CCL18 aggravates atherosclerosis by inducing CCR6-dependent T-cell influx and polarization.

Introduction: The CC chemokine ligand 18 (CCL18) is a chemokine highly expressed in chronic inflammation in humans. Recent observations of elevated CCL18 plasma levels in patients with acute cardiovascular syndromes prompted an investigation into the role of CCL18 in the pathogenesis of human and mouse atherosclerosis. Methods and results: CCL18 was profoundly upregulated in ruptured human atherosclerotic plaque, particularly within macrophages. Repeated administration of CCL18 in Western-type diet-fed ApoE -/- mice or PCSK9mut-overexpressing wild type (WT) mice led to increased plaque burden, enriched in CD3+ T cells. In subsequent experimental and molecular modeling studies, we identified CCR6 as a functional receptor mediating CCL18 chemotaxis, intracellular Ca2+ flux, and downstream signaling in human Jurkat and mouse T cells. CCL18 failed to induce these effects in vitro in murine spleen T cells with CCR6 deficiency. The ability of CCR6 to act as CCL18 receptor was confirmed in vivo in an inflammation model, where subcutaneous CCL18 injection induced profound focal skin inflammation in WT but not in CCR6-/- mice. This inflammation featured edema and marked infiltration of various leukocyte subsets, including T cells with a Th17 signature, supporting CCR6's role as a Th17 chemotactic receptor. Notably, focal overexpression of CCL18 in plaques was associated with an increased presence of CCR6+ (T) cells. Discussion: Our studies are the first to identify the CCL18/CCR6 axis as a regulator of immune responses in advanced murine and human atherosclerosis.

Detection of blaKPC gene among carbapenemase producing Klebsiella pneumoniae isolated from different clinical specimens at tertiary care hospital of Nepal.

BACKGROUND: Klebsiella pneumoniae infections have become a major cause of hospital acquired infection worldwide with the increased rate of acquisition of resistance to antibiotics. Carbapenem resistance mainly among Gram negative is an ongoing problem which causes serious outbreaks dramatically limiting treatment options. This prospective cross-sectional study was designed to detect blaKPC gene from carbapenem resistant K. pneumoniae. MATERIALS AND METHODS: A totally of 1118 different clinical specimens were screened and confirmed for KPC producing K. pneumoniae phenotypically using Meropenem (10 µg) disc. The blaKPC gene was amplified from the isolates of K. pneumoniae to detect the presence of this gene. RESULT: Of the total samples processed, 18.6% (n = 36) were K. pneumoniae and among 36 K. pneumoniae, 61.1% (n = 22/36) were meropenem resistant. This study demonstrated the higher level of MDR 91.7% (n = 33) and KPC production 47.2% (n = 17) among K. pneumoniae isolates. The blaKPC gene was detected in 8.3% (n = 3) of meropenem resistant isolates. CONCLUSION: Since the study demonstrates the higher level of MDR and KPC producing K. pneumoniae isolates that has challenged the use of antimicrobial agents, continuous microbiology, and molecular surveillance to assist early detection and minimize the further dissemination of blaKPC should be initiated. We anticipate that the findings of this study will be useful in understanding the prevalence of KPC-producing K. pneumoniae in Nepal.

Characterization of Thermostable Cellulase from Bacillus licheniformis PANG L Isolated from the Himalayan Soil.

This study aimed to isolate, purify, and characterize a potential thermophilic cellulase-producing bacterium from the Himalayan soil. Eleven thermophilic bacteria were isolated, and the strain PANG L was found to be the most potent cellulolytic producer. Morphological, physiological, biochemical, and molecular characterization identified PANG L as Bacillus licheniformis. This is the first study on the isolation of thermostable cellulase-producing Bacillus licheniformis from the Himalayan soil. This bacterium was processed for the production of cellulase enzyme. The optimum conditions for cellulase production were achieved at 45°C after 48 h of incubation at pH 6.5 in media-containing carboxymethyl cellulose (CMC) and yeast extract as carbon and nitrogen sources, respectively, in a thermo-shaker at 100 rpm. The enzyme was partially purified by 80% ammonium sulphate precipitation followed by dialysis, resulting in a 1.52-fold purification. The optimal activity of partially purified cellulase was observed at a temperature of 60°C and pH 5. The cellulase enzyme was stable within the pH ranges of 3-5 and retained 67% of activity even at 55°C. Cellulase activity was found to be enhanced in the presence of metal ions such as Cd2+, Pb2+, and Ba2+. The enzyme showed the highest activity when CMC was used as a substrate, followed by cellobiose. The Km and Vmax values of the enzyme were 1.8 mg/ml and 10.92 µg/ml/min, respectively. The cellulase enzyme obtained from Bacillus licheniformis PANG L had suitable catalytic properties for use in industrial applications.

Extended-spectrum ß-lactamases producing Enterobacteriaceae (ESBL-PE) prevalence in Nepal: A systematic review and meta-analysis.

