Current Microbiology

Isolation of extracellular enzyme degrading aerobic bacteria in the digestive tracts of fresh water prawn was carried out. Gut bacteria were isolated on carboxymethyl cellulose agar plates, starch agar media plates, gelatine peptone agar media plates. The isolated strain was qualitatively screen based on their extracellular enzyme on selective media. The strain was characterised based on morphological, physiological and biochemical characterization identified Bacillus species. Bacillus species was isolated by colony characterization carried out by using Gelatin agar Media, carboxymethyl cellulose media and congo red CMC media and starch agar media for different enzyms. Isolates are capable of hydrolysing proteins and carbohydrates indicating their importance in fish nutrition.

The stringent response has far-reaching consequences, with links to antimicrobial resistance, stress and virulence. This study assessed the role of relA in nutritional, environmental and antimicrobial stress in Klebsiella pneumoniae, the extent of polysaccharide capsule and biofilm formation, and the pathogenic effect on Drosophila melanogaster. Two single mutants (KP03DrelA and KPPR1DrelA) obtained using the Lambda Red Recombinase Technique were the focus of analyses. We assessed environmental (ethanol, osmotic, heat) and nutritional stress (carbon, phosphate, amino acid) tolerance, capsule formation, and cell size determination in wild-type (WT) and mutant bacteria. Biofilm and agar plate susceptibility assays were performed on both starved and non-starved strains using gentamicin and ceftazidime, and stress response genes were analyzed. Drosophila melanogaster was used to examine the pathogenic effect of the presence or deletion of relA on fly survival. DrelA mutants had reduced fitness to environmental and nutritional stress compared to WT strains. Mutant strains cell lengths were elongated and lacked a capsule versus WT and non-starved strains, and mutant strains exhibited enhanced biofilm formation or survival in the presence of ceftazidime and gentamicin. Stress response genes rpoD, phoR, phoU and pstB were absent during starvation, but rpoS was detected in C-, C+PO+ and serine hydroxamate (SHX) media; mrkA gene was not detected during starvation. Lastly, the animal model proved to be effective in showing infection levels associated with the presence of the relA gene.

With the growing antibiotic resistance in mycobacterial species posing a significant threat globally, there is an urgent need to find alternative solutions. Bacteriophage-derived endolysins aid in releasing phage progeny from the host bacteria by attacking the cell wall at the end of their life cycle. Endolysins are attractive antibacterial candidates due to their rapid lytic action, specificity and low risk of resistance development. In mycobacteria, owing to the complex, hydrophobic cell wall, mycobacteriophages usually synthesize two endolysins: LysinA, which hydrolyzes peptidoglycan; LysinB, which delinks mycolylarabinogalactan from peptidoglycan and releases mycolic acid. In this study, we conducted domain analysis and functional characterization of a recombinant LysinB from RitSun, an F2 sub-cluster mycobacteriophage. Several properties of RitSun LysinB are important as an antimycobacterial agent: its ability to lyse Mycobacterium smegmatis from without, a specific activity of 1.36 U/mg, higher than the reported ones and its inhibitory effect on biofilm formation. Given the impervious nature of the mycobacterial cell envelope, native endolysins ability to damage cells on exogenous applications warrants further investigation. A molecular dissection of RitSun LysinB to identify its cell wall destabilizing sequence could be utilized to engineer other native lysins as fusion proteins and expand their activity profile.

A highly active alkaline phosphatase (ALP) from the mollusk strain of the marine bacterium Cobetia amphilecti KMM 296 (CmAP) was found to remove phosphorus from the Escherichia coli lipopolysaccharides (LPS). Phylogenetic analysis of the amino acid sequences of ALPs found in 36 available Cobetia genomes revealed that CmAP and its homologues from nine strains clustered together with the human and squid LPS-detoxifying enzymes. Each strain of the genus Cobetia has a variety of ALPs mostly of the PhoD and PhoX families. The PhoA gene encoding for the CmAP-like ALP is characteristic for the subspecies C. amphilecti, with a complete set of four ALP families, including PafA and two PhoD structures (5 genes). However, a single strain of the species Cobetia crustatorum JO1T from fermented shrimp, phylogenetically distant from C. amphilecti and C. marina, among four ALPs contains a CmAP homologue carrying an inactive mutation. Apparently, the multiplicity of ALPs in bacteria of the genus Cobetia is a trait of incredible adaptation to a phosphorus-depleted environment and a specialty of organophosphate destructor in eco-niches to which they once emerged, including Zostera spp. roots. The ALP clusterization and an identity level of the genus-specific biosynthetic genes encoding for ectoine and polyketide cluster T1PKS, responsible for sulfated extracellular polysaccharide synthesis, coincide with a new whole genome-based taxonomic classification of the genus Cobetia. The LPS-dephosphorylating property of the PhoA family C. amphilecti ALP CmAP may be used in the development of anti-inflammatory drugs.

