A comparative whole genome analysis of Helicobacter pylori from a human dense South Asian setting.

Helicobacter pylori, a Gram-negative bacterium, is associated with a wide range of gastric diseases such as gastritis, duodenal ulcer, and gastric cancer. The prevalence of H pylori and risk of disease vary in different parts of the world based on the prevailing bacterial lineage. Here, we present a contextual and comparative genomics analysis of 20 clinical isolates of H pylori from patients in Bangladesh. Despite a uniform host ethnicity (Bengali), isolates were classified as being part of the HpAsia2 (50%) or HpEurope (50%) population. Out of twenty isolates, eighteen isolates were cagA positive, with two HpEurope isolates being cagA negative, three EPIYA motif patterns (AB, AB-C, and ABC-C) were observed among the cagA-positive isolates. Three vacA genotypes were observed with the s1m1i1dic1 genotype observed in 75% of isolates; the s1m2i1d1c2 and s2m2i2d2c2 genotypes were found to be 15% and 10% of isolates, respectively. The non-virulent genotypes s2m2i2d2c2 was only observed in HpEurope population isolates. Genotypic analysis of oipA gene, present in all isolates, revealed five different patterns of the CT repeat; all HpAsia2 isolates were in "ON" while 20% of HpEurope isolates were genotypically "OFF." The three blood group antigen binding adhesins encoded genes (bab genes) examined and we observed that the most common genotype was (babA/babB/-) found in eight isolates, notably six were HpAsia2 isolates. The babA gene was found in all HpAsia2 isolates but present in only half of the HpEurope isolates. In silico antibiotic susceptibility analysis revealed that 40% of the strains were multi-drug resistant. Mutations associated with resistance to metronidazole, fluoroquinolone, and clarithromycin were detected 90%, 45%, and 5%, respectively, in H pylori strain. In conclusion, it is evident that two populations of H pylori with similar antibiotic profiles are predominant in Bangladesh, and it appears that genotypically the HpAisa2 isolates are potentially more virulent than the HpEurope isolates.

Design of an inhibitor of Helicobacter pylori cholesteryl-α-glucoside transferase critical for bacterial colonization.

BACKGROUND: Fifty percent of the world's population surves as a host for Helicobacter pylori, gastric cancer causing bacteria, that colonizes the gastric region of digestive tract. It has a remarkable capacity to infect the host stomach for the entire lifetime despite an activated host immune response. METHODS: In this study, we have performed the virtual screening analysis of protein-inhibitor binding between the glycosyl transferase enzymes of Helicobacter pylori (CapJ or HP0421) and a corresponding library of inhibitors in the known substrate-binding pockets. We have docked our library of ligands consisting of cholesterol backbone with CapJ protein and identified several ligands' interacting amino acid residues present in active site pocket(s) of the protein. RESULTS: In most of the cases, the ligands showed an interaction with the residues of the same pocket of the enzyme. Top three (03) hits were filtered out from the whole data set, which might act as potent inhibitors of the enzyme-substrate reaction. CONCLUSIONS: This study describes a new possibility by which colonization of H. pylori can be limited. The reported evidence suggests that comprehensive knowledge and wet laboratory validation of these inhibitors are needed in order to develop them as lead molecules.

Genetic diversity of Koala retrovirus env gene subtypes: insights into northern and southern koala populations.

Koala retrovirus (KoRV) is a recently endogenized retrovirus associated with neoplasia and immunosuppression in koala populations. The virus is known to display sequence variability and to be present at varying prevalence in different populations, with animals in southern Australia displaying lower prevalence and viral loads than northern animals. This study used a PCR and next-generation sequencing strategy to examine the diversity of the KoRV env gene in both proviral DNA and viral RNA forms in two distinct populations representative of the 'northern' and 'southern' koala genotypes. The current study demonstrated that the full range of KoRV subtypes is present across both populations, and in both healthy and sick animals. KoRV-A was the predominant proviral subtype in both populations, but there was marked diversity of DNA and RNA subtypes within individuals. Many of the northern animals displayed a higher RNA viral diversity than evident in their proviral DNA, indicating relatively higher replication efficiency of non-KoRV-A subtypes. The southern animals displayed a lower absolute copy number of KoRV than the northern animals as reported previously and a higher preponderance of KoRV-A in individual animals. These discrepancies in viral replication and diversity remain unexplained but may indicate relative protection of the southern population from KoRV replication due to either viral or host factors and may represent an important protective effect for the host in KoRV's ongoing entry into the koala genome.

