Muhammad Akram Hossain

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 17u2009kDa 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.

Rickettsia felis Infection among Humans, Bangladesh, 2012-2013.

To the Editor: Rickettsia felis, which belongs to the spotted fever group of rickettsiae, causes febrile illness in humans. The main vector of this bacterium is the cat flea (Ctenocephalides felis). Since publication of reports of R. felis as a putative pathogen of humans in the United States in 1994, R. felis infection in humans worldwide has been increasingly described, especially in the Americas, Europe, Africa, and eastern Asia (1,2). R. felis infection is common among febrile patients (≈15%) in tropical Africa (3) and among apparently healthy persons in eastern coastal provinces of China (4). However, little is known about prevalence of R. felis infection of humans in southern Asia, although 3 serologically diagnosed cases in Sri Lanka have been described (5) and R. felis has been detected in rodent fleas in Afghanistan (6). Hence, we conducted a cross-sectional study in Bangladesh to explore the presence of rickettsial pathogens among patients with fever of unknown origin. n nStudy participants were 150 patients at Mymensingh Medical College (MMC) hospital in Mymensingh, north-central Bangladesh, from July 2012 through January 2014, and 30 healthy control participants from the staff at the same college. Selected patients met the following criteria: 1) fever (axillary temperature >37.5°C) for >15 days that did not respond to common antimicrobial drug therapy; 2) any additional clinical features including headache, rash, lymphadenopathy, myalgia, and eschars on skin; and 3) titer according to the Weil-Felix test (antibodies against any of 3 Proteus antigens) of >1:80. Patients with evident cause of fever (e.g., malaria diagnosed by blood smear or immunochromatography) were excluded from the study. This research was approved by the college institutional review board, and informed consent was obtained from patients (or guardians) and healthy controls before their entry into the study. n nVenous blood samples were aseptically collected from the patients, and DNA was extracted by conventional method by using proteinase K and sodium dodecyl sulfate. Nested PCR selective for the 17-kDa antigen gene was used to screen for rickettsiae according to the method described previously (7); ≈100 ng of DNA in a 50-μL reaction mixture was used. For each PCR, a negative control (water) was included and utmost care was taken to avoid contamination. Among the 150 samples tested, results were positive with a 232-bp amplified product for 69 (46%) and negative for all controls. n nPCR products from 20 samples were randomly selected for sequence analysis. All nucleotide sequences from the 17-kDa antigen gene (186-bp) were identical to that of reference strain R. felis URRWXCa12 (GenBank accession no. {type:entrez-nucleotide,attrs:{text:CP000053,term_id:67003925,term_text:CP000053}}CP000053). Among all 17-kDa–positive samples, positivity was further confirmed by PCR detection of the R. felis 16S rRNA gene and gltA in 95% and 75% of samples, respectively. Partial 16S rRNA gene sequences (305-bp) from 12 samples were 100% or 99% (10 and 2 samples, respectively) identical to that of R. felis URRWXCa12. The complete open reading frames of ompA (1773-bp), partial ompB (413-bp), and gltA (611-bp) sequences determined for 3, 3, and 5 samples, respectively, were also identical to those of R. felis URRWXCa12. The 5 gene sequences were determined for samples from 3 patients (2-year-old girl, 8-year-old boy, 17-year-old boy). The 5 gene sequences from the 2-year-old girl (strain Ric-MMC7) and 2 partial sequences of 16S rRNA (Ric-MMC71 and Ric-MMC133) were deposited in GenBank under accession nos. {type:entrez-nucleotide-range,attrs:{text:KP318088-KP318094,start_term:KP318088,end_term:KP318094,start_term_id:880804642,end_term_id:880804651}}KP318088-KP318094. n nAccording to PCR, the positivity rate for the R. felis 17-kDa antigen gene was higher among male (54%, 40/74) than among female (38%, 29/76) patients and higher among patients in young and old age groups (0–15 years, 57%; 45–60 years, 62%) than among patients in other age groups (15–30 years, 41%; 30–45 years, 44%). During the study period, rates of R. felis positivity were highest during the late rainy season of 2012 (September [59%] and October [52%]) and lowest (0%) from December 2012 through April 2013 (Figure). The rate was significantly higher among farmers (76%, 13/17) than among persons of other occupations (e.g., housewives, teachers, students) (42%, 56/133); p = 0.016. Among the 69 rickettsiae-positive patients, headache and myalgia were reported by 29 (42%) and 17 (25%), respectively, whereas rash was detected in only 2 (3%) patients, both of whom were female. n n n nFigure n nNumber of patients with fever of unknown origin and Rickettsia felis–positive cases in the Mymensingh Medical College hospital, Bangladesh, 2012–2013. Numbers in parentheses indicate rates of R. felis positivity for each month; dashed ... n n n nThis study demonstrated R. felis infection in patients in Bangladesh with unidentified febrile illness. The high prevalence (46%) of R. felis infection suggests that this infection is endemic to the north-central area of this country and might be associated with contact between humans of low socioeconomic status and the large number of stray cats and dogs. In contrast, the number of genetically confirmed cases of R. felis infection in humans reported to date in China, Taiwan, Thailand, and Laos have been very few (1,2,4,8–10), although widespread presence of this bacterium in cat fleas has been documented. For further confirmation of spread of this infectious disease, the prevalence of R. felis infections among humans, vectors, and reservoirs in other areas in Bangladesh and in other countries in southern Asia should be investigated.

