Genetic characterization of methicillin-resistant / susceptible Staphylococcus aureus (MRSA/MSSA) and Staphylococcus argenteus clinical isolates in Bangladesh: Dominance of ST6-MRSA-IV/t304 and detection of cfr/fexA in ST8-MSSA/t008.

Objectives: Coagulase-positive staphylococcus (CoPS), represented by Staphylococcus aureus, is a major cause of infections in humans. This study aimed to investigate molecular epidemiological characteristics, antimicrobial resistance, and their trends of CoPS in Bangladesh. Methods: Clinical isolates of CoPS were collected from two medical institutions in Bangladesh for a 2-year period and analyzed for their species, genotypes, virulence factors, antimicrobial susceptibility, and resistance determinants. Results: 172 CoPS isolates collected were identified as S. aureus or S. argenteus (170 and two, respectively). Methicillin-resistant S. aureus (MRSA) accounted for 36% (n = 61), having Staphylococcal cassette chromosome mec (SCCmec)-IV (82%) or V (18%). Panton-Valentine leukocidin (PVL) genes were detected at higher rate in methicillin-susceptible S. aureus (MSSA) (62%) than MRSA (26%). MRSA comprised 11 STs, including a dominant type ST6 (46%) associated with mostly SCCmec-IVa/spa-t304, and one isolate had genetic features of the USA300 clone (ST8/SCCmec-IVa/coa-IIIa/spa-t008/ACME-I/ΦSa2USA). STs of CC1, CC88, and CC398 were common in MSSA, with CC88 showing the highest PVL-positive rate. One MSSA isolate (ST8/spa-t008) harbored fexA and cfr showing susceptibility to linezolid. S. argenteus was methicillin-susceptible and belonged to ST2250/coa-XId. Conclusions: Genetic characteristics of current MRSA/MSSA in Bangladesh were revealed, with first identification of S. argenteus at low prevalence.

Genetic Characterization of the Dengue Virus Type 3 Genotype I Prevailing in Dhaka, Bangladesh, 2021.

Background: In Bangladesh, dengue has been prevalent since its resurgence in 2018, and the dominant causative virus in 2019 was considered dengue virus serotype 3 (DENV-3). However, limited information is available for DENV serotype/genotype circulating after 2020. Materials and Methods: Viral RNA was extracted from NS1 antigen-positive blood samples of febrile patients in Dhaka, in 2021. DENV gene was detected by semi-nested RT-PCR, and sequences of envelope (E) gene and C-prM gene were determined by direct sequencing of RT-PCR products for genetic analysis. Results: Among 172 NS1-positive samples collected, 91 samples were assigned to DENV-3 and DENV-2 (88 and 3 samples, respectively) by RT-PCR targeting the C-prM gene. Phylogenetic analysis of the E gene for the 17 representative DENV-3 samples showed that all the viruses belonged to genotype I, forming a cluster (B-cluster) with those of DENV-3 reported in Bangladesh in 2017. Analysis of the deduced amino acid sequences of E protein revealed 16 amino acid substitutions, including two novel ones (G221W, L285P), and a substitution T223I that was specifically found in DENV-3 B-cluster. Conclusion: This study showed the persistent predominance of DENV-3 genotype I in Bangladesh having unique genetic traits in the E gene. (Approval number: MMC/IRB/2022/468).

Clonal Diversity of Candida auris, Candida blankii, and Kodamaea ohmeri Isolated from Septicemia and Otomycosis in Bangladesh as Determined by Multilocus Sequence Typing.

Candida auris, Candida blankii, and Kodamaea ohmeri have been regarded as emerging fungal pathogens that can cause infections with high mortality. For genotyping of C. auris, a multilocus sequence typing (MLST) scheme based on four locus sequences has been reported, while there is no typing scheme for C. blankii and K. ohmeri. In the present study, the existing MLST scheme of C. auris was modified by adding more locus types deduced from sequence data available in the GenBank database. Furthermore, MLST schemes of C. blankii and K. ohmeri were developed using the four cognate loci (ITS, RPB1, RPB2, D1/D2) and similar sequence regions to those of C. auris. These MLST schemes were applied to identify the ST (sequence type) of clinical isolates of C. auris (n = 7), C. blankii (n = 9), and K. ohmeri (n = 6), derived from septicemia or otomycosis in Bangladesh in 2021. All the C. auris isolates were classified into a single ST (ST5) and clade I, having a Y132F substitution in ERG11p, which is associated with azole resistance. Similarly, all the C. blankii isolates belonged to a single type (ST1). In contrast, six K. ohmeri isolates were assigned to five types (ST1-ST5), suggesting its higher genetic diversity. These findings revealed the availability of MLST schemes for these three fungal species for understanding their clonal diversity among clinical isolates.

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.