Potential Virulence Factors Research Articles

Heterogeneity and Genomic Plasticity of Acinetobacter baumannii and Acinetobacter nosocomialis Isolates Recovered from Clinical Samples in India.

Acinetobacter baumannii and Acinetobacter nosocomialis are the imperious pathogens in the intensive care units. We aimed to explore the genomic features of these pathogens to understand the factors influencing their plasticity. Using next-generation sequencing, two carbapenem-resistant A. baumannii (AbaBS-3, AbaETR-4) isolates and a pan-susceptible A. nosocomialis (AbaAS-5) isolate were characterised. All genomes exhibited 94% similarity with a degree of heterogeneity. AbaBS-3 and AbaETR-4 harboured antibiotic resistance gene (ARG) repertoire to most antibiotic classes. Carbapenem resistance was due to blaOXA-23 and blaOXA-66 besides the antibiotic efflux pumps. Diverse mobile genetic elements (MGE), insertion sequences (IS), prophages and virulence determinants with a plethora of stress response genes were identified in all three genomes. Class-1 integron in AbaETR-4, encoded genes that confer resistance to aminoglycosides, phenicol, sulfonamides and disinfectants. Substitutions in LpxACD and PmrCAB of AbaETR-4 confirmed the colistin resistance in vitro. Novel mutations in piuA, responsible for transporting cefiderocol, were found in AbaBS-3 and AbaETR-4. Plasmids carrying toxin-antitoxin systems, ARGs and ISs were present in these genomes. All three genomes harboured diverse protein secretion systems, virulence determinants related to immune evasion, adherence, biofilm formation and iron acquisition systems. AbaAS-5 exclusively harboured serine protease pkf, and CpaA substrate of type-II secretion system but lacked the acinetobactin-iron acquisition system. Our work delivers a holistic genome characterization of A. baumannii, coupled with a trailblazing attempt to study A. nosocomialis from India. The presence of ARGs and potential virulence factors interspersed with MGE is a cause for concern, depicting the dynamic adaptability mediated by genetic recombination.

VmSpm1: a secretory protein from Valsa mali that targets apple's abscisic acid receptor MdPYL4 to suppress jasmonic acid signaling and enhance infection.

Pathogenic fungi such as Valsa mali secrete effector proteins to manipulate host defenses and facilitate infection. Subtilases are identified as potential virulence factors, yet their specific roles in fruit tree pathogens, such as those affecting apple trees, are poorly understood. Our research shows VmSpm1 as a virulence factor in V. mali. Knocking it out decreased virulence, whereas its heterologous expression in apple led to reduced disease resistance. Using Y2H, BiFC, SLC, and Co-IP techniques, we demonstrated an interaction between VmSpm1 and MdPYL4. MdPYL4 levels increased during V. mali infection. The stable transgenic apple lines inoculation experiment showed that MdPYL4 correlates with enhanced resistance to Apple Valsa canker when overexpressed in apples. Furthermore, through in vitro and in vivo assays, we showed the degradative role of VmSpm1 on MdPYL4. MdPYL4 promotes the synthesis of jasmonic acid (JA) in apples in an abscisic acid-dependent manner. The degradation of MdPYL4 leads to a reduction in JA content in apples during V. mali infection, thereby impairing JA signal transduction and decreasing disease resistance in apple plants. In summary, this study reveals how V. mali utilizes VmSpm1 to subvert JA signaling, shedding light on fungal manipulation of plant hormones to disrupt immunity.