Genome sequence-based identification of two strains (3986T and 51.81) isolated from rabbits in France in 1972 and 1981 and deposited in the Collection of Institut Pasteur (CIP) has led to the description of a novel species in the genus Neisseria. The cells of both strains were non-motile, Gram-stain-negative and diplococcobacilli. Optimal growth on trypticase soy agar was recorded at 37{degrees}C and pH 8.5 in aerobic conditions. Phylogeny based on 16S rRNA gene placed the strains close to Neisseria bacilliformis ATCC BAA-1200T (96.38%) nesting with the members of Neisseriaceae family. Furthermore, the phylogenetic analysis based on bac120 gene set from the Genome Taxonomy Database (GTDB) placed both strains within the monophyletic Neisseria clade, which also included type strains of Morococcus cerebrosus, Bergeriella denitrificans, Kingella potus, Uruburuella suis and Uruburuella testudinis. However, Neisseria shayeganii strain 871T was placed outside Neisseria clade and close to the members of Eikenella genus. Strains 3986T and 51.81 were placed in a branch distinct from all species of the genus Neisseria and exhibited the average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values below the species demarcation values. In contrast, ANI value within the two strains was 96.9% confirming that they represent same species. The genomic DNA G+C content of strain 3986T was 56.92%. Based on the phylogenetic and phenotypic data, the strains 3986T and 51.81 represent a novel species of the genus Neisseria, for which the name Neisseria leonis sp. nov. is proposed (type strain 3986T = CIP 109994T = LMG 32907T). Additionally, based on phylogenetic analysis, DUS dialect and average amino acid identity (AAI) values, we also proposed the reclassification of Morococcus cerebrosus, Bergeriella denitrificans, Kingella potus, Uruburuella suis and Uruburuella testudinis into Neisseria genus and Neisseria shayeganii into Eikenella genus. Author NotesThe GenBank accession numbers for the 16S rRNA gene sequence of strains 3986T and 51.81 are respectively OQ121838.1 and OQ428162.1. The draft genome sequences have been deposited in GenBank under the accession numbers JAPQFK000000000 (strain 3986T) and JAPQFL000000000 (strain 51.81). Further explanations mentioned in the article as well as 7 supplementary tables and 7 supplementary figures are available with the online version of this article.

Horizontal gene transfer (HGT) significantly influences prokaryotic genome evolution. Bacillus cereus and Bacillus thuringiensis are nearly identical at the chromosomal level, except for B. thuringiensis producing parasporal crystals. The genes for these crystal proteins (e.g., cry1A), along with other encoded insecticidal proteins (e.g., vip3A), are located on megaplasmids and can be horizontally transferred. Recently, Sauka et al. (2022) reported a Bacillus toyonensis strain that produces parasporal crystals with dual insecticidal activity. This strain was classified as Bacillus toyonensis biovar Thuringiensis (NCBI: txid2923195) following Carroll et al.s (2020) nomenclature. Misclassified B. toyonensis strains, previously identified as B. thuringiensis (e.g., strain MC28), encode cry and cyt genes toxic to lepidopteran and dipteran insects. Advances in genome sequencing and bioinformatics tools now reduce misidentifications, enabling accurate reclassification in databases like GenBank. These findings highlight the need for genome-based taxonomic reassessment within the Bacillus cereus group and clarify the chromosomal placement of crystal-forming B. toyonensis strains.

