The Adaptation of Botrytis cinerea Extracellular Vesicles Proteome to Surrounding Conditions: Revealing New Tools for Its Infection Process.

Extracellular vesicles (EVs) are membranous particles released by different organisms. EVs carry several sets of macromolecules implicated in cell communication. EVs have become a relevant topic in the study of pathogenic fungi due to their relationship with fungal-host interactions. One of the essential research areas in this field is the characterization protein profile of EVs since plant fungal pathogens rely heavily on secreted proteins to invade their hosts. However, EVs of Botrytis cinerea are little known, which is one of the most devastating phytopathogenic fungi. The present study has two main objectives: the characterization of B. cinerea EVs proteome changes under two pathogenic conditions and the description of their potential role during the infective process. All the experimental procedure was conducted in B. cinerea growing in a minimal salt medium supplemented with glucose as a constitutive stage and deproteinized tomato cell walls (TCW) as a virulence inductor. The isolation of EVs was performed by differential centrifugation, filtration, ultrafiltration, and sucrose cushion ultracentrifugation. EVs fractions were visualised by TEM using negative staining. Proteomic analysis of EVs cargo was addressed by LC-MS/MS. The methodology used allowed the correct isolation of B. cinerea EVs and the identification of a high number of EV proteins, including potential EV markers. The isolated EVs displayed differences in morphology under both assayed conditions. GO analysis of EV proteins showed enrichment in cell wall metabolism and proteolysis under TCW. KEGG analysis also showed the difference in EVs function under both conditions, highlighting the presence of potential virulence/pathogenic factors implicated in cell wall metabolism, among others. This work describes the first evidence of EVs protein cargo adaptation in B. cinerea, which seems to play an essential role in its infection process, sharing crucial functions with the conventional secretion pathways.

Selective killing of the human gastric pathogen Helicobacter pylori by mitochondrial respiratory complex I inhibitors.

Respiratory complex I is a multicomponent enzyme conserved between eukaryotic cells and many bacteria, which couples oxidation of electron donors and quinone reduction with proton pumping. Here, we report that protein transport via the Cag type IV secretion system, a major virulence factor of the Gram-negative bacterial pathogen Helicobacter pylori, is efficiently impeded by respiratory inhibition. Mitochondrial complex I inhibitors, including well-established insecticidal compounds, selectively kill H. pylori, while other Gram-negative or Gram-positive bacteria, such as the close relative Campylobacter jejuni or representative gut microbiota species, are not affected. Using a combination of different phenotypic assays, selection of resistance-inducing mutations, and molecular modeling approaches, we demonstrate that the unique composition of the H. pylori complex I quinone-binding pocket is the basis for this hypersensitivity. Comprehensive targeted mutagenesis and compound optimization studies highlight the potential to develop complex I inhibitors as narrow-spectrum antimicrobial agents against this pathogen.

Intraspecific Comparative Analysis Reveals Genomic Variation of Didymella arachidicola and Pathogenicity Factors Potentially Related to Lesion Phenotype.

Didymella arachidicola is one of the most important fungal pathogens, causing foliar disease and leading to severe yield losses of peanuts (Arachis hypogaea L.) in China. Two main lesion phenotypes of peanut web blotch have been identified as reticulation type (R type) and blotch type (B type). As no satisfactory reference genome is available, the genomic variations and pathogenicity factors of D. arachidicola remain to be revealed. In the present study, we collected 41 D. arachidicola isolates from 26 geographic locations across China (33 for R type and 8 for B type). The chromosome-scale genome of the most virulent isolate (YY187) was assembled as a reference using PacBio and Hi-C technologies. In addition, we re-sequenced 40 isolates from different sampling sites. Genome-wide alignments showed high similarity among the genomic sequences from the 40 isolates, with an average mapping rate of 97.38%. An average of 3242 SNPs and 315 InDels were identified in the genomic variation analysis, which revealed an intraspecific polymorphism in D. arachidicola. The comparative analysis of the most and least virulent isolates generated an integrated gene set containing 512 differential genes. Moreover, 225 genes individually or simultaneously harbored hits in CAZy-base, PHI-base, DFVF, etc. Compared with the R type reference, the differential gene sets from all B type isolates identified 13 shared genes potentially related to lesion phenotype. Our results reveal the intraspecific genomic variation of D. arachidicola isolates and pathogenicity factors potentially related to different lesion phenotypes. This work sets a genomic foundation for understanding the mechanisms behind genomic diversity driving different pathogenic phenotypes of D. arachidicola.