Screening and in silico characterization of prophages in Helicobacter pylori clinical strains

The increase of antibiotic resistance calls for alternatives to control Helicobacter pylori, a Gram-negative bacterium associated with various gastric diseases. Bacteriophages (phages) can be highly effective in the treatment of pathogenic bacteria. Here, we develop a method to identify prophages in H. pylori genomes aiming at a future use in therapy. A PCR-based technique tested five primer pairs on 74 clinical H. pylori strains. With the PCR screening, 14 strains most likely to carry prophages were fully sequenced. After that, a more holistic approach was taken by studying the complete genome of the strains. The work allowed us to identify 12 intact prophage sequences, which were then characterized concerning their morphology, virulence, and antibiotic-resistance genes. To understand the variability of prophages, a phylogenetic analysis using the sequences of all H. pylori phages reported to date was performed. Using a PCR-based technique we increased the efficiency of identifying complete prophages to 54.1 %. Genes with homology to potential virulence factors were identified in some new prophages. Phylogenetic analysis revealed a close relationship among H. pylori-phages, although there are phages with different geographical origins. This study provides a deeper understanding of H. pylori-phages, providing valuable insights into their potential use in therapy.

Whole genome sequencing and characterization of Corynebacterium isolated from the healthy and dry eye ocular surface

BackgroundThe purpose of this study was to characterize Corynebacterium isolated from the ocular surface of dry eye disease patients and healthy controls. We aimed to investigate the pathogenic potential of these isolates in relation to ocular surface health. To this end, we performed whole genome sequencing in combination with biochemical, enzymatic, and antibiotic susceptibility tests. In addition, we employed deferred growth inhibition assays to examine how Corynebacterium isolates may impact the growth of potentially competing microorganisms including the ocular pathogens Pseudomonas aeruginosa and Staphylococcus aureus, as well as other Corynebacterium present on the eye.ResultsThe 23 isolates were found to belong to 8 different species of Corynebacterium with genomes ranging from 2.12 mega base pairs in a novel Corynebacterium sp. to 2.65 mega base pairs in C. bovis. Whole genome sequencing revealed the presence of a range of antimicrobial targets present in all isolates. Pangenome analysis showed the presence of 516 core genes and that the pangenome is open. Phenotypic characterization showed variously urease, lipase, mucinase, protease and DNase activity in some isolates. Attention was particularly drawn to a potentially new or novel Corynebacterium species which had the smallest genome, and which produced a range of hydrolytic enzymes. Strikingly the isolate inhibited in vitro the growth of a range of possible pathogenic bacteria as well as other Corynebacterium isolates. The majority of Corynebacterium species included in this study did not seem to possess canonical pathogenic activity.ConclusionsThis study is the first reported genomic and biochemical characterization of ocular Corynebacterium. A number of potential virulence factors were identified which may have direct relevance for ocular health and contribute to the finding of our previous report on the ocular microbiome, where it was shown that DNA libraries were often dominated by members of this genus. Particularly interesting in this regard was the observation that some Corynebacterium, particularly new or novel Corynebacterium sp. can inhibit the growth of other ocular Corynebacterium as well as known pathogens of the eye.

Microbial extracellular polymeric substances in the environment, technology and medicine.

Microbial biofilms exhibit a self-produced matrix of extracellular polymeric substances (EPS), including polysaccharides, proteins, extracellular DNA and lipids. EPS promote interactions of the biofilm with other cells and sorption of organics, metals and chemical pollutants, and they facilitate cell adhesion at interfaces and ensure matrix cohesion. EPS have roles in various natural environments, such as soils, sediments and marine habitats. In addition, EPS are relevant in technical environments, such as wastewater and drinking water treatment facilities, and water distribution systems, and they contribute to biofouling and microbially influenced corrosion. In medicine, EPS protect pathogens within the biofilm against the host immune system and antimicrobials, and emerging evidence suggests that EPS can represent potential virulence factors. By contrast, EPS yield a wide range of valuable products that include their role in self-repairing concrete. In this Review, we aim to explore EPS as a functional unit of biofilms in the environment, in technology and in medicine.

Characterization Studies on the sugC Gene of Streptococcus suis Serotype 2 in Adhesion, Invasion, and Virulence in Mice.