Two yellow pigmented bacterial strains were isolated from healthy rice seeds. The strains designated as PPL560T and PPL568 were identified as members of genus Xanthomonas based on analysis of biochemical and 16S rRNA gene sequence retrieved from whole genome sequence. Isolates formed a distinct monophyletic lineage with X. sontii and X. sacchari as the closest relatives in the phylogenetic tree based on core gene content shared by reported species in genus Xanthomonas. Pairwise ortho Average Nucleotide Identity and digital DNA-DNA hybridisation values calculated against other species of Xanthomonas were below their respective cut-offs. In planta studies revealed that PPL560T and PPL568 are non-pathogenic to rice plants upon leaf clip inoculation. Absence of type III secretion system related genes and effectors further supported their non-pathogenic status. Herein, we propose Xanthomonas indica sp. nov. as novel species of genus Xanthomonas with PPL560T=MTCC13185 as its type strain and PPL568 as another constituent member.

L-asparaginase is a promising enzyme for cancer treatment and is found in plants, animals and microbes. This enzyme is of great medical and industrial importance. It is used with inside the remedy of acute lymphoblastic leukemia and helps in reducing the acryl amide substances found in fried and baked foods. Its source varies from bacteria to yeast and fungi. This study aimed to screen the potential L-asparaginase producing E. coli isolates among river water and cafeteria sewage samples near the Gubryie area, SNNPR-Ethiopia. In this study, E. coli isolates were isolated from sewage from the Wabe River and Wolkite University student cafeteria. During the study, 14 isolates, 11 from cafeteria sewage and 3 from Wabe River, were confirmed to be E. coli using IMViC, TSI, SCA, and Gram tests. For the E. coli-positive samples, screening of L-asparaginase was performed using the phenol red indicator. The change in color from yellow to pink in M9 media due to the acidic environment created when L-asparagine was degraded to urea indicates the presence of L-asparginase in the potent E. coli cells. The production of L-asparaginase was carried out using submerged fermentation method. Mechanical cell disruption method, high speed centrifugation, was used to separate the secreted enzyme from cells. The potential of the E. coli cells to produce L-asparginase was also checked using a rapid plate assay method with the indicator dye phenol red. The zone of inhibition for the intracellular enzyme activity ranges from 16.5 mm up to 22.25 mm while that of extracellular enzyme ranged from 7.5 mm up to 9 mm. The commonly used software system is SPSS version 23. The PCR result depicted that the ansA gene presence was confirmed in 50% of the isolates. The result confirmed that the E.coli isolates from sewage showed better L-asparaginase production potency than the Wabe River isolates. This study indicated that E. coli strains are promising sources of L-Asparginase for food and pharmacological companies if the scale-up of this work has been completed in the future.

Citrus is an economically important fruit crop grown in all provinces of Pakistan, while Punjab accounting for 95 percent of total production due to its favorable climate for citrus production. Commercially grown varieties in Pakistan include sweet oranges, grapefruits, Mandarine, Lime, and lemon. The goal of this research was to see how diverse the cultivable bacterial populations are found in citrus cultivars. Out of 90 isolated cultures, 37 endophytic bacterial species and 15 different genera of bacteria were characterized based on morphological, biochemical, and molecular methods from citrus leaves. All the isolated bacteria were subjected to PCR amplification through 16S rDNA followed by sequencing. RDP base classification revealed that class Bacilli has the largest percentage of isolates, whereas class Alpha, Beta, and Gamma Proteobacteria have the lowest percentage among all genotypes used. According to the findings, the phylum Firmicutes contains a common genus (Brevibacterium, 1%; Enterococcus, 6%; Staphylococcus, 7% and Bacillus, 60%). Alpha (Rhizobium) beta (Burkholderia cepacia; Comamonas terigena) gamma Proteobacteria (Enterobacter hermachei (1%), Klebsiella pneumoniae (1%), Proteus mirabilis (8%), Pseudomonas aeruginosa (5%), Psychrobacter pulmonis and Yersinia molalretti (1%) respectively. These results revealed that cultivars of the plants might contribute to the structure and endophytic bacterial communities associated with citrus. Endophytes extracted from leaf samples of different citrus cultivars in Pakistan are reported for the first time. The idea of employing endophytes bacteria to produce enzymes stimulate plant growth, and its purpose as a biological control agent will be investigated in the future.