Identification of a Novel Lipase with AHSMG Pentapeptide in Hypocreales and Glomerellales Filamentous Fungi.

Lipases are enzymes that hydrolyze triglycerides to fatty acids and glycerol. A typical element in lipases is a conserved motif of five amino acids (the pentapeptide), most commonly G-X-S-X-G. Lipases with the pentapeptide A-X-S-X-G are present in species of Bacillus, Paucimonas lemoignei, and the yeast Trichosporon asahii; they are usually thermotolerant and solvent resistant. Recently, while searching for true lipases in the Trichoderma harzianum genome, one lipase containing the pentapeptide AHSMG was identified. In this study, we cloned from T. harzianum strain B13-1 the lipase ID135964, renamed here as ThaL, which is 97.65% identical with the reference. We found that ThaL is a lid-containing true lipase of cluster III that belongs to a large family comprising highly conserved proteins in filamentous fungi in the orders Hypocreales and Glomerellales, in which predominantly pathogenic fungi are found. ThaL was expressed in conidia, as well as in T. harzianum mycelium, where it was cultured in liquid minimal medium. These results-together with the amino acid composition, absence of a signal peptide, mitochondrial sorting prediction, disordered regions in the protein, and lineage-specific phylogenetic distribution of its homologs-suggest that ThaL is a non-canonical effector. In summary, AHSMG-lipase is a novel lipase family in filamentous fungi, and is probably involved in pathogenicity.

Pathogenicity Factors of Botryosphaeriaceae Associated with Grapevine Trunk Diseases: New Developments on Their Action on Grapevine Defense Responses.

Botryosphaeriaceae are a family of fungi associated with the decay of a large number of woody plants with economic importance and causing particularly great losses in viticulture due to grapevine trunk diseases. In recent years, major advances in the knowledge of the pathogenicity factors of these pathogens have been made possible by the development of next-generation sequencing. This review highlights the knowledge gained on genes encoding small secreted proteins such as effectors, carbohydrate-associated enzymes, transporters and genes associated with secondary metabolism, their representativeness within the Botryosphaeriaceae family and their expression during grapevine infection. These pathogenicity factors are particularly expressed during host-pathogen interactions, facilitating fungal development and nutrition, wood colonization, as well as manipulating defense pathways and inducing impacts at the cellular level and phytotoxicity. This work highlights the need for further research to continue the effort to elucidate the pathogenicity mechanisms of this family of fungi infecting grapevine in order to improve the development of control methods and varietal resistance and to reduce the development and the effects of the disease on grapevine harvest quality and yield.

Horizontal Transfer of Virulence Factors by Pathogenic Enterobacteria to Marine Saprotrophic Bacteria during Co-Cultivation in Biofilm.

Environmental problems associated with marine pollution and climate warming create favorable conditions for the penetration and survival of pathogenic bacteria in marine ecosystems. These microorganisms have interspecific competitive interactions with marine bacteria. Co-culture, as an important research strategy that mimics the natural environment of bacteria, can activate silent genes or clusters through interspecies interactions. The authors used modern biotechnology of co-cultivation to dynamically study intercellular interactions between different taxa of bacteria-pathogenic enterobacteria Yersinia pseudotuberculosis and Listeria monocytogenes and saprotrophic marine bacteria Bacillus sp. and Pseudomonas japonica isolated in summer from the coastal waters of the recreational areas of the Sea of Japan. The results of the experiments showed that during the formation of polycultural biofilms, horizontal transfer of genes encoding some pathogenicity factors from Y. pseudotuberculosis and L. monocytogenes to marine saprotrophic bacteria with different secretion systems is possible. It was previously thought that this was largely prevented by the type VI secretion system (T6SS) found in marine saprotrophic bacteria. The authors showed for the first time the ability of marine bacteria Bacillus sp. and P. japonica to biofilm formation with pathogenic enterobacteria Y. pseudotuberculosis and L. monocytogenes, saprophytic bacteria with type III secretion system (T3SS). For the first time, a marine saprotrophic strain of Bacillus sp. Revealed manifestations of hyaluronidase, proteolytic and hemolytic activity after cultivation in a polycultural biofilm with listeria. Saprotrophic marine bacteria that have acquired virulence factors from pathogenic enterobacteria, including antibiotic resistance genes, could potentially play a role in altering the biological properties of other members of the marine microbial community. In addition, given the possible interdomain nature of intercellular gene translocation, acquired virulence factors can be transferred to marine unicellular and multicellular eukaryotes. The results obtained contribute to the paradigm of the epidemiological significance and potential danger of anthropogenic pollution of marine ecosystems, which creates serious problems for public health and the development of marine culture as an important area of economic activity in coastal regions.

