Bovine clinical mastitis (CM) is one of the most prevalent diseases caused by a wide range of resident microbes. The emergence of antimicrobial resistance in CM bacteria is well-known, however, the genomic resistance composition (the resistome) at the microbiome-level is not well characterized. In this study, we applied whole metagenome sequencing (WMS) to characterize the resistome of the CM microbiome, focusing on antibiotics and metals resistance, biofilm formation (BF), and quorum sensing (QS) along with in vitro resistance assays of six selected pathogens isolated from the same CM samples. The WMS generated an average of 21.13 million reads (post-processing) from 25 CM samples that mapped to 519 bacterial strains, of which 30.06% were previously unreported. We found a significant (P = 0.001) association between the resistomes and microbiome composition with no association with cattle breed, despite significant differences in microbiome diversity among breeds. The in vitro investigation determined that 76.2% of six selected pathogens considered "biofilm formers" actually formed biofilms and were also highly resistant to tetracycline, doxycycline, nalidixic acid, ampicillin, and chloramphenicol and remained sensitive to metals (Cr, Co, Ni, Cu, Zn) at varying concentrations. We also found bacterial flagellar movement and chemotaxis, regulation and cell signaling, and oxidative stress to be significantly associated with the pathophysiology of CM. Thus, identifying CM microbiomes, and analyzing their resistomes and genomic potentials will help improve the optimization of therapeutic schemes involving antibiotics and/or metals usage in the prevention and control of bovine CM.

The SARS‐CoV‐2 coronavirus is responsible for the current COVID‐19 pandemic, with an ongoing toll of over 5 million infections and 333 thousand deaths worldwide within the first 5 months. Insight into the phylodynamics and mutation variants of this virus is vital to understanding the nature of its spread in different climate conditions. The incidence rate of COVID‐19 is increasing at an alarming pace within subtropical Southeast Asian nations with high temperatures and humidity. To understand this spread, we analyzed 444 genome sequences of SARS‐CoV‐2 available on the GISAID platform from 6 Southeast Asian countries. Multiple sequence alignments and maximum likelihood phylogenetic analyses were performed to analyze and characterize the non‐synonymous mutant variants circulating in this region. Global mutation distribution analysis showed that the majority of the mutations found in this region are also prevalent in Europe and North America, and the concurrent presence of these mutations at a high frequency in other countries indicate possible transmission routes. Unique spike protein and non‐structural protein mutations were observed circulating within confined area of a given country. We divided the circulating viral strains into 4 major groups and 3 sub‐groups on the basis of the most frequent non‐synonymous (NS) mutations. Strains with a unique set of 4 co‐evolving mutations were found to be circulating at a high frequency within India, specifically. Group 2 strains characterized by two co‐evolving NS mutants which alter in RdRp (P323L) and spike (S) protein (D614G), were found to be common in Europe and North America. These European and North American variants have rapidly emerged as dominant strains within Southeast Asia, increasing from a 0% prevalence in January to an 81% by May 2020. These variants may have an evolutionary advantage over their ancestral types and could present a large threat to Southeast Asia for the coming winter.

Background We evaluated the impact of low-cost water, sanitation, handwashing (WSH) and child nutrition interventions on enteropathogen carriage in the WASH Benefits cluster-randomized controlled trial in rural Bangladesh. Methods We analyzed 1411 routine fecal samples from children 14±2 months old in the WSH (n = 369), nutrition counseling plus lipid-based nutrient supplement (n = 353), nutrition plus WSH (n = 360), and control (n = 329) arms for 34 enteropathogens using quantitative PCR. Outcomes included the number of co-occurring pathogens; cumulative quantity of four stunting-associated pathogens; and prevalence and quantity of individual pathogens. Masked analysis was by intention-to-treat. Results 326 (99.1%) control children had one or more enteropathogens detected (mean 3.8±1.8). Children receiving WSH interventions had lower prevalence and quantity of individual viruses than controls (prevalence difference for norovirus: -11% [95% confidence interval [CI], -5 to -17%]; sapovirus: -9% [95%CI, -3 to -15%]; and adenovirus 40/41: -9% [95%CI, -2 to - 15%]). There was no difference in bacteria, parasites, or cumulative quantity of stunting-associated pathogens between controls and any intervention arm. Conclusions WSH interventions were associated with fewer enteric viruses in children aged 14 months. Different strategies are needed to reduce enteric bacteria and parasites at this critical young age.