The sugC gene of Streptococcus suis (S. suis) is a coding gene for the ATP-binding transporter-associated protein with strong pathogenicity. In order to reveal the effect of the sugC gene on the virulence of S. suis serotype 2, a wild-type strain of TJS75, isolated from fattening pigs' brain tissue samples, was used as a parent strain, and a knockout sugC gene (ΔsugC) and complementary strain (CΔsugC) were successfully constructed via homologous recombination technology. The biological characteristics of TJS75, ΔsugC and CΔsugC were compared and analyzed through growth curves, biochemical characteristics, hemolysis characteristics, cell infection tests and pathogenicity tests on BALB/c mice. The results of the growth characteristic experiments in vitro showed that the plateau stage growth period of ΔsugC was delayed compared to the TJS75 strain, but there was no difference in the total number of bacteria. The biochemical characteristics and hemolysis ability of ΔsugC in sheep blood had no difference compared with TJS75, but its adhesion and invasion abilities in PK-15 cells were decreased. Knockout of the sugC gene had no impact on the expression levels of adhesion-related genes in TJS75 in real-time PCR analysis. In addition, the LD50 of ΔsugC in BALB/c mice was 1.47 × 108 CFU, seven times higher than that of TJS75 (LD50 = 2.15 × 107 CFU). These results illustrate that the deletion of sugC reduced the virulence of TJS75 to BALB/c mice, but its role in the adhesion and invasion of PK-15 cells in this strain needs to be further explored. In summary, this study provides evidence that the sugC gene is a virulence-related gene in the S. suis serotype 2 strain and plays a crucial role in the adhesion and invasion of S. suis. This study lays a foundation for the further exploration of the potential virulence factors and pathogenesis of S. suis.

Highly Repetitive Genome of Coniella granati (syn. Pilidiella granati), the Causal Agent of Pomegranate Fruit Rot, Encodes a Minimalistic Proteome with a Streamlined Arsenal of Effector Proteins.

This study describes the first genome sequence and analysis of Coniella granati, a fungal pathogen with a broad host range, which is responsible for postharvest crown rot, shoot blight, and canker diseases in pomegranates. C. granati is a geographically widespread pathogen which has been reported across Europe, Asia, the Americas, and Africa. Our analysis revealed a 46.8 Mb genome with features characteristic of hemibiotrophic fungi. Approximately one third of its genome was compartmentalised within 'AT-rich' regions exhibiting a low GC content (30 to 45%). These regions primarily comprised transposable elements that are repeated at a high frequency and interspersed throughout the genome. Transcriptome-supported gene annotation of the C. granati genome revealed a streamlined proteome, mirroring similar observations in other pathogens with a latent phase. The genome encoded a relatively compact set of 9568 protein-coding genes with a remarkable 95% having assigned functional annotations. Despite this streamlined nature, a set of 40 cysteine-rich candidate secreted effector-like proteins (CSEPs) was predicted as well as a gene cluster involved in the synthesis of a pomegranate-associated toxin. These potential virulence factors were predominantly located near repeat-rich and AT-rich regions, suggesting that the pathogen evades host defences through Repeat-Induced Point mutation (RIP)-mediated pseudogenisation. Furthermore, 23 of these CSEPs exhibited homology to known effector and pathogenicity genes found in other hemibiotrophic pathogens. The study establishes a foundational resource for the study of the genetic makeup of C. granati, paving the way for future research on its pathogenicity mechanisms and the development of targeted control strategies to safeguard pomegranate production.

Pheno- and genotypic epidemiological characterization of Staphylococcus aureus isolated from bulk tank milk in Colombia

Staphylococcus aureus is recognized for causing contagious mastitis in cattle. Elimination of the organism in affected animals is challenging due to its potential antibiotic resistance mechanisms and virulence factors. The aim of the study was the pheno- and genotypic epidemiological characterization of S. aureus isolated from bulk tank milk samples obtained from three municipalities in the northern region of Antioquia. Twenty-one S. aureus isolates from 150 bulk tank milk samples were characterized by evaluating antimicrobial susceptibility to 17 antibiotics using the VITEK 2, and identifying several genes encoding for virulence factors, enterotoxins, and the agr groups by conventional PCR. The clonal relationship between the isolates was assessed using macrorestriction fragment analysis (MRFA) of chromosomal DNA by pulsed-field gel electrophoresis (PFGE). Most of the isolates showed susceptibility to the tested antibiotics; 19% exhibited resistance to tetracycline and all isolates showed beta-lactamases. Molecular typing revealed that 76% carried the agrI and cap5 genes. Seven isolates harbored genes coding for staphylococcal enterotoxins (SE), with seh being the most identified gene. Furthermore, MRFA demonstrated high heterogeneity among the isolates, resulting in the assignment of 18 different MRFA patterns. These results indicate phenotypic resistance to tetracycline and beta-lactams, the presence of genes encoding for staphylococcal enterotoxins, and high heterogeneity among S. aureus isolates from bulk tank milk.