Comparative genomics of plant pathogenic Diaporthe species and transcriptomics of Diaporthe caulivora during host infection reveal insights into pathogenic strategies of the genus.

BACKGROUND: Diaporthe caulivora is a fungal pathogen causing stem canker in soybean worldwide. The generation of genomic and transcriptomic information of this ascomycete, together with a comparative genomic approach with other pathogens of this genus, will contribute to get insights into the molecular basis of pathogenicity strategies used by D. caulivora and other Diaporthe species. RESULTS: In the present work, the nuclear genome of D. caulivora isolate (D57) was resolved, and a comprehensive annotation based on gene expression and genomic analysis is provided. Diaporthe caulivora D57 has an estimated size of 57,86 Mb and contains 18,385 predicted protein-coding genes, from which 1501 encode predicted secreted proteins. A large array of D. caulivora genes encoding secreted pathogenicity-related proteins was identified, including carbohydrate-active enzymes (CAZymes), necrosis-inducing proteins, oxidoreductases, proteases and effector candidates. Comparative genomics with other plant pathogenic Diaporthe species revealed a core secretome present in all Diaporthe species as well as Diaporthe-specific and D. caulivora-specific secreted proteins. Transcriptional profiling during early soybean infection stages showed differential expression of 2659 D. caulivora genes. Expression patterns of upregulated genes and gene ontology enrichment analysis revealed that host infection strategies depends on plant cell wall degradation and modification, detoxification of compounds, transporter activities and toxin production. Increased expression of effectors candidates suggests that D. caulivora pathogenicity also rely on plant defense evasion. A high proportion of the upregulated genes correspond to the core secretome and are represented in the pathogen-host interaction (PHI) database, which is consistent with their potential roles in pathogenic strategies of the genus Diaporthe. CONCLUSIONS: Our findings give novel and relevant insights into the molecular traits involved in pathogenicity of D. caulivora towards soybean plants. Some of these traits are in common with other Diaporthe pathogens with different host specificity, while others are species-specific. Our analyses also highlight the importance to have a deeper understanding of pathogenicity functions among Diaporthe pathogens and their interference with plant defense activation.

Sequence Variability of pXO1-Located Pathogenicity Genes of Bacillus anthracis Natural Strains of Different Geographic Origin.

The main pathogenic factor of Bacillus anthracis is a three-component toxin encoded by the pagA, lef, and cya genes, which are located on the pXO1 plasmid. The atxA gene, which encodes the primary regulator of pathogenicity factor expression, is located on the same plasmid. In this work, we evaluated the polymorphism of the pagA, lef, cya, and atxA genes for 85 B. anthracis strains from different evolutionary lineages and canSNP groups. We have found a strong correlation of 19 genotypes with the main evolutionary lineages, but the correlation with the canSNP group of the strain was not as strong. We have detected several genetic markers indicating the geographical origin of the strains, for example, their source from the steppe zone of the former USSR. We also found that strains of the B.Br.001/002 group caused an anthrax epidemic in Russia in 2016 and strains isolated during paleontological excavations in the Russian Arctic have the same genotype as the strains of the B.Br.CNEVA group circulating in Central Europe. This data could testify in favor of the genetic relationship of these two groups of strains and hypothesize the ways of distribution of their ancestral forms between Europe and the Arctic.

Identification and Comparison of Colletotrichum Secreted Effector Candidates Reveal Two Independent Lineages Pathogenic to Soybean.

