background: VIM (Verona Integron-encoded Metallo-beta-lactamase) is a member of the Metallo-Beta-Lactamases (MBLs), and is able to hydrolyze all beta-lactams antibiotics, except for monobactams, and including carbapenems. Here we characterize a VIM-producing IncA plasmid isolated from a clinical ST69 Escherichia coli strain from [...] Read more.

Background: VIM (Verona Integron-encoded Metallo-beta-lactamase) is a member of the Metallo-Beta-Lactamases (MBLs), and is able to hydrolyze all beta-lactams antibiotics, except for monobactams, and including carbapenems. Here we characterize a VIM-producing IncA plasmid isolated from a clinical ST69 Escherichia coli strain from an Italian Long-Term Care Facility (LTCF) inpatient. Methods: An antimicrobial susceptibility test and conjugation assay were carried out, and the transferability of the bla_VIM-type gene was confirmed in the transconjugant. Whole-genome sequencing (WGS) of the strain 550 was performed using the Sequel I platform. Genome assembly was performed using “Microbial Assembly”. Genomic analysis was conducted by uploading the contigs to ResFinder and PlasmidFinder databases. Results: Assembly resulted in three complete circular contigs: the chromosome (4,962,700 bp), an IncA plasmid (p550_IncA_VIM_1; 162,608 bp), harboring genes coding for aminoglycoside resistance (aac(6′)-Ib4, ant(3″)-Ia, aph(3″)-Ib, aph(3′)-XV, aph(6)-Id), beta-lactam resistance (bla_SHV-12, bla_VIM-1), macrolides resistance (mph(A)), phenicol resistance (catB2), quinolones resistance (qnrS1), sulphonamide resistance (sul1, sul2), and trimethoprim resistance (dfrA14), and an IncK/Z plasmid (p550_IncB_O_K_Z; 100,306 bp), free of antibiotic resistance genes. Conclusions: The increase in reports of IncA plasmids bearing different antimicrobial resistance genes highlights the overall important role of IncA plasmids in disseminating carbapenemase genes, with a preference for the bla_VIM-1 gene in Italy. Full article

(This article belongs to the Special Issue Bacterial Responses to Environmental Stress and Their Specific Contribution to Escherichia coli and Vibrio spp. Survival and Virulence)

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14 pages, 1363 KiB

Open AccessArticle

Enhancement of the Molecular and Serological Assessment of Hepatitis E Virus in Milk Samples

by Ibrahim M. Sayed, Ahmed R. A. Hammam, Mohamed Salem Elfaruk, Khalid A. Alsaleem, Marwa A. Gaber, Amgad A. Ezzat, Eman H. Salama, Amal A. Elkhawaga and Mohamed A. El-Mokhtar

Microorganisms 2020, 8(8), 1231; https://doi.org/10.3390/microorganisms8081231 - 12 Aug 2020

Cited by 14 | Viewed by 3685

Abstract

Hepatitis E virus (HEV) infection is endemic in developing and developed countries. HEV was reported to be excreted in the milk of ruminants, raising the possibility of transmission of HEV infection through the ingestion of contaminated milk. Therefore, the detection of HEV markers [...] Read more.