There is a barnacle larva settlement model in which barnacle cryprid selectively locates itself only on a specific marine biofilm area that contains specific microbes. That means a local biofilm site with specific characteristics can attract barnacle larva to the maximum. If this is true, a barnacle already growing up shall still press down a chunk of biofilm area where it settles when it was a crypid. The chunk of biofilm should be at the center of barnacle footprint and may still contain most of the microbes at the attachment site. By this consideration, a group of such chunks of barnacle cement (with about 2mm diameter) was collected from the center of barnacle footprints, followed by genomic DNA extraction, PCR amplification with primers representing prokaryotes, eukaryotes, archaea and fungus, DNA sequencing and species determination. The most abundant 13 species were preliminarily determined (mainly fungi). Whether they are really wanted target microbes largely depends on future investigations on whether they possess some common features that can attract barnacle crypids.

Phycosphere harbors cross-kingdom interactions with significant ecological relevance for harmful algal blooms (HAB) and phycotoxins biosynthesis. Previously, a new red-pigmented bacterium designated as strain LZ-28 was isolated from phycosphere microbiota of typical HAB dinoflagellate Alexandrium catenella LZT09 which is a vitamin B12 auxotroph and produces high levels of paralytic shellfish poisoning toxins (PST). Strain LZ-28 exhibited obvious growth-promoting activity toward its algal host, along with the production of active bioflocculanting exopolysaccharides (EPS). But the phylogenetic affiliation and genomic potential of this versatile bacterium has not yet been elucidated. In this study, we carried out combined taxonomic and phylogenomic analysis to clarify the taxonomic classification of strain LZ-28. The obtained 16S rRNA phylogeny revealed close taxonomic relationship between strain LZ-28 and other Mameliella alba members. Additional calculations of key phylogenomic parameters, average nucleotide identity (ANI), the average amino acid identity (AAI) and the digital DNA-DNA hybridization (dDDH) values based on genomes of strain LZ-28 and type strain of Mameliella alba were all exceeded the limit of species circumscription. Collectively considering the phenotypic and biochemical characterizations, strain LZ-28 was therefore identified as a new member of Mameliella alba. Furthermore, based on the genomic evidence, potential algae-bacteria interactions of strain LZ-28 with host algae LZT09 were elucidated through the associations with photosynthetic and antioxidant carotenoids, supplying of bacterial VB12 to auxotroph host, and versatile EPS serving for bacterial colonization and nutrient exchange during their interactions, along with stress response systems to defense oxidative stress and quorum sensing (QS) signals benefited survival for bacteria in the symbiotic system. Comparative genomics shed light on similar genomic features between M. alba strains, revealed potential close associations of strain LZ-28 with its algae host, and further enriched the genomic repertoire of interactions between phycosphere microbiota and algal host LZT09.

This study aimed to investigate the antibiotic susceptibility, antibiotic resistance genes (ARGs), mobile genetic elements (MGEs), virulence genes, and molecular epidemiology of carbapenem-resistant Klebsiella pneumoniae (CRKP) strains isolated from giant pandas. The screening of 178 nonduplicated Klebsiella pneumoniae strains identified eight CRKP strains, with the most abundant ARGs observed in ampC/blaDHA, blaSHV-01, blaSHV-02, tetB-01, tetB-02, tetC-01, and tetC-02. MGE analysis revealed the presence of intI1 in all strains, while other MGEs exhibited varying detection rates. Strain 24 exhibited the highest diversity in terms of MGE species. Seven virulence genes including wabG, uge, ycf, entB, kpn, alls, and wcaG, showed positive results with different proportions across the strains. Molecular epidemiology analysis using pulsed-field gel electrophoresis (PFGE) patterns indicated a high level of genetic diversity among the CRKP strains. Multi-locus sequence typing (MLST) analysis classified the strains into different sequence types (STs). In conclusion, this study highlighted the diverse nature of CRKP strains found in giant pandas, which exhibited varying levels of antibiotic resistance along with multiple ARGs and virulence genes present. These findings emphasized the importance of monitoring and researching antibiotic resistance within wildlife populations to safeguard the health status of these endangered animals.

Rice seed microbiota play a vital role throughout their growth and developmental stages. Xanthomonas sontii is known as an abundant, core-vertically transmitted rice seed endophyte with probiotic properties. Studies reveal a less explored world of non-pathogenic Xanthomonas (NPX) species like X. sontii and X. indica that are associated with healthy rice seeds. The present study reports the isolation, biochemical characterization, and in-depth taxonogenomic evidence for a third NPX species, X. protegens, from rice seeds, a taxonomic outlier of X. sontii. It also highlights the importance of using multiple taxonogenomic indices to identify taxonomic outlier species. Similar to the other two NPX species, the new member species is also non-pathogenic and provides in vivo protection to rice plants from Xanthomonas oryzae pv. oryzae upon leaf clip inoculation. The pangenome investigation suggested a set of unique genes to the novel species that might be important in adaptation to rice plants and differentiate it from other NPX species. The study will allow the design of markers for the identification of these NPX members, targeted culturomics of a particular NPX species and in the discovery of other novel NPX species from healthy rice seed microbiome for further exploitation in rice health.