Colletotrichum is one of the most important plant pathogenic genus of fungi due to its scientific and economic impact. A wide range of hosts can be infected by Colletotrichum spp., which causes losses in crops of major importance worldwide, such as soybean. Soybean anthracnose is mainly caused by C. truncatum, but other species have been identified at an increasing rate during the last decade, becoming one of the most important limiting factors to soybean production in several regions. To gain a better understanding of the evolutionary origin of soybean anthracnose, we compared the repertoire of effector candidates of four Colletotrichum species pathogenic to soybean and eight species not pathogenic. Our results show that the four species infecting soybean belong to two lineages and do not share any effector candidates. These results strongly suggest that two Colletotrichum lineages have acquired the capability to infect soybean independently. This study also provides, for each lineage, a set of candidate effectors encoding genes that may have important roles in pathogenicity towards soybean offering a new resource useful for further research on soybean anthracnose management.

Comparative transcriptomic analysis of races 1, 2, 5 and 6 of Fusarium oxysporum f.sp. pisi in a susceptible pea host identifies differential pathogenicity profiles.

BACKGROUND: The fungal pathogen Fusarium oxysporum f.sp. pisi (Fop) causes Fusarium wilt in peas. There are four races globally: 1, 2, 5 and 6 and all of these races are present in Australia. Molecular infection mechanisms have been studied in a few other F. oxysporum formae speciales; however, there has been no transcriptomic Fop-pea pathosystem study. RESULTS: A transcriptomic study was carried out to understand the molecular pathogenicity differences between the races. Transcriptome analysis at 20 days post-inoculation revealed differences in the differentially expressed genes (DEGs) in the Fop races potentially involved in fungal pathogenicity variations. Most of the DEGs in all the races were engaged in transportation, metabolism, oxidation-reduction, translation, biosynthetic processes, signal transduction, proteolysis, among others. Race 5 expressed the most virulence-associated genes. Most genes encoding for plant cell wall degrading enzymes, CAZymes and effector-like proteins were expressed in race 2. Race 6 expressed the least number of genes at this time point. CONCLUSION: Fop races deploy various factors and complex strategies to mitigate host defences to facilitate colonisation. This investigation provides an overview of the putative pathogenicity genes in different Fop races during the necrotrophic stage of infection. These genes need to be functionally characterised to confirm their pathogenicity/virulence roles and the race-specific genes can be further explored for molecular characterisation.

Genome-wide analysis of Claviceps paspali: insights into the secretome of the main species causing ergot disease in Paspalum spp.

BACKGROUND: The phytopatogen Claviceps paspali is the causal agent of Ergot disease in Paspalum spp., which includes highly productive forage grasses such as P. dilatatum. This disease impacts dairy and beef production by affecting seed quality and producing mycotoxins that can affect performance in feeding animals. The molecular basis of pathogenicity of C. paspali remains unknown, which makes it more difficult to find solutions for this problem. Secreted proteins are related to fungi virulence and can manipulate plant immunity acting on different subcellular localizations. Therefore, identifying and characterizing secreted proteins in phytopathogenic fungi will provide a better understanding of how they overcome host defense and cause disease. The aim of this work is to analyze the whole genome sequences of three C. paspali isolates to obtain a comparative genome characterization based on possible secreted proteins and pathogenicity factors present in their genome. In planta RNA-seq analysis at an early stage of the interaction of C. paspali with P. dilatatum stigmas was also conducted in order to determine possible secreted proteins expressed in the infection process. RESULTS: C. paspali isolates had compact genomes and secretome which accounted for 4.6-4.9% of the predicted proteomes. More than 50% of the predicted secretome had no homology to known proteins. RNA-Seq revealed that three protein-coding genes predicted as secreted have mayor expression changes during 1 dpi vs 4 dpi. Also, three of the first 10 highly expressed genes in both time points were predicted as effector-like. CAZyme-like proteins were found in the predicted secretome and the most abundant family could be associated to pectine degradation. Based on this, pectine could be a main component affected by the cell wall degrading enzymes of C. paspali. CONCLUSIONS: Based on predictions from DNA sequence and RNA-seq, unique probable secreted proteins and probable pathogenicity factors were identified in C. paspali isolates. This information opens new avenues in the study of the biology of this fungus and how it modulates the interaction with its host. Knowledge of the diversity of the secretome and putative pathogenicity genes should facilitate future research in disease management of Claviceps spp.

Responses to Ecopollutants and Pathogenization Risks of Saprotrophic Rhodococcus Species.