Comparison of Hypervirulent and Non-Hypervirulent Carbapenem-Resistant Acinetobacter baumannii Isolated from Bloodstream Infections: Mortality, Potential Virulence Factors, and Combination Therapy In Vitro.

Hypervirulent carbapenem-resistant Acinetobacter baumannii (hv-CRAB) has emerged in bloodstream infections (BSI). Cases of BSI caused by hv-CRAB (hv-CRAB-BSI) had posed a significant threat to hospitalized patients. In this study, 31 CRAB strains isolated from Chinese BSI patients were analyzed, of which 24 were identified as hv-CRAB-BSI and 7 as non-hv-CRAB-BSI, using the Galleria mellonella infection model. Patients with hv-CRAB-BSI had higher rates of septic shock (79.2% vs. 14.3%, p = 0.004) and mortality (66.7% vs. 14.3%, p = 0.028). All strains were resistant to most antibiotics but sensitive to colistin. Hv-CRAB-BSI showed lower resistance to minocycline than non-hv-CRAB-BSI (54.2% vs. 100%, p = 0.03). Whole-genome sequencing revealed that the detection rates of immune modulation genes ptk and epsA in hv-CRAB-BSI were significantly higher than in non-hv-CRAB-BSI (91.7% vs. 28.6%, p = 0.002). Additionally, all ST457 hv-CRAB-BSI lacked abaR, and all ST1486 non-hv-CRAB-BSI lacked adeG. The checkerboard dilution method assessed the efficacies of various antibiotic combinations, revealing that although synergism was rarely observed, the combination of colistin and minocycline showed the best efficacy for treating CRAB-BSI, regardless of whether the infections were hv-CRAB-BSI or non-hv-CRAB-BSI. These findings highlight the importance of analyzing molecular characteristics and exploring effective treatment strategies for hv-CRAB-BSI.

Genome analysis of the esca-associated Basidiomycetes Fomitiporia mediterranea, Fomitiporia polymorpha, Inonotus vitis, and Tropicoporus texanus reveals virulence factor repertoires characteristic of white-rot fungi.

Some Basidiomycete fungi are important plant pathogens, and certain species have been associated with the grapevine trunk disease esca. We present the genomes of 4 species associated with esca: Fomitiporia mediterranea, Fomitiporia polymorpha, Tropicoporus texanus, and Inonotus vitis. We generated high-quality phased genome assemblies using long-read sequencing. The genomic and functional comparisons identified potential virulence factors, suggesting their roles in disease development. Similar to other white-rot fungi known for their ability to degrade lignocellulosic substrates, these 4 genomes encoded a variety of lignin peroxidases and carbohydrate-active enzymes (CAZymes) such as CBM1, AA9, and AA2. The analysis of gene family expansion and contraction revealed dynamic evolutionary patterns, particularly in genes related to secondary metabolite production, plant cell wall decomposition, and xenobiotic degradation. The availability of these genomes will serve as a reference for further studies of diversity and evolution of virulence factors and their roles in esca symptoms and host resistance.