Hepatitis E virus (HEV) infection is endemic in developing and developed countries. HEV was reported to be excreted in the milk of ruminants, raising the possibility of transmission of HEV infection through the ingestion of contaminated milk. Therefore, the detection of HEV markers in milk samples becomes pivotal. However, milk includes inhibitory components that affect HEV detection assays. Previously it was reported that dilution of milk matrix improves the performance of HEV molecular assay, however, the dilution of milk samples is not the best strategy especially when the contaminated milk sample has a low HEV load. Therefore, the objective of this study is to compare the effect of extraction procedures on the efficiency of HEV RNA detection in undiluted milk samples. In addition, we assessed the effect of the removal of milk components such as fats and casein on the performance of the molecular and serological assays of HEV. Phosphate buffered saline (PBS) and different milk matrices (such as whole milk, skim milk, and milk serum) were inoculated with different HEV inoculums and subjected to two different extraction procedures. Method A includes manual extraction using spin column-based extraction, while method B includes silica-based automated extraction. Method A was more sensitive than method B in the whole milk and skim milk matrices with a LoD_95% of 300 IU/mL, and virus recovery yield of 47%. While the sensitivity and performance of method B were significantly improved using the milk serum matrix, with LoD_95% of 96 IU/mL. Interestingly, retesting HEV positive milk samples using the high sensitivity assay based on method B extraction and milk serum matrix increased the HEV RNA detection rate to 2-fold. Additionally, the performance of HEV serological assays such as anti-HEV IgG and HEV Ag in the milk samples was improved after the removal of the fat globules from the milk matrix. In conclusion, HEV RNA assay is affected by the components of milk and the extraction procedure. Removal of inhibitory substances, such as fat and casein from the milk sample increased the performance of HEV molecular and serological assays which will be suitable for the low load HEV milk with no further dilutions. Full article

(This article belongs to the Special Issue Enteric Virus Detection: Recent Developments and Application in Food and Waters)

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11 pages, 3136 KiB

Open AccessArticle

Involvement of the cbb₃-Type Terminal Oxidase in Growth Competition of Bacteria, Biofilm Formation, and in Switching between Denitrification and Aerobic Respiration

by Igor Kučera and Vojtěch Sedláček

Microorganisms 2020, 8(8), 1230; https://doi.org/10.3390/microorganisms8081230 - 12 Aug 2020

Cited by 8 | Viewed by 2692

Abstract

Paracoccus denitrificans has a branched electron transport chain with three terminal oxidases transferring electrons to molecular oxygen, namely aa₃-type and cbb₃-type cytochrome c oxidases and ba₃-type ubiquinol oxidase. In the present study, we focused on strains expressing [...] Read more.

Paracoccus denitrificans has a branched electron transport chain with three terminal oxidases transferring electrons to molecular oxygen, namely aa₃-type and cbb₃-type cytochrome c oxidases and ba₃-type ubiquinol oxidase. In the present study, we focused on strains expressing only one of these enzymes. The competition experiments showed that possession of cbb₃-type oxidase confers significant fitness advantage during oxygen-limited growth and supports the biofilm lifestyle. The aa₃-type oxidase was shown to allow rapid aerobic growth at a high oxygen supply. Activity of the denitrification pathway that had been expressed in cells grown anaerobically with nitrate was fully inhibitable by oxygen only in wild-type and cbb₃ strains, while in strains aa₃ and ba₃ dinitrogen production from nitrate and oxygen consumption occurred simultaneously. Together, the results highlight the importance of the cbb₃-type oxidase for the denitrification phenotype and suggest a way of obtaining novel bacterial strains capable of aerobic denitrification. Full article

(This article belongs to the Section Microbial Biotechnology)

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17 pages, 5678 KiB

Open AccessArticle

Import of Entamoeba histolytica Mitosomal ATP Sulfurylase Relies on Internal Targeting Sequences

by Herbert J. Santos, Yoko Chiba, Takashi Makiuchi, Saki Arakawa, Yoshitaka Murakami, Kentaro Tomii, Kenichiro Imai and Tomoyoshi Nozaki

Microorganisms 2020, 8(8), 1229; https://doi.org/10.3390/microorganisms8081229 - 12 Aug 2020

Cited by 2 | Viewed by 3531

Abstract

Mitochondrial matrix proteins synthesized in the cytosol often contain amino (N)-terminal targeting sequences (NTSs), or alternately internal targeting sequences (ITSs), which enable them to be properly translocated to the organelle. Such sequences are also required for proteins targeted to mitochondrion-related organelles (MROs) that [...] Read more.