Three novel bioluminescent bacterial strains, VNB-15T, VNB-16 and SChm4, were isolated from water of the Black Sea (Russia) and intestines of the Black Sea horse mackerel. Cells of the isolated strains are motile Gram negative slightly curved rods with single polar flagellum. The temperature range for growth was 10-35{degrees}C, the optimum being 20-25{degrees}C. The pH range for growth was 6.0-9.0, the optimum being 7.0-8.0. The bacteria were able to grow in the presence of 0.5 to 5.0% NaCl (w/v), the optimum being 1.0-4.0% (w/v). Phylogenetic analysis based on comparison of 16S rRNA sequences shows these strains to have kinship with the species Vibrio jasicida, Vibrio hyugaensis, Vibrio alginolyticus, Vibrio campbelli, Vibrio rotiferianus, Vibrio harveyi and Vibrio owensii with sequence similarity from 99.6 to 98.0%. Phylogenetic analysis based on comparison of the sequences of genes gyrB, recA, pyrH, gapA, rpoA, mreB, ftsZ, topA shows that the strains VNB-15T, VNB-16 and SChm4 to form a cluster within the V. harveyi clade and belong to a new species of the Vibrio genus. Comparison of the complete genomic sequence of VNB-15T with typical strains of nearby species also indicates that VNB-15T belongs to a separate species (maximum similarity 98% with V. hyugaensis and 96% with V. jasicida). VNB-15T differs from closely related species by its ability to utilize glucose, mannitol, inositol, sorbitol, rhamnose and sucrose, and to form lysine decarboxylase, ornithine decarboxylase, lipase, acid phosphatase, -glucosidase, {beta}-glucosidase and N-acetyl-{beta}-D-glucosaminidase enzymes. Based on phylogenetic analysis and phenotypic characteristics, Vibrio aquamarinus sp. nov. is proposed. The type strain is VNB-15T (= VKPM B-11245T = DSM 26054 T). RepositoriesThe GenBank accession numbers for the gapA, 16S rRNA, gyrB, pyrH, rpoA, recA, mreB, ftsZ, topA genes sequences of strain VNB-15T are JQ319116-JQ319121, KX242381, KX 242384, KX242387, respectively. The GenBank accession numbers for the gapA, gyrB, pyrH, recA, rpoA,16S rRNA, mreB, ftsZ, topA genes sequences of strain VNB-16 are KP221561-KP221566, KX242382, KX 242385, KX242388 respectively. The GenBank accession numbers for the 16S rRNA, gapA gyrB, rpoA, recA, pyrH, mreB, ftsZ, topA genes sequences of strain SChm4 are KX242375-KX242380, KX242383, KX 242386, KX242389, respectively. The GenBank/EMBL/DDBJ accession numbers for the housekeeping gene sequences used in this study are detailed in supplementary Table S1, Figures S1-S8. The genome of Vibrio aquamarinus sp. nov. VNB-15T, comprising two chromosomes and a plasmid, has been assembled and deposited in the NCBI database under the submission number SUB14585067

Many severe mosquito-borne diseases are transmitted by the Aedes sp. mosquitoes. Control efforts have been strengthened by the implementation of novel integrated vector management techniques such as alginate hydrogel beads and appealing toxic sugar bait. These techniques help to control mosquitoes by taking advantage of their attraction to sugar. Different types of sugar that mosquitoes ingest during feeding can affect the makeup of microbiome in the midgut. Immune priming and baseline immune activity are maintained by mosquito midgut microbiome. Thus, this study is currently focusing on using microbial communities for vector control measures with a particular emphasis on how they consume various forms of sugar. Both wild and lab strain Ae. aegypti and Ae. albopictus mosquito samples were reared and fed with attractive targeted sugar baits (ATSBs) infused with Chrysanthemum, mango, mix and control solutions. Then, the impact on bacterial communities was assessed by using 16S rRNA gene sequences. According to our findings, the majority of the bacterial species in mango and mix treatments belonged to the Enterobacteriaceae family. A total of 24 various bacterial species were found in Aedes mosquitoes that fed on mango ATSBs. All isolates were members of three phyla from Actinobacteria (4.16%), Firmicutes (54.17%), and Proteobacteria (41.67%). Data reveals that different species, strains and diet affect the midgut bacterial diversity in the mosquitoes. In addition to improving our knowledge concerning the way this bacterium shapes the microbial community, a thorough investigation of the prevalence of the midgut bacterial community is essential for alerting present and future mosquito and disease control initiatives.

