Novel insights into the co-selection of metal-driven antibiotic resistance in bacteria: a study of arsenic and antibiotic co-exposure.

The simultaneous development of antibiotic resistance in bacteria due to metal exposure poses a significant threat to the environment and human health. This study explored how exposure to both arsenic and antibiotics affects the ability of an arsenite oxidizer, Achromobacter xylosoxidans CAW4, to transform arsenite and its antibiotic resistance patterns. The bacterium was isolated from arsenic-contaminated groundwater in the Chandpur district of Bangladesh. We determined the minimum inhibitory concentration (MIC) of arsenite, cefotaxime, and tetracycline for A. xylosoxidans CAW4, demonstrating a multidrug resistance (MDR) trait. Following this determination, we aimed to mimic an environment where A. xylosoxidans CAW4 was exposed to both arsenite and antibiotics. We enabled the strain to grow in sub-MIC concentrations of 1 mM arsenite, 40 µg/mL cefotaxime, and 20 µg/mL tetracycline. The expression dynamics of the arsenite oxidase (aioA) gene in the presence or absence of antibiotics were analyzed. The findings indicated that simultaneous exposure to arsenite and antibiotics adversely affected the bacteria's capacity to metabolize arsenic. However, when arsenite was present in antibiotics-containing media, it promoted bacterial growth. The study observed a global downregulation of the aioA gene in arsenic-antibiotic conditions, indicating the possibility of increased susceptibility through co-resistance across the entire bacterial population of the environment. This study interprets that bacterial arsenic-metabolizing ability can rescue the bacteria from antibiotic stress, further disseminating environmental cross-resistance. Therefore, the co-selection of metal-driven antibiotic resistance in bacteria highlights the need for effective measures to address this emerging threat to human health and the environment.

Microbiome signature and diversity regulates the level of energy production under anaerobic condition.

The microbiome of the anaerobic digester (AD) regulates the level of energy production. To assess the microbiome diversity and composition in different stages of anaerobic digestion, we collected 16 samples from the AD of cow dung (CD) origin. The samples were categorized into four groups (Group-I, Group-II, Group-III and Group-IV) based on the level of energy production (CH4%), and sequenced through whole metagenome sequencing (WMS). Group-I (n = 2) belonged to initial time of energy production whereas Group-II (n = 5), Group-III (n = 5), and Group-IV (n = 4) had 21-34%, 47-58% and 71-74% of CH4, respectively. The physicochemical analysis revealed that level of energy production (CH4%) had significant positive correlation with digester pH (r = 0.92, p < 0.001), O2 level (%) (r = 0.54, p < 0.05), and environmental temperature (°C) (r = 0.57, p < 0.05). The WMS data mapped to 2800 distinct bacterial, archaeal and viral genomes through PathoScope (PS) and MG-RAST (MR) analyses. We detected 768, 1421, 1819 and 1774 bacterial strains in Group-I, Group-II, Group-III and Group-IV, respectively through PS analysis which were represented by Firmicutes, Bacteroidetes, Proteobacteria, Actinobacteria, Spirochaetes and Fibrobacteres phyla (> 93.0% of the total abundances). Simultaneously, 343 archaeal strains were detected, of which 95.90% strains shared across four metagenomes. We identified 43 dominant species including 31 bacterial and 12 archaeal species in AD microbiomes, of which only archaea showed positive correlation with digester pH, CH4 concentration, pressure and temperature (Spearman correlation; r > 0.6, p < 0.01). The indicator species analysis showed that the species Methanosarcina vacuolate, Dehalococcoides mccartyi, Methanosarcina sp. Kolksee and Methanosarcina barkeri were highly specific for energy production. The correlation network analysis showed that different strains of Euryarcheota and Firmicutes phyla exhibited significant correlation (p = 0.021, Kruskal-Wallis test; with a cutoff of 1.0) with the highest level (74.1%) of energy production (Group-IV). In addition, top CH4 producing microbiomes showed increased genomic functional activities related to one carbon and biotin metabolism, oxidative stress, proteolytic pathways, membrane-type-1-matrix-metalloproteinase (MT1-MMP) pericellular network, acetyl-CoA production, motility and chemotaxis. Importantly, the physicochemical properties of the AD including pH, CH4 concentration (%), pressure, temperature and environmental temperature were found to be positively correlated with these genomic functional potentials and distribution of ARGs and metal resistance pathways (Spearman correlation; r > 0.5, p < 0.01). This study reveals distinct changes in composition and diversity of the AD microbiomes including different indicator species, and their genomic features that are highly specific for energy production.