Under conditions of increasing environmental pollution, true saprophytes are capable of changing their survival strategies and demonstrating certain pathogenicity factors. Actinobacteria of the genus Rhodococcus, typical soil and aquatic biotope inhabitants, are characterized by high ecological plasticity and a wide range of oxidized organic substrates, including hydrocarbons and their derivatives. Their cell adaptations, such as the ability of adhering and colonizing surfaces, a complex life cycle, formation of resting cells and capsule-like structures, diauxotrophy, and a rigid cell wall, developed against the negative effects of anthropogenic pollutants are discussed and the risks of possible pathogenization of free-living saprotrophic Rhodococcus species are proposed. Due to universal adaptation features, Rhodococcus species are among the candidates, if further anthropogenic pressure increases, to move into the group of potentially pathogenic organisms with "unprofessional" parasitism, and to join an expanding list of infectious agents as facultative or occasional parasites.

Substitution Arg140Gly in Hemagglutinin Reduced the Virulence of Highly Pathogenic Avian Influenza Virus H7N1.

The H7 subtype of avian influenza viruses (AIV) stands out among other AIV. The H7 viruses circulate in ducks, poultry and equines and have repeatedly caused outbreaks of disease in humans. The laboratory strain A/chicken/Rostock/R0p/1934 (H7N1) (R0p), which was previously derived from the highly pathogenic strain A/FPV/Rostock/1934 (H7N1), was studied in this work to ascertain its biological property, genome stability and virulent changing mechanism. Several virus variants were obtained by serial passages in the chicken lungs. After 10 passages of this virus through the chicken lungs we obtained a much more pathogenic variant than the starting R0p. The study of intermediate passages showed a sharp increase in pathogenicity between the fifth and sixth passage. By cloning these variants, a pair of strains (R5p and R6p) was obtained, and the complete genomes of these strains were sequenced. Single amino acid substitution was revealed, namely reversion Gly140Arg in HA1. This amino acid is located at the head part of the hemagglutinin, adjacent to the receptor-binding site. In addition to the increased pathogenicity in chicken and mice, R6p differs from R5p in the shape of foci in cell culture and an increased affinity for a negatively charged receptor analogue, while maintaining a pattern of receptor-binding specificity and the pH of conformational change of HA.

Genetic response to nitrogen starvation in the aggressive Eucalyptus foliar pathogen Teratosphaeria destructans.

Teratosphaeria destructans is one of the most aggressive foliar pathogens of Eucalyptus. The biological factors underpinning T. destructans infections, which include shoot and leaf blight on young trees, have never been interrogated. Thus, the means by which the pathogen modifies its host environment to overcome host defences remain unknown. By applying transcriptome sequencing, the aim of this study was to compare gene expression in a South African isolate of T. destructans grown on nitrogen-deficient and complete media. This made it possible to identify upregulated genes in a nitrogen-starved environment, often linked to the pathogenicity of the fungus. The results support the hypothesis that nitrogen starvation in T. destructans likely mirrors an in planta genetic response. This is because 45% of genes that were highly upregulated under nitrogen starvation have previously been reported to be associated with infection in other pathogen systems. These included several CAZymes, fungal effector proteins, peptidases, kinases, toxins, lipases and proteins associated with detoxification of toxic compounds. Twenty-five secondary metabolites were identified and expressed in both nitrogen-deficient and complete conditions. Additionally, the most highly expressed genes in both growth conditions had pathogenicity-related functions. This study highlights the large number of expressed genes associated with pathogenicity and overcoming plant defences. As such, the generated baseline knowledge regarding pathogenicity and aggressiveness in T. destructans is a valuable reference for future in planta work.

Corynebacterium spp. - problematic pathogens of the human respiratory tract (review of literature).