Mycobacterium tuberculosis DosR regulon gene Rv2004c contributes to streptomycin resistance and intracellular survival.

Tuberculosis (TB) is the deadly infectious disease challenging the public health globally and its impact is further aggravated by co-infection with HIV and the emergence of drug resistant strains of Mycobacterium tuberculosis. In this study, we attempted to characterise the Rv2004c encoded protein, a member of DosR regulon, for its role in drug resistance. In silico docking analysis revealed that Rv2004c binds with streptomycin (SM). Phosphotransferase assay demonstrated that Rv2004c possibly mediates SM resistance through the aminoglycoside phosphotransferase activity. Further, E. coli expressing Rv2004c conferred resistance to 100µM of SM in liquid broth cultures indicating a mild aminoglycoside phosphotransferase activity of Rv2004c. Moreover, we investigated the role of MSMEG_3942 (an orthologous gene of Rv2004c) encoded protein in intracellular survival, its effect on in-vitro growth and its expression in different stress conditions by over expressing it in Mycobacterium smegmatis (M. smegmatis). MSMEG_3942 overexpressing recombinant M. smegmatis strains grew faster in acidic medium and also showed higher bacillary counts in infected macrophages when compared to M. smegmatis transformed with vector alone. Our results are likely to contribute to the better understanding of the involvement of Rv2004c in partial drug resistance, intracellular survival and adaptation of bacilli to stress conditions.

Novel insights into viral infection and oncogenesis from koala retrovirus (KoRV) infection of HEK293T cells.

Koala retrovirus is thought to be an underlying cause of high levels of neoplasia and immunosuppression in koalas. While epidemiology studies suggest a strong link between KoRV and disease it has been difficult to prove causality because of the complex nature of the virus, which exists in both endogenous and exogenous forms. It has been difficult to identify koalas completely free of KoRV, and infection studies in koalas or koala cells are fraught with ethical and technical difficulties, respectively. This study uses KoRV infection of the susceptible human cell line HEK293T and RNAseq to demonstrate gene networks differentially regulated upon KoRV infection. Many of the pathways identified are those associated with viral infection, such as cytokine receptor interactions and interferon signalling pathways, as well as viral oncogenesis pathways. This study provides strong evidence that KoRV does indeed behave similarly to infectious retroviruses in stimulating antiviral and oncogenic cellular responses. In addition, it provides novel insights into KoRV oncogenesis with the identification of a group of histone family genes that are part of several oncogenic pathways as upregulated in KoRV infection.

Genome Dynamics of Vibrio cholerae Isolates Linked to Seasonal Outbreaks of Cholera in Dhaka, Bangladesh.