Evidence of combined effect of amino acid substitutions within G-H and B-C loops of VP1 conferring serological heterogeneity in foot-and-mouth disease virus serotype A.

Foot-and-mouth disease virus (FMDV) serotype A exhibits a higher degree of genetic and antigenic diversity resulting in frequent vaccine failure due to serological mismatch between the vaccine and heterologous strains. Currently, knowledge on the molecular basis of antigenic relationships among the FMDVs is limited; nevertheless, intratype antigenic variation due to mutation(s) is widely considered as the main hurdle to appropriate FMD vaccine development. Here, we studied genetic and antigenic variations of four FMDV serotype A isolates, BAN/GA/Sa-197/2013 (BAN-197), BAN/CH/Sa-304/2016 (BAN-304), BAN/DH/Sa-307/2016 (BAN-307) and BAN/DH/Sa-310/2017 (BAN-310) circulating in Bangladesh during 2013-2017. Initially, antigenic relationships (r1 -values) of the field isolates were evaluated by the two-dimensional microneutralization test (2D-MNT) using the hyperimmune antisera raised in cattle against the vaccine strain, BAN-304. Interesingly, the results showed protective serological cross-reactivity (r1 -values > 0.4) between the vaccine strain and the field isolates, BAN-307 and BAN-310, except BAN-197 that substantially mismatched (r1  = 0.129 ± 0.043) with the BAN-304. Although VP1-based phylogeny grouped all the isolates within the same sublineage C (a subgroup of VP3Δ59 variant) under the lineage A/ASIA/G-VII, strikingly, computational analyses of the viral capsid proteins demonstrated significant deviation at the VP1 G-H loop of BAN-197 from the vaccine strain, while VP(2-4) of both isolates were structurally conserved. To bridge the gap of how the distortion of the G-H loop and consequent antigenic hetergeneity occurred in BAN-197, we performed in silico combinatorial substitutions of the VP1 mutant amino acids (aa) of BAN-197 with the respective residues in BAN-304. Remarkably, our analyses revealed that two substitutions of distantly located aa at B-C (T48I:threonine â†’ isoleucine) and G-H (A143V:alanine â†’ valine) loops, in combination, distorted the VP1 G-H loop. Overall, this work contributes to understanding the molecular basis of antigenic relationships operating in serotype A FMDVs and the selection of suitable vaccine strain(s) for effective prophylaxis of FMD based on VP1-based analyses.

Epitope-based chimeric peptide vaccine design against S, M and E proteins of SARS-CoV-2, the etiologic agent of COVID-19 pandemic: an in silico approach.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the etiologic agent of the ongoing pandemic of coronavirus disease 2019 (COVID-19), a public health emergency of international concerns declared by the World Health Organization (WHO). An immuno-informatics approach along with comparative genomics was applied to design a multi-epitope-based peptide vaccine against SARS-CoV-2 combining the antigenic epitopes of the S, M, and E proteins. The tertiary structure was predicted, refined and validated using advanced bioinformatics tools. The candidate vaccine showed an average of ≥90.0% world population coverage for different ethnic groups. Molecular docking and dynamics simulation of the chimeric vaccine with the immune receptors (TLR3 and TLR4) predicted efficient binding. Immune simulation predicted significant primary immune response with increased IgM and secondary immune response with high levels of both IgG1 and IgG2. It also increased the proliferation of T-helper cells and cytotoxic T-cells along with the increased IFN-γ and IL-2 cytokines. The codon optimization and mRNA secondary structure prediction revealed that the chimera is suitable for high-level expression and cloning. Overall, the constructed recombinant chimeric vaccine candidate demonstrated significant potential and can be considered for clinical validation to fight against this global threat, COVID-19.