AbstractThe avian pathogenic Escherichia coli (APEC) strains are the chief etiology of avian colibacillosis worldwide. The present study investigated the circulating phylotypes, existence of virulence genes (VGs) and antimicrobial resistance (AMR) in 392 APEC isolates, obtained from 130 samples belonged to six farms using both phenotypic and PCR-based molecular approaches. Congo red binding (CRB) assay confirmed 174 APEC isolates which were segregated into 10, 9 and 8 distinctive genotypes by RAPD assay (discriminatory index, D=0.8707), BOX-PCR (D=0.8591) and ERIC-PCR (D=0.8371), respectively. The combination of three phylogenetic markers (chuA, yjaA and DNA fragment TspE4.C2) classified APEC isolates into B23 (37.36%), A1 (33.91%), D2 (11.49%), B22 (9.20%) and B1 (8.05%) phylotypes. Majority of the APEC isolates (75-100%) harbored VGs (ial, fimH, crl, papC and cjrC), and of them, VGs (papC and cjrC) and phylotypes (D2 and B2) of APEC had significant (p=0.004) association with colibacillosis. Phylogenetic analysis showed two distinct clades (Clade A and Clade B) of APEC where Clade A had 98-100.0% similarity with E. coli APEC O78 and E. coli EHEC strains, and Clade B had closest relationship with E. coli O169:H41 strain. Interestingly, phylogroups B2 and D2 were found in the APEC strains of both clades while the strains from phylogroups A1 and B1 were found in clade A only. In this study, 81.71% of the isolates were biofilm formers and possessed plasmids of varying ranges (1.0 to 54 kb). In vitro antibiogram profiling revealed that 100.0% isolates were resistant to ≥3 antibiotics, of which61.96%, 55.24, 53.85, 51.16 and 45.58 % isolates in phylotypes B1, D2, B22, B23 and A1, respectively were these antimicrobials. The resistance patterns varied among different phylotypes, notably in phylotype B22 showing the highest resistance to ampicillin (90.91%), nalidixic acid (90.11%), tetracycline (83.72%) and nitrofurantoin (65.12%). Correspondence analysis also showed significant correlation of phylotypes with CRB (p=0.008), biofilm formation (p=0.02), drug resistance (p=0.03) and VGs (p=0.06). This report demonstrated that B2 and A1 phylotypes are dominantly circulating APEC phylotypes in Bangladesh; however, B2 and D2 are strongly associated with the pathogenicity. A high prevalence of antibiotic resistant APEC strains from different phylotypes suggest to use of organic antimicrobial compounds, and/or metals, and the rotational use of antibiotics in the poultry farms in Bangladesh.

The novel coronavirus, SARS-CoV-2, has caused the most unfathomable pandemic in the history of humankind. Bangladesh is also a victim of this critical situation. To investigate the genomic features of the pathogen from Bangladesh, the first complete genome of the virus has very recently been published. Therefore, long-awaited ques

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is continuously spreading worldwide at an unprecedented scale in 2020. Within the first six months of the COVID-19 pandemic, it has evolved into six clades according to GISAID where three (G, GH, and GR) are now globally prevalent (>75%). Here we report the prevalence of these dominant clades, both individually and in combination, with disease progression and death-case scenario that leads to infer fitness of the SARS-CoV-2 by compromising its virulence. Unlike G or GH clades, the GR clade strains represent a significant negative association with the death-case ratio (R= -0.558, p=0.019). Docking analysis revealed the molecular scenario behind more infectiousness of S protein D614G mutation and reasoned more favorable binding of G614 with the elastase-2. Viral RNA-dependent-RNA-polymerase (RdRp) mutation p.P323L facilitated significantly higher (p<0.0001) genome-wide mutations because more flexible RdRp (mutant)-NSP8 interaction may accelerate replication. Superior RNA stability and structural variation at NSP3:C241T might change the protein’s conformation with a speculated impact on 5'UTR, nucleocapsid, and replication complex interactions. Another silent 5'UTR:C241T mutation might affect translational efficiency and viral packaging. These G-featured coevolving mutations might together increase the viral load, quicker cell death, and potentially a stronger immune response within the host, hence can modulate intra-host genomic plasticity. In addition, viroporin ORF3a:p.Q57H mutation of GH-clade prevents ion permeability by constricting the channel pore more tightly due to additional ionic interaction with the cysteine (C81) of transmembrane-domain-2, which possibly reduces viral release and immune response. GR strains (four G clade mutations with N:p.RG203-204KR) would have maintained more stability with stronger RNA interaction, a flexible linker region, and the molecular effect of hypo-phosphorylation at SR-stretch. These empirical assumptions need further retrospective and prospective studies to understand detailed molecular and evolutionary events featuring the fitness and virulence of SARS-CoV-2.

Clinical Waste Water (CWW) possesses the risks of spreading antibiotic resistant bacteria in the environment. In Bangladesh, liquid discharge is directly released into the municipal sewage system and pollutes the surroundings water bodies/rivers. Liquid samples were collected from the three CWW discharge sites connected to DMCH (Dhaka Medical College Hospital) and from one control group non-connected to DMCH. About 32, 30 and 26 times higher ciprofloxacin, cefixime and multi-drug resistant bacterial count were found in CWW over control samples. Among the isolates, two Escherichia spp. isolates, denoted 26N and 28N, were found to be resistant against fluoroquinolone (MBC of Ciprofloxacin > 1024 μg/ml), cephalosporin, glycopeptide, monobactam, penicillin, tetracycline, rifampicin, macrolides, sulfonamide and nitrofuran classes of drugs and were also ESBL positive through phenotypic assay. Plasmid curing experiment detected possible plasmid mediated resistance of fluoroquinolone, cephalosporin, tetracycline, lincosamide and nitrofuran classes of antibiotics. Phylogenetically, isolate 26N and 28N were characterized as Escherichia coli and Escherichia fergusonii. These MDR and ESBL positive bacteria are potent to disseminate resistant determinants in the surrounding environments.

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