Expression of Fungal and Host Markers in Models of Dermatophytosis on Mice and Human Epidermis

Increasing resistance of dermatophytes against antifungals creates global public health problems, rendering essential a better understanding of virulence mechanisms and factors determining host-specificity of dermatophytes. Since dermatophytes switch from a saprophytic to a parasitic lifestyle by reprogramming gene expression, reliable experimental models are needed to investigate the pathogenesis of dermatophytosis. Here, a relevant mouse model of Trichophyton benhamiae dermatophytosis was assessed, together with a model based on reconstructed human epidermis (RHE), allowing their respective validation regarding fungal gene expressed during infection. The use of a standardized inoculum induced a natural-like superficial infection in mice. The severity and persistence of lesions enabled the assessment of infection markers, including mouse-specific pro-inflammatory molecules and fungal genes previously reported as potential virulence factors. Upregulated expression of fungal genes, including those encoding subtilisins, in infected RHE revealed that dermatophytes deploy similar processes as those observed during in vivo infection. The RHE model was then used to compare infections by anthropophilic Trichophyton rubrum and zoophilic T. benhamiae. Therefore, these two models represent complementary analytical tools to study the pathogenesis of acute dermatophytoses. In addition, we have identified certain fungal markers of infection and highlighted the existence of different mechanisms deployed by zoophilic versus anthropophilic dermatophytes.

Comparative genomics between Trichomonas tenax and Trichomonas vaginalis: CAZymes and candidate virulence factors.

The oral trichomonad Trichomonas tenax is increasingly appreciated as a likely contributor to periodontitis, a chronic inflammatory disease induced by dysbiotic microbiota, in humans and domestic animals and is strongly associated with its worst prognosis. Our current understanding of the molecular basis of T. tenax interactions with host cells and the microbiota of the oral cavity are still rather limited. One laboratory strain of T. tenax (Hs-4:NIH/ATCC 30207) can be grown axenically and two draft genome assemblies have been published for that strain, although the structural and functional annotation of these genomes is not available. GenSAS and Galaxy were used to annotate two publicly available draft genomes for T. tenax, with a focus on protein-coding genes. A custom pipeline was used to annotate the CAZymes for T. tenax and the human sexually transmitted parasite Trichomonas vaginalis, the most well-characterized trichomonad. A combination of bioinformatics analyses was used to screen for homologs of T. vaginalis virulence and colonization factors within the T. tenax annotated proteins. Our annotation of the two T. tenax draft genome sequences and their comparison with T. vaginalis proteins provide evidence for several candidate virulence factors. These include candidate surface proteins, secreted proteins and enzymes mediating potential interactions with host cells and/or members of the oral microbiota. The CAZymes annotation identified a broad range of glycoside hydrolase (GH) families, with the majority of these being shared between the two Trichomonas species. The presence of candidate T. tenax virulence genes supports the hypothesis that this species is associated with periodontitis through direct and indirect mechanisms. Notably, several GH proteins could represent potential new virulence factors for both Trichomonas species. These data support a model where T. tenax interactions with host cells and members of the oral microbiota could synergistically contribute to the damaging inflammation characteristic of periodontitis, supporting a causal link between T. tenax and periodontitis.

Extracellular BSA-degrading SAPs in the rare pathogen Meyerozyma guilliermondii strain SO as potential virulence factors in candidiasis

Meyerozyma guilliermondii (Candida guilliermondii) is one of the Candida species associated with invasive candidiasis. With the potential for expressing industrially important enzymes, M. guilliermondii strain SO possessed 99 % proteome similarity with the clinical ATCC 6260 isolate and showed pathogenicity towards zebrafish embryos. Recently, three secreted aspartyl proteinases (SAPs) were computationally identified as potential virulence factors in this strain without in vitro verification of SAP activity. The quantification of Candida SAPs activity in liquid broth were also scarcely reported. Thus, this study aimed to characterize M. guilliermondii strain SO's ability to produce SAPs (MgSAPs) in different conditions (morphology and medium) besides analyzing its growth profile. MgSAPs' capability to cleave bovine serum albumin (BSA) was also determined to propose that MgSAPs as the potential virulence factors compared to the avirulent Saccharomyces cerevisiae. M. guilliermondii strain SO produced more SAPs (higher activity) in yeast nitrogen base-BSA-dextrose broth compared to yeast extract-BSA-dextrose broth despite insignificantly different SAP activity in both planktonic and biofilm cells. FeCl3 supplementation significantly increased the specific protein activity (∼40 %). The BSA cleavage by MgSAPs at an acidic pH was proven through semi-quantitative SDS-PAGE, sharing similar profile with HIV-1 retropepsin. The presented work highlighted the MgSAPs on fungal cell wall and extracellular milieu during host infection could be corroborated to the quantitative production in different growth modes presented herein besides shedding lights on the potential usage of retropepsin's inhibitors in treating candidiasis. Molecular and expression analyses of MgSAPs and their deletion should be further explored to attribute their respective virulence effects.