Development and serology based efficacy assessment of a trivalent foot-and-mouth disease vaccine.

Foot-and-mouth disease (FMD) is a highly contagious disease of cloven-hoofed animals throughout the world. The endemicity of this disease in Bangladesh has been causing high economic loss and an impediment to the full potential surge of livestock industries. In Bangladesh, vaccination using imported or locally produced FMD vaccines is the existing practice of controlling the disease, although vaccine failure cases are very common. Hence, to address the problem, the present study was envisaged to develop an effective FMD vaccine tailored to the circulating indigenous foot-and-mouth disease virus (FMDV) strains. Three local circulating FMDVs O/BAN/TA/Dh-301/2016 (MK088170.1), A/BAN/CH/Sa-304/2016 (MK088171.1) and Asia1/BAN/DH/Sa-318/2018 (MH457186.1) isolates were selected as vaccine strains based on recent epidemiology, genetic and antigenic analyses. These serotype O, A and Asia1 vaccine strains showed strong antigenic relationship (r1 > 0.3) with 100% to 75% of the respective circulating viruses. The candidate viruses were successfully inactivated by 3.0 mM binary ethylenimine within 7-10 h after the onset of inactivation. Extrapolation of inactivation kinetics confirmed < 1 log10 TCID50 in a 10000-liter batch liquid preparation after 24 h inactivation cycle. The inactivated virus particles were significantly (p < 0.05) concentrated and the trivalent vaccine was formulated using 6 µg per dose per serotype antigen payload. The trivalent vaccine was administered in divided doses in different groups of cattle. All doses of the vaccine elicited significantly (p < 0.05) higher levels of antibodies as early as 14-day post-vaccination (dpv) and peak antibody titers were achieved in 28 dpv. The 'full dose' (6.0 µg per dose per serotype) vaccine elicited antibody titers expected to confer protection in 100% cattle of the respective group and maintained such level of antibodies beyond 180 dpv. Thus, the trivalent FMD vaccine prepared with 6.0 µg antigen per dose per serotype of the selected candidate viruses will confer protection against circulating FMDVs of Bangladesh and its neighboring countries.

Complete genome sequence of a potential foot-and-mouth disease virus serotype O vaccine strain from Bangladesh.

One of the six sublineages of the dominant O/ME-SA/Ind2001 lineage of foot-and-mouth disease virus (FMDV), Ind2001BD1 has already spread throughout 14 countries, including Bangladesh. Here, we report the complete genome sequence of the potential serotype O vaccine strain BAN/TA/Dh-301/2016, which has been shown to provide protection against all the circulating serotype O viruses in Bangladesh. The viral genome is 8,211 nucleotide (nt) long with an open reading frame (ORF) of 6999 nt. The ORF is flanked by a 1098-nt-long 5'-UTR and a 114-nt-long 3'-UTR. Compared to the Indian FMDV serotype O vaccine strain O/India/R2/75 (AF204276), ten mutations were identified in the major antigenic sites of BAN/TA/Dh-301/2016 (MK088170.1).

Emergence of novel lineage of foot-and-mouth disease virus serotype Asia1 BD-18 (G-IX) in Bangladesh.