K. pneumoniae is a non-motile, encapsulated bacterium from the Enterobacteriaceae family. The illnesses related to this opportunistic pathogen are pneumonia, Urinary Tract Infection (UTI), pyogenic liver abscess, endophthalmitis, and meningitis. Among them, UTI is predominant due to its biofilm formation leading to the mortality of 150 million people worldwide. The function of monovalent and divalent ions on Klebsiella biofilm, aside from physiochemical variables, remains unclear. Hence the present study was performed to analyze the role of K+, Ca2+, Na+, Mg2+, NH+4 in biofilm formation and its influence on biofilm-related genes. Among the tested cations, K+ and Ca2+ yielded strong biofilm in clinical and environment isolates in pH between 6.5 to 9.5. Increasing Ca2+ ions concertation reduced the Klebsiella biofilm. When compared to the hypercalciuria condition (Ca2+ level > 5 mM), 2.5 mM resulted in high biofilm mass. Cations concurrently enhanced the size of the capsule and cell density of Klebsiella but were not correlated with biofilm mass. Expression of the LPS gene (WbaG) either in planktonic or biofilm stage promoted biofilm formation in the presence of K+, Ca2+ and Na+. Whereas, expression of fimbriae genes (FimH and mrkD) was co- regulation, and capsule genes (RmpA and Wcab) were absent. Stating, the primary component needed for the Klebsiella biofilm is not the capsule or fimbriae, rather LPS. Resulting as a potent target in the treatment of Klebsiella biofilm in UTI. To the best of our knowledge, this is the first kind of study on the effect of cation ions on biofilm and planktonic cells of Klebsiella spp. and demonstrating the role of LPS biosynthesis gene (WbaG) in biofilm development.

Aeromonas hydrophila is human-animal commensal bacterium that seriously threatens the development of aquaculture and human health. Type IV pili (T4P) are essential for bacterial physiological function. The relationship between the molecular mechanism of flpL gene affiliated to T4P and the pathogenicity is still unclear. In the research, A. hydrophila with a genetically stable deletion of flpL gene ({Delta}flpL) was constructed. The median lethal dose of {Delta}flpL for Crucian carp was 4.87 times higher compared to wild-type strain, suggesting that {Delta}flpL significantly reduced pathogenicity. The attenuation may be attributed to a reduced capacity for biofilm formation and downregulation of the expression levels of virulence-related genes in {Delta}flpL. Meanwhile, the significant increase in swimming capacity and adhesion can be related to the upregulation of flpC, tapM, tapB and tapP genes. In conclusion, flpL gene plays essential role in the pathogenicity, adhesion, motility, and capacity for biofilm formation in A. hydrophila. The research aims to clarify the pathogenesis of A. hydrophila, and lay the foundation for developing live attenuated vaccines through the targeted modification flpL gene.