The role of cytolethal distending toxin in Glaesserella parasuis JS0135 strain infection: Cytotoxicity, phagocytic resistance and pathogenicity

Glaesserella parasuis is an important porcine pathogen that commonly colonizes the upper respiratory tract of pigs and is prone to causing Glässer’s disease under complex conditions. As yet, the disease has led to serious economic losses to the swine industry worldwide. Studies so far have found that several virulence factors are associated with the pathogenicity of G. parasuis, but the pathogenic mechanism is still not fully understood. Cytolethal distending toxin (CDT), a potential virulence factor in G. parasuis, is involved in cytotoxicity, serum resistance, adherence to and invasion of host cells in vitro. Here, to further investigate the pathogenic role of CDT during G. parasuis infection in vitro and in vivo, a double cdt1 and cdt2 deletion mutant (Δcdt1Δcdt2) without selectable marker was first generated in G. parasuis JS0135 strain by continuous natural transformations and replica plating. Morphological observation and lactate dehydrogenase assay showed that the Δcdt1Δcdt2 mutant was defective in cytotoxicity. Additionally, the Δcdt1Δcdt2 mutant was more susceptible to phagocytosis caused by 3D4/2 macrophages compared to the wild-type JS0135 strain. Moreover, by focusing on clinical signs, necropsy, bacterial recovery and pathological observation, we found that the deletion of cdt1 and cdt2 genes led to a significant attenuation of virulence in G. parasuis. Taken together, these findings suggest that as an important virulence factor, CDT can significantly affect the pathogenicity of G. parasuis.

Mycolyltransferase is important for biofilm formation and pathogenesis of Tsukamurella keratitis

ABSTRACT Tsukamurella, a group of multi-drug resistant, Gram-positive, aerobic, and partially acid-fast bacteria, are emerging causes of bacterial conjunctivitis and keratitis. However, the pathogenesis of Tsukamurella keratitis is largely unknown. To address this, we used New Zealand White rabbits to develop the first eye infection model and conducted in vitro tests to study the pathogenesis mechanisms of Tsukamurella. There is increasing evidence that biofilms play a significant role in ocular infections, leading us to hypothesize that biofilm formation is crucial for effective Tsukamurella infection. In order to look for potential candidate genes which are important in biofilm formation and Tsukamurella keratitis. We performed genome sequencing of two ocular isolates, T. pulmonis-PW1004 and T. tyrosinosolvens-PW899, to identify potential virulence factors. Through in vitro and in vivo studies, we characterized their biological roles in mediating Tsukamurella keratitis. Our findings confirmed that Tsukamurella is an ocular pathogen by fulfilling Koch's postulates, and using genome sequence data, we identified tmytC, encoding a mycolyltransferase, as a crucial gene in biofilm formation and causing Tsukamurella keratitis in the rabbit model. This is the first report demonstrating the novel role of mycolyltransferase in causing ocular infections. Overall, our findings contribute to a better understanding of Tsukamurella pathogenesis and provide a potential target for treatment. Specific inhibitors targeting TmytC could serve as an effective treatment option for Tsukamurella infections.