Foot-and-mouth disease virus (FMDV) is a highly evolutionary divergent pathogen causing great economic havoc in many countries. Among its seven existing serotypes, Asia1 is the least divergent with a single topotype both genetically and antigenically. It is reported sporadically in Indian subcontinent and was classified under lineage G-VIII. In 2018, serotype Asia1 re-emerged in Bangladesh after 2013, along with circulation of a novel serotype Asia1 BD-18 (G-IX) lineage. VP1 phylogeny and sequence variation clearly demonstrated the novel strains which was estimated to have at least >5% nucleotide divergence with distinct clade formation. Also, the Bayesian phylogeographic inferences traced back to the origin time of lineage G-IX in early 2017 and a possible origin in Bangladesh. Mutational analysis considering established eight lineages revealed that the virus strains belonged to lineage G-IX contained a unique mutation at 44 position in the B-C loop region of VP1. Inappropriate vaccination and inefficient outbreak surveillance possibly contributed to the current episode of emergence. Therefore, active surveillance and continued vigilance are essential to assess and timely detect the occurrence, extent and distribution of this novel Asia1 strains in Bangladesh and the neighbouring countries.

Near-Complete Genome Sequence of a Potential Foot-and-Mouth Disease Virus Serotype A Vaccine Strain Isolated from Bangladesh.

The near-complete genome sequence of foot-and-mouth disease virus (FMDV) serotype A potential vaccine strain BAN/CH/Sa-304/2016 is reported here. Its genome revealed antigenic heterogeneity with the current Indian vaccine strain IND40/00, with four amino acid substitutions in antigenically critical sites of the VP1 protein.

Near-Complete Genome Sequence of a Representative Strain within a Rare Foot-and-Mouth Disease Virus O/ME-SA/Ind2001BD2 Sublineage from Bangladesh.

The near-complete genome sequence of a foot-and-mouth disease virus (FMDV), strain O/ME-SA/Ind2001BD2, isolated exclusively from Bangladesh, is reported here. Amino acid substitutions at critical antigenic sites of the capsid were identified compared to the surface proteins of existing vaccine strain O/India/R2/75 and contemporary FMDV serotype O isolates of Bangladesh.

Adaptation of metal and antibiotic resistant traits in novel ß-Proteobacterium Achromobacter xylosoxidans BHW-15.

Chromosomal co-existence of metal and antibiotic resistance genes in bacteria offers a new perspective to the bacterial resistance proliferation in contaminated environment. In this study, an arsenotrophic bacterium Achromobacter xylosoxidans BHW-15, isolated from Arsenic (As) contaminated tubewell water in the Bogra district of Bangladesh, was analyzed using high throughput Ion Torrent Personal Genome Machine (PGM) complete genome sequencing scheme to reveal its adaptive potentiality. The assembled draft genome of A. xylosoxidans BHW-15 was 6.3 Mbp containing 5,782 functional genes, 1,845 pseudo genes, and three incomplete phage signature regions. Comparative genome study suggested the bacterium to be a novel strain of A. xylosoxidans showing significant dissimilarity with other relevant strains in metal resistance gene islands. A total of 35 metal resistance genes along with arsenite-oxidizing aioSXBA, arsenate reducing arsRCDAB, and mercury resistance merRTPADE operonic gene cluster and 20 broad range antibiotic resistance genes including ß-lactams, aminoglycosides, and multiple multidrug resistance (MDR) efflux gene complex with a tripartite system OM-IM-MFP were found co-existed within the genome. Genomic synteny analysis with reported arsenotrophic bacteria revealed the characteristic genetic organization of ars and mer operonic genes, rarely described in ß-Proteobacteria. A transposon Tn21 and mobile element protein genes were also detected to the end of mer (mercury) operonic genes, possibly a carrier for the gene transposition. In vitro antibiotic susceptibility assay showed a broad range of resistance against antibiotics belonging to ß-lactams, aminoglycosides, cephalosporins (1st, 2nd, and 3rd generations), monobactams and even macrolides, some of the resistome determinants were predicted during in silico analysis. KEGG functional orthology analysis revealed the potential of the bacterium to utilize multiple carbon sources including one carbon pool by folate, innate defense mechanism against multiple stress conditions, motility, a proper developed cell signaling and processing unit and secondary metabolism-combination of all exhibiting a robust feature of the cell in multiple stressed conditions. The complete genome of the strain BHW-15 stands as a genetic basis for the evolutionary adaptation of metal and the antibiotic coexistence phenomenon in an aquatic environment.