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.

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.

Molecular characterization of chikungunya virus causing the 2017 outbreak in Dhaka, Bangladesh

Chikungunya viruses from the 2017 outbreak in Dhaka, Bangladesh, were analysed phylogenetically. E1 sequences from 21 strains belonged to the Indian Ocean clade of the East/Central/South African (ECSA) genotype, forming a novel cluster with latest South Asian strains. They lacked the A226V substitution.

Nationwide prevalence of Rickettsia felis infections in patients with febrile illness in Bangladesh

From July 2015 to December 2016, the presence of rickettsial pathogens was investigated for 414 patients with unknown fever in eight places in all the divisions of Bangladesh. Rickettsia felis was identified in blood samples from all the regions (overall detection rate, 19.6%), suggesting nationwide prevalence of R. felis infections.

Antimicrobial susceptibility pattern of enterococci isolated from clinical specimens at mymensingh medical college hospital, Mymensingh, Bangladesh

Enterococci are part of the normal intestinal flora of humans and animals but have also emerged as important pathogens responsible for serious infections in hospital and community acquired infections. The aim of this study was to investigate the prevalence of Enterococcus with their antimicrobialsusceptibility pattern from patients ofMymensingh Medical College Hospital, Bangladesh during the period from July 2015 to June 2016.Samples were collected and identification of enterococcus was done by standard microbiological procedure and biochemical tests.Multiplex PCR was performed by using species specific primers for Enterococcus faecalis (E. faecalis) and Enterococcus faecium (E. faecium),vanA, vanB, vanC1, vanC2/C3 genes for vancomycin resistance.A total of 105 enterococci were isolated from 1201 different clinical specimens (from urine (931), pus (175) and wound swab (95) respectively) from July, 2015 to June, 2016.The prevalence of enterococci was 12.34% in urine, 17.17% in pus and 1.58% in wound samples. Out of 105 enterococci,80 (77.14%) were Enterococcus faecalis,1 (0.95%) was E. gallinaramand 24 (22.86%) were other species. All the species were highly resistant to aminoglycosides (Gentamicin) (75.24%) and ciprofloxacin (73.33%). Regarding antibiotic sensitivity, all the enterococci were (100%) sensitive to linezolidand vancomycin followed by nitrofurantoin 93 (88.57%) and imipenem 89 (84.76%). No vancomycin resistant enterococci were identified by disk-diffusion method. But by PCR, vanB was found in 1/80 (1.25%) of the E. faecalis isolates and 01 (100%) of the E. gallinarum.vanC1 was detected in 1 (100%) ofE. gallinarum isolates. The vanA and vanC2/C3 gene was not found in any isolates. For the first time, one isolate of E. gallinarum has been found harboringthe vanB gene in our hospital.The presence of multidrug resistant enterococci should be considered as danger alarm for serious enterococcal infections and further study in large scale is needed.