“Phylogenomic insights into brucellaceae: The Pseudochrobactrum algeriensis case”

The genus Pseudochrobactrum encompasses free-living bacteria phylogenetically close to Ochrobactrum opportunistic pathogens and to Brucella, facultative intracellular parasites causing brucellosis, a worldwide-extended and grave zoonosis. Recently, Pseudochrobactrum strains were isolated from Brucella natural hosts on Brucella selective media, potentially causing diagnostic confusions. Strikingly, P. algeriensis was isolated from cattle lymph nodes, organs that are inimical to bacteria. Here, we analyse P. algeriensis potential virulence factors in comparison with Ochrobactrum and Brucella. Consistent with genomic analyses, Western-Blot analyses confirmed that P. algeriensis lacks the ability to synthesize the N-formylperosamine O-polysaccharide characteristic of the lipopolysaccharide (LPS) of smooth Brucella core species. However, unlike other Pseudochrobactrum but similar to some early diverging brucellae, P. algeriensis carries genes potentially synthetizing a rhamnose-based O-polysaccharide LPS. Lipid A analysis by MALDI-TOF demonstrated that P. algeriensis LPS bears a lipid A with a reduced pathogen-associated molecular pattern, a trait shared with Ochrobactrum and Brucella that is essential to generate a highly stable outer membrane and to delay immune activation. Also, although not able to multiply intracellularly in macrophages, the analysis of P. algeriensis cell lipid envelope revealed the presence of large amounts of cationic aminolipids, which may account for the extremely high resistance of P. algeriensis to bactericidal peptides and could favor colonization of mucosae and transient survival in Brucella hosts. However, two traits critical in Brucella pathogenicity are either significantly different (T4SS [VirB]) or absent (erythritol catabolic pathway) in P. algeriensis. This work shows that, while diverging in other characteristics, lipidic envelope features relevant in Brucella pathogenicity are conserved in Brucellaceae. The constant presence of these features strongly suggests that reinforcement of the envelope integrity as an adaptive advantage in soil was maintained in Brucella because of the similarity of some environmental challenges, such as the action of cationic peptide antibiotics and host defense peptides. This information adds knowledge about the evolution of Brucellaceae, and also underlines the taxonomical differences of the three genera compared.

Genomic and molecular characterization of a cyprinid herpesvirus 2 YC-01 strain isolated from gibel carp

Cyprinid herpesvirus 2 (CyHV-2) is the pathogen of herpesviral hematopoietic necrosis (HVHN), causing the severe economic losses in farmed gibel carp (Carassius gibelio). Further exploration of the genome structure and potential molecular pathogenesis of CyHV-2 through complete genome sequencing, comparative genomics, and molecular characterization is required. Herein, the genome of a CyHV-2 YC-01 strain isolated from diseased gibel carp collected in Yancheng, Jiangsu Province, China was sequenced, then we analyzed the genomic structure, genetic properties, and molecular characterization. First, the complete YC-01 genome comprises 275,367 bp without terminal repeat (TR) regions, with 151 potential open reading frames (ORFs). Second, compared with other representative published strains of the genus Cyvirus, several evident variations are found in YC-01, particularly the orientation and position of ORF25 and ORF25B. ORF107 and ORF156 are considered as potential molecular genetic markers for YC-01. ORF55 (encoding thymidine kinase) might be used to distinguish YC-01 and ST-J1 from other CyHV-2 isolates. Third, phylogenetically, YC-01 clusters with the members of the genus Cyvirus (together with the other six CyHV-2 isolates). Fourth, 43 putative proteins are predicted to be functional and are mainly divided into five categories. Several conserved motifs are found in nucleotide, amino acid, and promoter sequences including cis-acting elements identification of YC-01. Finally, the potential virulence factors and linear B cell epitopes of CyHV-2 are predicted to supply possibilities for designing novel vaccines rationally. Our results provide insights for further understanding genomic structure, genetic evolution, and potential molecular mechanisms of CyHV-2.

Genome Sequence Comparisons between Small and Large Colony Phenotypes of Equine Clinical Isolates of Arcanobacterium hippocoleae.