Mitochondrial matrix proteins synthesized in the cytosol often contain amino (N)-terminal targeting sequences (NTSs), or alternately internal targeting sequences (ITSs), which enable them to be properly translocated to the organelle. Such sequences are also required for proteins targeted to mitochondrion-related organelles (MROs) that are present in a few species of anaerobic eukaryotes. Similar to other MROs, the mitosomes of the human intestinal parasite Entamoeba histolytica are highly degenerate, because a majority of the components involved in various processes occurring in the canonical mitochondria are either missing or modified. As of yet, sulfate activation continues to be the only identified role of the relic mitochondria of Entamoeba. Mitosomes influence the parasitic nature of E. histolytica, as the downstream cytosolic products of sulfate activation have been reported to be essential in proliferation and encystation. Here, we investigated the position of the targeting sequence of one of the mitosomal matrix enzymes involved in the sulfate activation pathway, ATP sulfurylase (AS). We confirmed by immunofluorescence assay and subcellular fractionation that hemagluttinin (HA)-tagged EhAS was targeted to mitosomes. However, its ortholog in the δ-proteobacterium Desulfovibrio vulgaris, expressed as DvAS-HA in amoebic trophozoites, indicated cytosolic localization, suggesting a lack of recognizable mitosome targeting sequence in this protein. By expressing chimeric proteins containing swapped sequences between EhAS and DvAS in amoebic cells, we identified the ITSs responsible for mitosome targeting of EhAS. This observation is similar to other parasitic protozoans that harbor MROs, suggesting a convergent feature among various MROs in favoring ITS for the recognition and translocation of targeted proteins. Full article

(This article belongs to the Special Issue Virulence and Parasitism of Parasitic Protozoa)

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21 pages, 1485 KiB

Open AccessReview

COVID-19 Is a Multifaceted Challenging Pandemic Which Needs Urgent Public Health Interventions

by Carlo Contini, Elisabetta Caselli, Fernanda Martini, Martina Maritati, Elena Torreggiani, Silva Seraceni, Fortunato Vesce, Paolo Perri, Leonzio Rizzo and Mauro Tognon

Microorganisms 2020, 8(8), 1228; https://doi.org/10.3390/microorganisms8081228 - 12 Aug 2020

Cited by 33 | Viewed by 18448

Abstract

Until less than two decades ago, all known human coronaviruses (CoV) caused diseases so mild that they did not stimulate further advanced CoV research. In 2002 and following years, the scenario changed dramatically with the advent of the new more pathogenic CoVs, including [...] Read more.

Until less than two decades ago, all known human coronaviruses (CoV) caused diseases so mild that they did not stimulate further advanced CoV research. In 2002 and following years, the scenario changed dramatically with the advent of the new more pathogenic CoVs, including Severe Acute Respiratory Syndome (SARS-CoV-1), Middle Eastern respiratory syndrome (MERS)-CoV, and the new zoonotic SARS-CoV-2, likely originated from bat species and responsible for the present coronavirus disease (COVID-19), which to date has caused 15,581,007 confirmed cases and 635,173 deaths in 208 countries, including Italy. SARS-CoV-2 transmission is mainly airborne via droplets generated by symptomatic patients, and possibly asymptomatic individuals during incubation of the disease, although for the latter, there are no certain data yet. However, research on asymptomatic viral infection is currently ongoing worldwide to elucidate the real prevalence and mortality of the disease. From a clinical point of view, COVID-19 would be defined as “COVID Planet “ because it presents as a multifaceted disease, due to the large number of organs and tissues infected by the virus. Overall, based on the available published data, 80.9% of patients infected by SARS-CoV-2 develop a mild disease/infection, 13.8% severe pneumonia, 4.7% respiratory failure, septic shock, or multi-organ failure, and 3% of these cases are fatal, but mortality parameter is highly variable in different countries. Clinically, SARS-CoV-2 causes severe primary interstitial viral pneumonia and a “cytokine storm syndrome”, characterized by a severe and fatal uncontrolled systemic inflammatory response triggered by the activation of interleukin 6 (IL-6) with development of endothelitis and generalized thrombosis that can lead to organ failure and death. Risk factors include advanced age and comorbidities including hypertension, diabetes, and cardiovascular disease. Virus entry occurs via binding the angiotensin-converting enzyme 2 (ACE2) receptor present in almost all tissues and organs through the Spike (S) protein. Currently, SARS-CoV-2 infection is prevented by the use of masks, social distancing, and improved hand hygiene measures. This review summarizes the current knowledge on the main biological and clinical features of the SARS-CoV-2 pandemic, also focusing on the principal measures taken in some Italian regions to face the emergency and on the most important treatments used to manage the COVID-19 pandemic. Full article