Corynebacterium spp. - representatives of the normal microflora of the human body, but their role in the development of diseases in both immunocompromised and immunocompetent patients is known. Corynebacterim spp. (C. pseudodiphtheriticum, C. striatum, C. amycolatum, C. accolens, C. argentoratense, etc.) is associated with diseases of the respiratory tract: tracheitis, pharyngitis, rhinosinusitis, bronchitis, etc. They can be transmitted by airborne droplets, household contact, and possibly by hematogenic pathways. Corynebacterim spp. toxins do not produce, but are capable of adhesion and invasion, biofilm formation, production of neuraminidase, hyaluronidase, and hemolysin. It is necessary to take into account not so much the species, but the strain affiliation of isolates of Corynebacterium spp., since among the representatives of one species of non-diphtheria corynebacteria (for example, C. pseudodiphtheriticum), colonizing the respiratory tract, there may be strains that can exhibit not only pathogenic properties, but also probiotic activity. Microbiological diagnostics is based on their quantitative determination in biological material, phenotypic (culture study, test systems for biochemical identification, Vitek 2 automated systems) and genotypic (16SpRNA gene sequencing and rpoB) methods. It is possible to use mass spectrometric analysis (MALDI-ToF-MS). The greatest activity against Corynebacterium spp. in vitro studies preserve vancomycin, teicoplanin, and linezolid. Successful therapy with at least two of the following antimicrobial agents (AMP) has been reported: vancomycin, rifampicin, linezolid, and daptomycin. The sensitivity of isolates of Corynebacterium spp. to AMP is not related to the species, but is due to strain differences, and therefore it is necessary to test each isolated strain. Continuous monitoring of the sensitivity of Corynebacterium spp. strains to AMP is necessary due to the observed variability of these traits. Of particular importance is the identification of multidrug-resistant isolates that are currently considered highly pathogenic. When compiling the review, the databases Scopus, Web of Science, The Cochrane Library, CyberLeninka, RSCI were used.

Comparative proteomic analysis reveals molecular differences between incompatible and compatible interaction of Erysiphe pisi in garden pea.

Comparative proteome analysis of Erysiphe pisi-infected pea genotypes; JI-2480 carrying er2 resistant gene and Arkel, the susceptible genotype by liquid chromatography- mass spectrometry (LCMS/MS QTOF) at 72 h post inoculation (hpi) revealed several differentially abundant proteins (DAPs) of both the host and the pathogen. The functional annotation of proteins through gene enrichment and KEGG pathway analyses revealed strong up-regulation of pathogenesis related protein NPR1, proteins related to defense, transportation and signal transduction, hypersensitive response, cell wall modifications, phenylpropanoid and metabolic pathways in J-72. Significant abundance of membrane-related polypeptides, kinase domains and small GTPase signal transduction-related proteins suggested their major role in plant defense. The abundance of cellular antioxidant protein, catalase and its isozyme along with calreticulin-1 and 2 in J-72 confirmed their intervention in maintaining a redox balance in powdery mildew defense. High abundance levels of Glycolysis-related proteins indicated it as a major pathway for energy source during fungal growth. The majority of pathogenicity and virulence genes were downregulated in J-72 compared to A-72, while four EKA (Effectors homologues to Avk1 and Avra10) like avirulence proteins were significantly upregulated in incompatible interaction suggesting their role in eliciting hypersensitive response in pea against E. pisi.

Comparative transcriptome of compatible and incompatible interaction of Erysiphe pisi and garden pea reveals putative defense and pathogenicity factors.

Comparative transcriptome analysis of Erysiphe pisi-infected pea (Pisum sativum) genotypes JI-2480 (resistant) and Arkel (susceptible) at 72 hours post-inoculation (hpi) was carried to detect molecular components involved in compatible and incompatible interactions. Differential gene expression was observed in Arkel and JI-2480 genotype at 72 hpi with E. pisi isolate (Ep01) using EdgeR software. Out of 32 217 transcripts, 2755 transcripts showed significantly altered gene expression in case of plants while 530 were related to E. pisi (P < 0.05). The higher transcript number of differentially expressed genes demonstrated peak activity of pathogenicity genes in plants at 72 hpi. Glycolysis was observed to be the major pathway for energy source during fungal growth. Differential gene expression of plant transcripts revealed significant expression of putative receptor and regulatory sequences involved in defense in the resistant, JI-2480 compared to susceptible, Arkel genotype. Expression of genes involved in defense and hormonal signaling, genes related to hypersensitive response, reactive oxygen species and phenylpropanoid pathway in JI-2480 indicated their crucial role in disease resistance against E. pisi. Down-regulation of transcription factors like-WRKY-28 and up-regulation of several putative pattern recognition receptors in JI-2480 compared to Arkel also suggested activation of host-mediated defense responses against E. pisi in pea.

Impact of a reduced water salinity on the composition of Vibrio spp. in recirculating aquaculture systems for Pacific white shrimp (Litopenaeus vannamei) and its possible risks for shrimp health and food safety.