Vaults are large cone-shaped and highly conservative ribonucleoprotein complexes present in the cells of most major eukaryote clades. However, despite their wide distribution, their functions and evolutionary dynamics still remain enigmatic. Several minor functions in modulating signaling cascades and multidrug resistance phenotypes were previously discovered for eukaryotic vaults, yet nothing is known about bacterial homologs of the major vault protein (MVP), a protein that comprises the entirety of vault external surface. Using gene and protein BLAST searches in NCBI and UniProt databases we identified a number of bacterial species in prokaryotic orders Myxococcales, Cytophagales and Oscillatoriales with >50% identity to eukaryotic MVP sequences. Interestingly, all of these species are characterized by one common feature - gliding type of motility. Secondary structures of the identified proteins were predicted using RAPTORX web service and aligned via jFATCAT-flexible algorithm in the RCSB PDB Java Structure Alignment tool to elucidate structural identity. Coiled coil domain at the MVP C-terminus of all studied bacterial species resembled TolA protein of Escherichia coli by both structure and sequence. We also showed that MVP sequences from chemotrophic bacteria Myxococcales and Cytophagales contain a domain homologous to eukaryotic band-7 domain, unlike cyanobacterial and eukaryotic major vault proteins. As expected, maximum-likelihood phylogenetic trees for MVP sequences separate studied taxa into two clades - first clade contains Oscillatoriales (Cyanobacteria) and Eukaryotes and the second one contains chemotrophic bacteria. In addition, binding prediction via RAPTORX showed great multiplicity GMP and CMP nucleoside monophosphate binding pockets in Myxococcales and Cytophagales MVP, unlike eukaryotic and cyanobacterial proteins which had much lower affinity to these substrates. Due to high similarity of eukaryotic and cyanobacterial MVP sequences and a pattern of its phylogenetic distribution, we can speculate that the most likely scenario for vault appearance in eukaryotes is horizontal gene transfer from cyanobacteria. Presence of GMP and CMP binding pockets in MVP could also point to a function in depleting cytosolic nucleotide concentration which would be beneficial, for instance, during a viral infection. Further research is necessary to uncover potential functions of this enigmatic protein in bacteria and to determine its evolutionary patterns. In addition, a correlation between MVP presence and gliding motility in bacteria could also lead to elucidating selective pressures on the early evolution of this protein.

Bacillus cereus is a food contaminant with widely varying enterotoxic potential of its virulence proteins. In this article, phylogenetic analysis of the whole-genome amino acid sequences of 41 strains, evolutionary distance calculation of the amino acid sequences of the virulence genes, and functional and structural prediction of the virulence proteins were performed to reveal the taxonomically diverse distribution of virulence factors. The genome evolution of the strains showed a clustering trend based on the coding virulence genes. The strains of B. cereus have evolved into non-toxic risk and toxic risk clusters with medium-high- and medium-low-risk clusters. The distances of evolutionary transfer relative to housekeeping genes of incomplete virulence genes were greater than those of complete virulence genes, and the distance values of HblACD were higher than those of nheABC and CytK among the complete virulence genes. Cytoplasmic localization was impossible for all the virulence proteins, and NheB, NheC, Hbl-B, and Hbl-L1 were extracellular according to predictive analysis. Nhe and Hbl proteins except CytK had similar spatial structures. The predicted structures of Nhe and Hbl mainly showed head and tail domains. The head of NheA and Hbl-B, including two -helices separated by {beta}-tongue strands, might play a special role in Nhe trimers and Hbl trimers, respectively. The cap of CytK, which includes two latches with many {beta}-sheets, formed a {beta}-barrel structure with pores, and a rim balanced the structure. The evolution of B. cereus strains showed a clustering tendency based on the coding virulence genes, and the complete virulence-gene operon combination had higher relative genetic stability. The beta-tongue or latch associated with {beta}-sheet folding might play an important role in the binding of virulence structures and pore-forming toxins in B. cereus.

AimsThe gut microbiota is referred to an extra organ and is ciritical in assisting the host in terms of nutrition and immunity. Environmental stressors could alter gut microbial community and cause gut inflammation. This study aimed to investigate and compare the gut microbiota community between healthy and diseased Tor tambroides. Methodology and resultsIn this study, such gut microbial alterations were explored using NGS-based 16S rDNA sequencing on the Malaysian mahseer (T. tambroides). Three adult healthy and three diseased adult Malaysian mahseers (showing signs of exophthalmia, coelomic distension and petechial haemorrhage) were obtained from LTT Aquaculture Sdn Bhd. Our results revealed significant differences in microbial diversity, composition and function between both populations of T. tambroides. Alpha diversity analysis depicts lower diversity of gut microbiota composition in diseased T. tambroides as compared to the healthy group. In particular, Enterobacteriaceae, Aeromonas, Bacteroides, Vibrio and Pseudomonas were found within gut microbiota of the diseased fishes. In addition, cellulose-degrading bacteria and protease-producing bacteria were identified from the gut of T. tambroides. Conclusion significance and impact of studyThus, our findings emphasised on the association between the alteration in gut microbiota composition and infectious abdominal dropsy (IAD) in T. tambroides. This finding is important to provide basic information for further diagnosis, prevention and treatment of intestinal diseases in fish.