(This article belongs to the Special Issue COVID-19: Focusing on Epidemiologic, Virologic, and Clinical Studies)

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17 pages, 2844 KiB

Open AccessArticle

Gene Expression and Photophysiological Changes in Pocillopora acuta Coral Holobiont Following Heat Stress and Recovery

by Rosa Celia Poquita-Du, Yi Le Goh, Danwei Huang, Loke Ming Chou and Peter A. Todd

Microorganisms 2020, 8(8), 1227; https://doi.org/10.3390/microorganisms8081227 - 12 Aug 2020

Cited by 17 | Viewed by 4652

Abstract

The ability of corals to withstand changes in their surroundings is a critical survival mechanism for coping with environmental stress. While many studies have examined responses of the coral holobiont to stressful conditions, its capacity to reverse responses and recover when the stressor [...] Read more.

The ability of corals to withstand changes in their surroundings is a critical survival mechanism for coping with environmental stress. While many studies have examined responses of the coral holobiont to stressful conditions, its capacity to reverse responses and recover when the stressor is removed is not well-understood. In this study, we investigated among-colony responses of Pocillopora acuta from two sites with differing distance to the mainland (Kusu (closer to the mainland) and Raffles Lighthouse (further from the mainland)) to heat stress through differential expression analysis of target genes and quantification of photophysiological metrics. We then examined how these attributes were regulated after the stressor was removed to assess the recovery potential of P. acuta. The fragments that were subjected to heat stress (2 °C above ambient levels) generally exhibited significant reduction in their endosymbiont densities, but the extent of recovery following stress removal varied depending on natal site and colony. There were minimal changes in chl a concentration and maximum quantum yield (Fv/Fm, the proportion of variable fluorescence (Fv) to maximum fluorescence (Fm)) in heat-stressed corals, suggesting that the algal endosymbionts’ Photosystem II was not severely compromised. Significant changes in gene expression levels of selected genes of interest (GOI) were observed following heat exposure and stress removal among sites and colonies, including Actin, calcium/calmodulin-dependent protein kinase type IV (Camk4), kinesin-like protein (KIF9), and small heat shock protein 16.1 (Hsp16.1). The most responsive GOIs were Actin, a major component of the cytoskeleton, and the adaptive immune-related Camk4 which both showed significant reduction following heat exposure and subsequent upregulation during the recovery phase. Our findings clearly demonstrate specific responses of P. acuta in both photophysiological attributes and gene expression levels, suggesting differential capacity of P. acuta corals to tolerate heat stress depending on the colony, so that certain colonies may be more resilient than others. Full article

(This article belongs to the Special Issue The Human Epoch: Cnidarians Holobiont Responses from Physiology to Epigenetic)

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11 pages, 565 KiB

Open AccessArticle

Diversity, Antibiotic Resistance, and Biofilm-Forming Ability of Enterobacteria Isolated from Red Meat and Poultry Preparations

by Rosa Capita, Ana Castaño-Arriba, Cristina Rodríguez-Melcón, Gilberto Igrejas, Patricia Poeta and Carlos Alonso-Calleja

Microorganisms 2020, 8(8), 1226; https://doi.org/10.3390/microorganisms8081226 - 12 Aug 2020

Cited by 10 | Viewed by 3755

Abstract

A total of 44 samples of beef, pork, and poultry preparations were tested. Average counts (log cfu/g) of enterobacteria were 1.99 ± 0.99 (beef preparations), 1.96 ± 1.44 (pork), 2.09 ± 0.92 (chicken), and 2.17 ± 1.06 (turkey) (p > 0.05). Two [...] Read more.