Tropical shrimp, like Litopenaeus vannamei, in land-based recirculating aquaculture systems (RAS) are often kept at low water salinities to reduce costs for artificial sea salt and the amount of salty wastewater. Although these shrimp are tolerant against low salinities, innate immunity suppression and changes in the microbial composition in the water can occur. As especially Vibrio spp. are relevant for shrimp health, alterations in the species composition of the Vibrio community were analysed in water from six RAS, run at 15‰ or 30‰. Additionally, pathogenicity factors including pirA/B, VPI, toxR, toxS, vhh, vfh, tdh, trh, flagellin genes and T6SS1/2 of V. parahaemolyticus were analysed. The Vibrio composition differed significantly depending on water salinity. In RAS at 15‰, higher numbers of the potentially pathogenic species V. parahaemolyticus, V. owensii and V. campbellii were detected, and especially in V. parahaemolyticus, various pathogenicity factors were present. A reduced salinity may therefore pose a higher risk of disease outbreaks in shrimp RAS. Because some of the detected pathogenicity factors are relevant for human health, this might also affect food safety. In order to produce healthy shrimp as a safe food for human consumption, maintaining high water salinities seems to be recommendable.

Identifying candidate Aspergillus pathogenicity factors by annotation frequency.

BACKGROUND: Members of the genus Aspergillus display a variety of lifestyles, ranging from saprobic to pathogenic on plants and/or animals. Increased genome sequencing of economically important members of the genus permits effective use of "-omics" comparisons between closely related species and strains to identify candidate genes that may contribute to phenotypes of interest, especially relating to pathogenicity. Protein-coding genes were predicted from 216 genomes of 12 Aspergillus species, and the frequencies of various structural aspects (exon count and length, intron count and length, GC content, and codon usage) and functional annotations (InterPro, Gene Ontology, and Kyoto Encyclopedia of Genes and Genomes terms) were compared. RESULTS: Using principal component analyses, the three sets of functional annotations for each strain were clustered by species. The species clusters appeared to separate by pathogenicity on plants along the first dimensions, which accounted for over 20% of the variance. More annotations for genes encoding pectinases and secondary metabolite biosynthetic enzymes were assigned to phytopathogenic strains from species such as Aspergillus flavus. In contrast, Aspergillus fumigatus strains, which are pathogenic to animals but not plants, were assigned relatively more terms related to phosphate transferases, and carbohydrate and amino-sugar metabolism. Analyses of publicly available RNA-Seq data indicated that one A. fumigatus protein among 17 amino-sugar processing candidates, a hexokinase, was up-regulated during co-culturing with human immune system cells. CONCLUSION: Genes encoding hexokinases and other proteins of interest may be subject to future manipulations to further refine understanding of Aspergillus pathogenicity factors.

Deciphering the Infectious Process of Colletotrichum lupini in Lupin through Transcriptomic and Proteomic Analysis.

The fungal phytopathogen Colletotrichum lupini is responsible for lupin anthracnose, resulting in significant yield losses worldwide. The molecular mechanisms underlying this infectious process are yet to be elucidated. This study proposes to evaluate C. lupini gene expression and protein synthesis during lupin infection, using, respectively, an RNAseq-based transcriptomic approach and a mass spectrometry-based proteomic approach. Patterns of differentially-expressed genes in planta were evaluated from 24 to 84 hours post-inoculation, and compared to in vitro cultures. A total of 897 differentially-expressed genes were identified from C. lupini during interaction with white lupin, of which 520 genes were predicted to have a putative function, including carbohydrate active enzyme, effector, protease or transporter-encoding genes, commonly described as pathogenicity factors for other Colletotrichum species during plant infection, and 377 hypothetical proteins. Simultaneously, a total of 304 proteins produced during the interaction were identified and quantified by mass spectrometry. Taken together, the results highlight that the dynamics of symptoms, gene expression and protein synthesis shared similarities to those of hemibiotrophic pathogens. In addition, a few genes with unknown or poorly-described functions were found to be specifically associated with the early or late stages of infection, suggesting that they may be of importance for pathogenicity. This study, conducted for the first time on a species belonging to the Colletotrichum acutatum species complex, presents an opportunity to deepen functional analyses of the genes involved in the pathogenicity of Colletotrichum spp. during the onset of plant infection.