Arcanobacterium hippocoleae is a Gram-positive fastidious bacterium and is occasionally isolated from the reproductive tract of apparently healthy mares (Equus caballus) or from mares with reproductive tract abnormalities. Apart from a few 16S rRNA gene-based GenBank sequences and one recent report on complete genome assembly, detailed genomic sequence and clinical experimental data are not available on the bacterium. Recently, we observed an unusual increase in the detection of the organism from samples associated with mare reproductive failures in Atlantic Canada. Two colony morphotypes (i.e., small, and large) were detected in culture media, which were identified as A. hippocoleae by MALDI-TOF mass spectrometry and 16S rRNA gene sequencing. Here, we report the whole genome sequencing and characterization of the morphotype variants. The genome length of the large phenotypes was between 2.42 and 2.43, and the small phenotype was 1.99 Mbs. The orthologous nucleotide identity between the large colony phenotypes was ~99%, and the large and small colony phenotypes was between 77.86 and 78.52%, which may warrant the classification of the two morphotypes into different species. Phylogenetic analysis based on 16S rRNA genes or concatenated housekeeping genes grouped the small and large colony variants into two different genotypic clusters. The UvrA protein, which is part of the nucleotide excision repair (NER) system, and 3-isopropoylmalate dehydratase small subunit protein expressed by the leuD gene were identified as potential virulence factors in the large and small colony morphotypes, respectively. However, detailed functional studies will be required to determine the exact roles of these and other identified hypothetical proteins in the cellular metabolism and potential pathogenicity of A. hippocoleae in mares.

Leptospiral LipL45 lipoprotein undergoes processing and shares structural similarities with bacterial sigma regulators.

Leptospirosis is a widespread zoonotic infectious disease of human and veterinary concern caused by pathogenic spirochetes of the genus Leptospira. To date, little progress towards understanding leptospiral pathogenesis and identification of virulence factors has been made, which is the main bottleneck for developing effective measures against the disease. Some leptospiral proteins, including LipL32, Lig proteins, LipL45, and LipL21, are being considered as potential virulence factors or vaccine candidates. However, their function remains to be established. LipL45 is the most expressed membrane lipoprotein in leptospires, upregulated when the bacteria are transferred to temperatures resembling the host, expressed during infection, suppressed after culture attenuation, and known to suffer processing in vivo and in vitro, generating fragments. Based on body of evidence, we hypothesized that the LipL45 processing might occur by an auto-cleavage event, deriving two fragments. The results presented here, based on bioinformatics, structure modeling analysis, and experimental data, corroborate that LipL45 processing probably includes a self-catalyzed non-proteolytic event and suggest the participation of LipL45 in cell-surface signaling pathways, as the protein shares structural similarities with bacterial sigma regulators. Our data indicate that LipL45 might play an important role in response to environmental conditions, with possible function in the adaptation to the host.

Genome-wide expanding of genetic evolution and potential pathogenicity in Vibrio alginolyticus

ABSTRACT Vibrio alginolyticus, an emergent species of Vibrio genus, exists in aquatic and marine environments. It has undergone genetic diversification, but its detailed genomic diversity is still unclear. Here, we performed a multi-dimensional comparative genomic analysis to explore the population phylogeny, virulence-related genes and potential drug resistance genes of 184 V. alginolyticus isolates. Although genetic diversity is complex, we analysed the population structure using three sub-datasets, including the subdivision for three lineages into sublineages and the distribution of strains in the marine ecological niche. Accessory genes, most of which reclassified V. alginolyticus genomes as different but with relatively close affinities, were nonuniformly distributed among these isolates. We demonstrated that the spread of some post-evolutionary isolates (mainly L3 strains isolated from Chinese territorial seas) was likely to be closely related to human activities, whereas other more ancestral strains (strains in the L1 and L2) tended to be locally endemic and formed clonal complex groups. In terms of pathogenicity, the potential virulence factors were mainly associated with toxin, adherence, motility, chemotaxis, and the type III secretion system (T3SS). We also found five types of antibacterial drug resistance genes. The prevalence of β-lactam resistance genes was 100%, which indicated that there may be a potential risk of natural resistance to β-lactam drugs. Our study reveals insights into genomic characteristics, evolution and potential virulence-associated gene profiles of V. alginolyticus.