A total of 44 samples of beef, pork, and poultry preparations were tested. Average counts (log cfu/g) of enterobacteria were 1.99 ± 0.99 (beef preparations), 1.96 ± 1.44 (pork), 2.09 ± 0.92 (chicken), and 2.17 ± 1.06 (turkey) (p > 0.05). Two hundred enterobacterial strains were identified and 13 genera (21 species) were distinguished, including species that are a significant cause of infection. The most common genera were Escherichia (32.5% of strains), Serratia (17.0%), Hafnia (12.5%), and Salmonella (12.0%). Isolates were screened by disc diffusion for susceptibility to 15 antibiotics. A total of 126 strains (63% of the isolates) were multirresistant (having resistance to two or more antibiotics), 46 (23%) were resistant to one antibiotic, and 28 (14%) were sensitive to all antibiotics. The average number of resistances per strain was 2.53 ± 2.05. A higher (p < 0.05) average number of resistances was observed in strains from turkey (3.14 ± 2.55) than in strains from beef (2.15 ± 1.22), pork (2.16 ± 1.39), or chicken (2.44 ± 2.22). At least 50% of strains showed resistance or reduced susceptibility to ampicillin, cefotaxime, ceftazidime, or streptomycin, considered to be “critically important” antimicrobial agents in human medicine. Seventy-nine strains (39.5%), 60 strains (30.0%), and 46 strains (23.0%) were weak, moderate, and strong biofilm producers (crystal violet assay), respectively. This investigation provides evidence that bacteria from red meat and poultry preparations pose major potential risk to consumers. Full article

(This article belongs to the Section Food Microbiology)

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Open AccessArticle

Lactobacillus Mucosae Strain Promoted by a High-Fiber Diet in Genetic Obese Child Alleviates Lipid Metabolism and Modifies Gut Microbiota in ApoE^-/- Mice on a Western Diet

by Tianyi Jiang, Huan Wu, Xin Yang, Yue Li, Ziyi Zhang, Feng Chen, Liping Zhao and Chenhong Zhang

Microorganisms 2020, 8(8), 1225; https://doi.org/10.3390/microorganisms8081225 - 12 Aug 2020

Cited by 25 | Viewed by 4333

Abstract

Supplementation of probiotics is a promising gut microbiota-targeted therapeutic method for hyperlipidemia and atherosclerosis. However, the selection of probiotic candidate strains is still empirical. Here, we obtained a human-derived strain, Lactobacillus mucosae A1, which was shown by metagenomic analysis to be promoted by [...] Read more.

Supplementation of probiotics is a promising gut microbiota-targeted therapeutic method for hyperlipidemia and atherosclerosis. However, the selection of probiotic candidate strains is still empirical. Here, we obtained a human-derived strain, Lactobacillus mucosae A1, which was shown by metagenomic analysis to be promoted by a high-fiber diet and associated with the amelioration of host hyperlipidemia, and validated its effect on treating hyperlipidemia and atherosclerosis as well as changing structure of gut microbiota in ApoE^-/- mice on a Western diet. L. mucosae A1 attenuated the severe lipid accumulation in serum, liver and aortic sinus of ApoE^-/- mice on a Western diet, while it also reduced the serum lipopolysaccharide-binding protein content of mice, reflecting the improved metabolic endotoxemia. In addition, L. mucosae A1 shifted the gut microbiota structure of ApoE^-/- mice on a Western diet, including recovering a few members of gut microbiota enhanced by the Western diet. This study not only suggests the potential of L. mucosae A1 to be a probiotic in the treatment of hyperlipidemia and atherosclerosis, but also highlights the advantage of such function-based rather than taxonomy-based strategies for the selection of candidate strains for the next generation probiotics. Full article

(This article belongs to the Special Issue The Human Gut Microbiome, Diets and Health)

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