Circulatory white spot syndrome virus in South-West region of Bangladesh from 2014 to 2017: molecular characterization and genetic variation.

White Spot Syndrome Virus (WSSV), the etiological agent of White Spot Disease (WSD) is a major impediment for shrimp aquaculture in the worldwide. A critical threshold level of WSSV load in infected shrimp is an important trait for disease manifestation and WSSV transmission in cultured shrimp and subsequently make outbreaks. The present study investigated 120 naturally infected cultured shrimp samples by SYBR Green based qPCR assay for WSD diagnosis and quantification of WSSV load. Among them, 94 samples resulted a variable count of WSSV load ranging from 2.1 × 108 to 2.64 × 1014 copies/g of shrimp tissue. The severity of WSSV infection was assessed based on the established critical threshold load of WSSV in shrimp tissue. Compared to the established critical threshold value of WSSV load in shrimp tissue, our findings showed the horrifying scenario of the severity of WSSV infection in cultured shrimps of Bangladesh that was found to be above the critical limit to initiate an outbreak in the Bangladeshi shrimp aquaculture industry. The latest phylogenetic pattern was altered from the former monophyletic history among WSSVs of Bangladesh due to a variation at 500th nucleotide of VP28 coding gene. Viruses characterized from recent outbreaks in 2015 and 2017 displayed amino acid substitution at position 167 (G→E) on the surface of VP28 protein which has demonstrated the probable replacement of indigenous virus pool. Therefore, it is imperative to take initiative for the management and prevention of WSSV outbreak to sustain shrimp aquaculture in South-West region of Bangladesh.

Complete Genome Sequence of the Circulatory Foot-and-Mouth Disease Virus Serotype Asia1 in Bangladesh.

The complete genome sequence of foot-and-mouth disease virus (FMDV) serotype Asia1 isolated from Bangladesh is reported here. Genome analysis revealed amino acid substitutions in the VP1 antigenic region and deletions in both the 5' and 3' untranslated regions (UTRs) compared to the genome of the existing vaccine strain (GenBank accession no. AY304994).

Complete Genome Sequence of Pig-Originated Foot-and-Mouth Disease Virus Serotype O from Bangladesh.

In this article, we document the first pig-isolated complete genome sequence of foot-and-mouth disease virus type O in Bangladesh. The complete viral genome revealed a potential serotypic recombination at the 5' untranslated region (UTR). Conventional amino acid deletion was lacking in 3A region, and antigenic heterogeneity to circulatory type O existed within the VP1 region.

Complete Genome Sequence of Foot-and-Mouth Disease Virus Type A Circulating in Bangladesh.

The complete genome sequence of a foot-and-and mouth disease virus (FMDV) type A strain (BAN/GA/Sa-197/2013), isolated from Gazipur in Bangladesh, revealed an 84-nucleotide insertion within the 5'-untranslated region (UTR), a lengthened poly(C) tract, and amino acid substitutions at the VP1 region compared to the available genome sequence of the vaccine strain (GenBank accession no. HM854025).

Complete Genome Sequence of Foot-and-Mouth Disease Virus Serotype O Isolated from Bangladesh.

Foot-and-mouth disease (FMD) is a highly infectious enzootic disease caused by FMD virus. The complete genome sequence of a circulatory FMD virus (FMDV) serotype O isolated from Natore, Bangladesh, is reported here. Genomic analysis revealed antigenic heterogeneity within the VP1 region, a fragment deletion, and insertions at the 5' untranslated region (UTR) and 3A region compared to the genome of the available vaccine strain.