The emergence of multidrug resistant (MDR) bacteria which is attributable to extended spectrum β-lactamases (ESBLs) production of CTX-M types is an obvious problem worldwide. This study is aimed at determining the prevalence of CTX-M β-lactamases producing multidrug resistant Escherichia coli and Klebsiella pneumoniae among patients attending Bir Hospital. A cross-sectional study was conducted between April and September 2019 at Bir Hospital, Kathmandu, and Department of Microbiology, National College, Kathmandu, Nepal. A total of 5,690 different clinical specimens were subjected to cultural, microscopic, and biochemical analyses for the identification of the isolates. Antimicrobial susceptibility testing of the isolates was done, and MDR isolates were selected and processed for further ESBL confirmation by the combination disks method. All confirmed ESBL isolates were screened for CTX-M type β-lactamases (blaCTX-M) by PCR. Of the total 345 isolates (227 Escherichia coli and 118 Klebsiella pneumoniae), 232 were MDR. All 232 (67.24%) MDR isolates were suspected as ESBL producers on the screening test. However, on the phenotypic test, 135 (58.18%) of total MDR bacteria were confirmed as ESBL producers with the highest proportion in K. pneumoniae (59.37%). The major source of ESBL producers was urine. ESBL producing isolates were mostly identified from outpatients and patients belonging to age group 41-60. Gentamicin was found to be effective against ESBL producers. The prevalence of blaCTX-M was (89.62%) with the highest frequency for E. coli (93.81%). High prevalence of ESBL of CTX-M types among MDR E. coli and K. pneumoniae was detected from clinical specimens of patients in Bir Hospital. This study warrants the need for the judicious use of antibiotics as well as emphasize the use of modern diagnostic tools for the early detection of MDR and ESBL producers to curb the emergence and spread of MDR and ESBL producing bacteria in the clinical settings of Nepal.

Pulmonary tuberculosis (PTB) is one of the major infectious diseases in developing countries. The objective of this study was to compare rapid diagnostics technique, GeneXpert MTB/RIF (GeneXpert) and Multiplex PCR assay (MPCR) targeting IS6110 segment and mpb64 gene for direct detection of Mycobacterium tuberculosis (MTB) in suspected PTB patients. A cross sectional study was carried among 105 sputum samples from suspected PTB patients to evaluate GeneXpert and Multiplex PCR who visited National Tuberculosis Center, Nepal. The patient's sputum samples were used directly for the GeneXpert whereas DNA extraction by CTAB method was followed to process the sample for MPCR. The sensitivity and specificity of GeneXpert and MPCR in smear positive cases was 78.6, 33.3, and 100.0%, 66.7%, respectively (P = 0.125). However, in smear negative cases sensitivity and specificity of both methods exhibited 90.9, 95.2, and 100.0%, 100.0% respectively (P = 0.625). Finally, the sensitivity and specificity of GeneXpert and MPCR were 82.9, 95.3 and 100.0%, 98.5% respectively, (P = 0.549) in pulmonary cases. Comparatively, we observed higher sensitivity and specificity for MPCR than GeneXpert for both smear positive and negative samples. Thus, we recommend MPCR alongside GeneXpert for the better diagnostic accuracy of PTB in a resource-limited country where tuberculosis is endemic.

Background: Pseudomonas aeruginosa is an opportunistic human pathogen and are reported to cause acute and chronic infectious diseases. Due to its high ability to acquire resistance to many antibiotics, it has become a global public health threat. It consists of some virulence genes that may lead to its pathogenicity. The main objective of this cross-sectional study was to detect the virulence genes and antibiotic susceptibility pattern of P. aeruginosa isolated from clinical specimens collected from governmental hospital of Nepal.Methods: A total of 7898 clinical specimens were analyzed for the period of six months from November 2018 to April 2019. The specimens were cultured on Nutrient agar, Blood agar, MacConkey agar, Chocolate agar, Cysteine-Lactose, Electrolyte Deficient agar plates and were incubated at 37°C for 24 hours. All the isolates were identified by standard biochemical tests and further confirmed by growth on Cetrimide agar plate. The antibiotic susceptibility testing was performed by modified Kirby-Bauer disc diffusion method following CLSI guideline. Multiplex-PCR was done to detect the virulence genes oprL and toxA. Statistical analysis was carried out using IBM SPSS Statistic ver. 25 and the p-value was calculated at significance level (0.05%) by using Chi square.Results: Out of these specimens investigated, 87 isolates were tentatively identified to be P. aeruginosa in which 20 (22.98 %) were found to be multidrug resistant. Comparatively, most of the P. aeruginosa were isolated from outpatients 63 (72.41 %) than inpatients 24 (27.58 %), from male 56 (64.36 %) than female 31 (35.63 %) and in age group 60-79 years (41.37 %). AST result showed the highest resistance of 100% with cefixime whereas susceptibilities of 83.9% and 81.6% with polymixin B and tobramycin were noticed respectively. The PCR results showed that all P. aeruginosa isolates carried oprL 87 (100%) and 83 (95.4 %) isolates showed toxA genes. Conclusion: The studies revealed that almost all P. aeruginosa harbors both oprL and toxA genes.

The multidrug- or extensively drug-resistant (MDR/XDR) Pseudomonas aeruginosa carrying some virulence genes has become a global public health threat. However, in Nepal, there is no existing report showing the prevalence of oprL and toxA virulence genes among the clinical isolates of P. aeruginosa. Therefore, this study was conducted for the first time in the country to detect the virulence genes (oprL and toxA) and antibiotic susceptibility pattern of P. aeruginosa. A total of 7,898 clinical specimens were investigated following the standard microbiological procedures. The antibiotic susceptibility testing was examined by the modified disc diffusion method, and virulence genes oprL and toxA of P. aeruginosa were assessed using multiplex PCR. Among the analyzed specimens, 87 isolates were identified to be P. aeruginosa of which 38 (43.68%) isolates were reported as MDR. A higher ratio of P. aeruginosa was detected from urine samples 40 (45.98%), outpatients’ specimens 63 (72.4%), and in patients of the age group of 60–79 years 36 (41.37%). P. aeruginosa was more prevalent in males 56 (64.36%) than in female patients 31 (35.63%). Polymyxin (83.90%) was the most effective antibiotic. P. aeruginosa (100%) isolates harboured the oprL gene, while 95.4% of isolates were positive for the toxA gene. Identification of virulence genes such as oprL and toxA carrying isolates along with the multidrug resistance warrants the need for strategic interventions to prevent the emergence and spread of antimicrobial resistance (AMR). The findings could assist in increasing awareness about antibiotic resistance and suggest the judicious prescription of antibiotics to treat the patients in clinical settings of Nepal.