The temporal switching of serotypes from serotype Ogawa to Inaba and back to Ogawa was identified in Vibrio cholerae O1, which was responsible for seasonal outbreaks of cholera in Dhaka during the period 2015 to 2018. In order to delineate the factors responsible for this serotype transition, we performed whole-genome sequencing (WGS) of V. cholerae O1 multidrug-resistant strains belonging to both the serotypes that were isolated during this interval where the emergence and subsequent reduction of the Inaba serotype occurred. The whole-genome-based phylogenetic analysis revealed clonal expansion of the Inaba isolates mainly responsible for the peaks of infection during 2016 to 2017 and that they might have evolved from the prevailing Ogawa strains in 2015 which coclustered with them. Furthermore, the wbeT gene in these Inaba serotype isolates was inactivated due to insertion of a transposable element at the same position signifying the clonal expansion. Also, V. cholerae isolates in the Inaba serotype dominant clade mainly contained classical ctxB allele and revealed differences in the genetic composition of Vibrioseventh pandemic island II (VSP-II) and the SXT integrative and conjugative element (SXT-ICE) compared to those of Ogawa serotype strains which remerged in 2018. The variable presence of phage-inducible chromosomal island-like element 1 (PLE1) was also noted in the isolates of the Inaba serotype dominant clade. The detailed genomic characterization of the sequenced isolates has shed light on the forces which could be responsible for the periodic changes in serotypes of V. cholerae and has also highlighted the need to analyze the mobilome in greater detail to obtain insights into the mechanisms behind serotype switching.IMPORTANCE The switching of serotype from Ogawa to Inaba and back to Ogawa has been observed temporally in Vibrio cholerae O1, which is responsible for endemic cholera in Bangladesh. The serospecificity is key for effective intervention and for preventing cholera, a deadly disease that continues to cause significant morbidity and mortality worldwide. In the present study, WGS of V. cholerae allowed us to better understand the factors associated with the serotype switching events observed during 2015 to 2018. Genomic data analysis of strains isolated during this interval highlighted variations in the genes ctxB, tcpA, and rtxA and also identified significant differences in the genetic content of the mobilome, which included key elements such as SXT ICE, VSP-II, and PLE. Our results indicate that selective forces such as antibiotic resistance and phage resistance might contribute to the clonal expansion and predominance of a particular V. cholerae serotype responsible for an outbreak.

Koala retrovirus viral load and disease burden in distinct northern and southern koala populations.

Koala retrovirus (KoRV) displays features of both an endogenous and exogenous virus and is linked to neoplasia and immunosuppression in koalas. This study explores the apparent differences in the nature and impact of KoRV infection between geographically and genetically separated "northern" and "southern" koala populations, by investigating the disease status, completeness of the KoRV genome and the proviral (DNA) and viral (RNA) loads of 71 northern and 97 southern koalas. All northern animals were positive for all KoRV genes (gag, pro-pol and env) in both DNA and RNA forms, whereas many southern animals were missing one or more KoRV genes. There was a significant relationship between the completeness of the KoRV genome and clinical status in this population. The proviral and viral loads of the northern population were significantly higher than those of the southern population (P < 0.0001), and many provirus-positive southern animals failed to express any detectable KoRV RNA. Across both populations there was a positive association between proviral load and neoplasia (P = 0.009). Potential reasons for the differences in the nature of KoRV infection between the two populations are discussed.

Genomic and Functional Characterization of Poultry Escherichia coli From India Revealed Diverse Extended-Spectrum ß-Lactamase-Producing Lineages With Shared Virulence Profiles.

Extended-spectrum ß-lactamases (ESBLs) form the most important resistance determinants prevalent worldwide. Data on ESBL-producing Escherichia coli from poultry and livestock are scarce in India. We present data on the functional and genomic characterization of ESBL-producing E. coli obtained from poultry in India. The whole genome sequences of 28 ESBL-producing E. coli were analyzed comprising of 12 broiler chicken E. coli isolates, 11 free-range chicken E. coli isolates, and 5 human extraintestinal pathogenic E. coli. All of the 28 ESBL-producing E. coli isolates were tested for antibiotic susceptibilities, in vitro conjugation, and virulence-associated phenotypic characteristics. A total of 13 sequence types were identified from the poultry E. coli, which included globally successful sequence types such as ST117 (9%), ST131 (4.3%), and ST10 (4.3%). The most common ESBL gene detected in poultry E. coli genomes was bla CTX-M-15 (17%). Also, FIB (73%) and FII (73%) were the most common plasmid replicons identified. Conjugation experiments demonstrated 54 (7/13), 30 (3/10), and 40% (2/5) of broiler, free-range, and human ExPEC E. coli to be able to transfer their ESBL genes, respectively. The in vitro virulence-associated phenotypic tests revealed the broiler, free-range, and human ExPEC isolates to be comparable in biofilm formation, resistance to serum bactericidal activity, adherence, and invasion capabilities. Our overall results showed prevalence of virulence phenotypes among the diverse ESBL-producing E. coli from poultry; while certain E. coli clones from broiler-poultry may indeed have the potential to cause infection in humans.