Drug Resistance Determinants in Clinical Isolates of Enterococcus faecalis in Bangladesh: Identification of Oxazolidinone Resistance Gene optrA in ST59 and ST902 Lineages.

Enterococcus faecalis is one of the major causes of urinary tract infection, showing acquired resistance to various classes of antimicrobials. The objective of this study was to determine the prevalence of drug resistance and its genetic determinants for E. faecalis clinical isolates in north-central Bangladesh. Among a total of 210 E. faecalis isolates, isolated from urine, the resistance rates to erythromycin, levofloxacin, and gentamicin (high level) were 85.2, 45.7, and 11.4%, respectively, while no isolates were resistant to ampicillin, vancomycin and teicoplanin. The most prevalent resistance gene was erm(B) (97%), and any of the four genes encoding aminoglycoside modifying enzyme (AME) were detected in 99 isolates (47%). The AME gene aac(6')-Ie-aph(2")-Ia was detected in 46 isolates (21.9%) and was diverse in terms of IS256-flanking patterns, which were associated with resistance level to gentamicin. Tetracycline resistance was ascribable to tet(M) (61%) and tet(L) (38%), and mutations in the quinolone resistance-determining region of both GyrA and ParC were identified in 44% of isolates. Five isolates (2.4%) exhibited non-susceptibility to linezolide (MIC, 4 µg/mL), and harbored the oxazolidinone resistance gene optrA, which was located in a novel genetic cluster containing the phenicol exporter gene fexA. The optrA-positive isolates belonged to ST59, ST902, and ST917 (CC59), while common lineages of other multiple drug-resistant isolates were ST6, ST28, CC16, and CC116. The present study first revealed the prevalence of drug resistance determinants of E. faecalis and their genetic profiles in Bangladesh.

Molecular Epidemiological Characterization of Methicillin-Susceptible and -Resistant Staphylococcus aureus Isolated from Skin and Soft Tissue Infections in Bangladesh.

Genetic background and molecular characteristics of Staphylococcus aureus collected from patients with skin and soft tissue infections were studied in the North-Central region of Bangladesh from 2015 to 2016. Among 430 clinical isolates, methicillin-resistant S. aureus (MRSA) accounted for 31% having SCCmec type IV (73%) and V (14%), and belonged mostly to coagulase (coa) genotypes IIa, IIIa, IVb, and XIa, while dominant coa type in methicillin-susceptible S. aureus (MSSA) was IIIa, followed by Va, IIa, and VIa. Panton-Valentine Leukocidin genes (pvl) were detected at higher rate in MSSA (54%) than in MRSA (24%). Based on multilocus sequence typing, pvl-positive MRSA isolates were classified into clonal complex 88 (CC88) (ST88, ST2884, ST4345), CC6 (ST6, ST4350), and CC1 (ST1, ST772), while pvl-negative MRSA into CC5, CC22, CC80, CC121, and CC672. The pvl-negative ST80 MRSA isolates had SCCmec-IVa (agr-III/coa-XIc, etd/edinB-positive, fusB-negative), indicating that they belong to the novel CC80 clade related to the European community-acquired MRSA clone. Among MSSA, genotypes ST121/spa-t645/coa-Va and ST2884 (CC88)/spa-t2393/coa-IIIa were identified in both pvl-positive and negative isolates, and all the ST772 isolates harbored pvl. All the ST121 isolates had a variant of elastin-binding protein gene (ebpS-v) with internal 180-nucleotide deletion. The present study suggested that CC88 (ST88, ST2884) and ST772 are the putative dominant lineages of pvl-positive MRSA/MSSA, while novel CC80 clade is one of the main pvl-negative MRSA lineages distributed endemically in Bangladesh.

Prevalence and Molecular Epidemiology of Clinical Isolates of Escherichia coli and Klebsiella pneumoniae Harboring Extended-Spectrum Beta-Lactamase and Carbapenemase Genes in Bangladesh.

Spread of Gram-negative bacteria producing extended-spectrum beta-lactamases (ESBLs) and carbapenemases constitutes a growing challenge in control of bacterial infections. In this study, prevalence and genetic characteristics of Escherichia coli and Klebsiella pneumoniae harboring ESBL and/or carbapenemase genes, with other beta-lactamase/resistance genes, were investigated for a total of 375 clinical isolates in Mymensingh located in north-central Bangladesh. The major ESBL gene was blaCTX-M-1 group, which was detected in 33.9% and 51.4% of E. coli and K. pneumoniae, respectively, with CTX-M-15 gene being dominant. SHV-type beta-lactamase genes, including newly identified alleles (SHV-201 and SHV-202) were detected at higher rate in K. pneumoniae (27%). Nine isolates of E. coli (3.9%) harbored carbapenemase genes; blaNDM-1 (phylogenetic group A-sequence type 2104 (A-ST2104), B2-ST73), blaNDM-5 (A-ST167, B2-ST38/ST2659-related STs), and blaNDM-7 (B1-ST101/ST224, D-ST6682). AmpC beta-lactamase genes (blaCMY-2 and blaCMY-42) were detected in E. coli, which mostly harbored blaCTX-M-15 and plasmid-mediated quinolone resistance (PMQR) determinants (aac6'-Ib-cr, qnrB, qnrS, qepA, and oqxAB). A new CMY allele (CMY-160) belonging to CMY-2 group was identified in phylogenetic group D E. coli. Among K. pneumoniae, carbapenemase gene was detected in three isolates (2%); blaNDM-1 in ST11 and ST1322, and blaOXA-181 in ST43 isolate. As well as higher rate of aac6'-Ib-cr in K. pneumoniae (39%), PMQR gene oqxAB was also commonly found among isolates analyzed. These findings indicated spread of blaNDM genes to diverse E. coli clones and emergence of blaOXA-181 in K. pneumoniae, with increased prevalence of ESBLs represented by CTX-M-15 in Bangladesh.

Distribution of two distinct rotavirus B (RVB) strains in the north-central Bangladesh and evidence for reassortment event among human RVB revealed by whole genomic analysis.

Human rotavirus B (RVB), a rare cause of diarrhea in several Asian countries, has been reported to be genetically highly conserved. However, 14 RVB strains with two distinct RNA electropherotypes E1 and E2 (11 and 3 strains, respectively) were detected in adult patients with diarrhea, in Mymensingh in the north-central Bangladesh in 2014. In this study, VP7 gene sequences of all the 14 strains and nearly full-length sequences of all the 11 RNA segments of four RVB (two strains each representing E1 and E2 types) were determined and analyzed phylogenetically. For all the gene segments, sequence identities among strains with the same RNA pattern were higher (99%-100%) than those between strains with different RNA patterns (94-98%). Although all the gene segments of RVB strains were grouped into Indian-Bangladeshi lineage, VP1-3, VP6, VP7, NSP1, NSP2 and NSP5 genes of strains with E1 and E2 types were assigned to distinct sublineages S1 and S2, respectively. E1-strains clustered with Bangladeshi RVB strains reported previously (e.g., Bang117), while E2-strains with those from India (e.g., NIV-1048101), Myanmar, and Nepal. In contrast, VP4, NSP3 and NSP4 genes of both E1 and E2 RVB strains were classified into sublineage S2. These findings indicated that two genetically distinct RVB strains were simultaneously circulating in Mymensingh, Bangladesh. RVB strains with E1 electropherotype were suggested to be reassortants acquiring three gene segments (VP4, NSP3 and NSP4 genes) from the foreign RVB in the genetic background of indigenous Bangladeshi RVB represented by the strain Bang117.

Molecular Detection of Rickettsia felis in Humans, Cats, and Cat Fleas in Bangladesh, 2013-2014.

High prevalence of Rickettsia felis in patients with fever of unknown origin was revealed in the north-central Bangladesh from 2012 to 2013. Subsequently, in this study, prevalence of R. felis in cats and cat fleas (Ctenocephalides felis), together with febrile patients, was studied by PCR detection of 17 kDa antigen gene and DNA sequencing. R. felis was detected in 28% (28/100) and 21% (14/68) of cat blood and cat flea samples, respectively, whereas 42% (21/50) of patients were positive for R. felis. R. felis-positive cat fleas were detected at significantly higher rate on R. felis-positive cats. The results suggested a potential role of cats and cat fleas for transmission of R. felis to humans in Bangladesh.

Whole genomic analysis of G2P[4] human Rotaviruses in Mymensingh, north-central Bangladesh.

Rotavirus A (RVA) is a dominant causative agent of acute gastroenteritis in children worldwide. G2P[4] is one of the most common genotypes among human rotavirus (HRV) strains, and has been persistently prevalent in South Asia including Bangladesh. In the present study, whole genome sequences of a total of 16 G2P[4] HRV strains (8 strains each in 2010 and 2013) detected in Mymensingh, north-central Bangladesh were determined. These strains had typical DS-1-like genotype constellation. Most of gene segments from DS-1 genogroup exhibited high level sequence identities to each other (>98%), while slight diversity was observed for VP1, VP3, and NSP4 genes. By phylogenetic analysis, individual RNA segments were classified into one (V) or two-three lineages (V-VI or V-VII). In terms of lineages (sublineages) of 11 gene segments, the 16 Bangladeshi strains could be further classified into four clades (A-D) containing 8 lineage constellations, revealing the presence of three clades (A-C) with three lineage constellations in 2010, and a single clade (D) with four constellations in 2013. Therefore, co-existence of multiple G2P[4] HRV strains with different lineage constellations, and change in clades for the study period were demonstrated. Although amino acids in the antigenic regions on VP7 and VP4 were mostly identical to those of global G2P[4] strains after 2000, VP4 of clade D RVAs in 2013 had alanine and proline at positions 88 and 114, respectively, which are novel substitutions compared with recent global G2P[4] strains. Replacement of lineage constellations associated with unique amino acid changes in the antigenic region in VP4 suggested continuous genetic evolutionary state for emerging new G2P[4] rotavirus strains in Bangladesh.