An alarming increase in the occurrence of extended-spectrum ß-lactamase-producing Enterobacteriaceae (ESBL-PE) has threatened the treatment and management of bacterial infections. This systematic review and meta-analysis aimed to provide a quantitative estimate of the prevalence of ESBL among the members of the Enterobacteriaceae family by analyzing the community-based and clinical studies published between 2011 and 2021 from Nepal and determine if ESBL-PE correlates with multidrug resistance (MDR). The Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines were followed for systematic review and meta-analysis and the articles' quality was assessed using the Newcastle-Ottawa scale. Of the 2529 articles screened, 65 articles were systematically reviewed, data extracted, and included in in-depth meta-analysis. The overall pooled prevalence of ESBL-producers in Enterobacteriaceae was 29 % (95 % CI: 26-32 %) with high heterogeneity (I2 = 96 %, p < 0.001). Escherichia coli was the predominant ESBL-producing member of the Enterobacteriaceae family, followed by Citrobacter spp. and Klebsiella spp. The prevalence of ESBL-PE increased from 18.7 % in 2011 to 29.5 % in 2021. A strong positive correlation (r = 0.98) was observed between ESBL production and MDR in Enterobacteriaceae. ESBL-PE isolates showed high resistance to ampicillin, cephalosporins, and amoxicillin-clavulanic acid, and blaCTX-M type was the most reported gene variant among ESBL-PE. In conclusion, this study demonstrated an increased prevalence of ESBL-PE in Nepal over the last decade, and such isolates showed a high level of MDR against the ß-lactams and non-ß-lactam antibiotics. Tackling the rising antibiotic resistance (AR) and MDR in ESBL-PE would require concerted efforts from all stakeholders to institute effective infection control programs in the community and clinical settings.

Detection of blaoxa-23 Gene from Carbapenem-resistant Acinetobacter Baumannii.

BACKGROUND: Antibiotic resistance is a great concern for public health and Acinetobacter baumannii-associated infections are increasing in many parts of the world, including Nepal. However, limited data is available on the prevalence of A. baumannii harboring blaOXA-23 from Nepal. METHODS: A hospital-based cross-sectional study was designed to detect the blaOXA-23 gene from carbapenem-resistant A. baumannii isolates in Nepal. A total of 380 clinical specimens were collected and processed following standard microbiological procedures. Antibiotic susceptibility test was performed as per the protocol of the Kirby-Bauer disk diffusion technique and the CLSI guidelines, while screening of carbapenemase production was assessed by the Modified Hodge Test using meropenem (10µg) disc. The presence of the blaOXA-23 gene in carbapenemase-positive A. baumannii was confirmed by PCR. RESULTS: Among 380 specimens analyzed, 210 (55.3%) samples were positive for bacterial growth, where 33(15.7% of total growth) of the isolates were A. baumannii, and most of them were isolated from the ICU patients (20/33, 60.6%) and sputum (16/33, 48.5%). Thirty-two isolates (97%) were colistin sensitive, while only four (12.1%) isolates were sensitive to meropenem and imipenem. Twenty-three (69.7%) of A. baumannii were carbapenemase positive as revealed by the Modified Hodge Test test, and 19 of them (57.6% of total A. baumannii) harbored the blaOXA-23 gene. CONCLUSIONS: A high prevalence of the blaOXA-23 gene among carbapenem-resistant A. baumannii isolates were found. Systematic network surveillance should be established to check the spread of such isolates, especially in the intensive care units of tertiary care hospitals in Nepal.

Cryptococcal meningitis in people living with human immunodeficiency virus in Nepal: Perspectives from resource limited setting.

Early diagnosis of cryptococcal meningitis among people living with HIV (PLHIV) is crucial for its therapeutic success. The objective of this study was to diagnose cryptococcal meningitis in PLHIV cases using the available laboratory techniques for its confirmation in resource limited setting. This cross-sectional prospective study was conducted among 72 PLHIV with clinical suspicion of meningitis. Each cerebrospinal fluid (CSF) sample received at the National Public Health Laboratory, Kathmandu was processed for India ink staining, cryptococcal antigen lateral flow assay, and fungal culture following standard protocols. The laboratory-confirmed cryptococcal meningitis cases were between 24 and 69 years of age (median age 39 years) with 87.5% (12/14) of cases being male. Cryptococcus was detected in 22.22% (16/72) by any of the three tests, 19.44% (14/72) by cryptococcal antigen lateral flow assay, 16.66% (12/72) by India ink staining, and 8.33% (6/72) by culture. High percentage of cryptococcal meningitis among PLHIV warrants early microbiological diagnosis for better case management. Cryptococcal antigen detection immunoassay should be the priority test for laboratory diagnosis of cryptococcal meningitis in PLHIV. Alternatively, very simple and economic India ink staining of CSF specimens could be used in resource limited settings.

Feasibility of one-shot dilation access in the pediatric age group.

Objective: To compare sequential fascial dilation (SFD) versus one-shot dilation (OSD) in the pediatric patients undergoing percutaneous nephrolithotomy. Methods: The present study is an observational study. The study subjects were divided into two groups. In group 1, renal dilation was done using the SFD and in group 2, renal dilation was done using the OSD. The amount of time exposed to radiation during access to pelvicalyceal system was estimated. Complications, stone free rates, ancillary procedures for residual stones and hospital stay were compared. Modified Clavien-Dindo classification was used for grading the complications. Results: Radiation exposure and operative time were less in OSD group (95% confidence interval (CI) 3.068 to 14.072, and 2.565 to 12.435, p<0.005). The mean drop of hematocrit was statistically less significant in OSD group (p=0.032). In both groups, complications, stone free rate and hospital stay were statistically insignificant. Conclusions: OSD is feasible in the children with reduced radiation exposure and shorter operative time. The outcome was similar to SFD.

Investigation of Visceral LeishmaniasisTransmission in Selected Districts of Nepal.

BACKGROUND: Visceral leishmaniasis is transmitted to humans by Leishmania donovani infected Phlebotomus argentipes sandflies. Nepal has successfully met the elimination target of less than 1 case per 10,000, although recently this threshold has been surpassed demonstrating ongoing transmission. The main objective of the present study was to investigate transmission of visceral leishmaniasis in 4 visceral leishmaniasis endemic districts of Nepal including Palpa, Morang, Saptari and Sarlahi. METHODS: Human blood samples (331), domestic animals blood samples [goats (n =67), dogs (n =1), cows (n = 6), buffaloes (n = 16), and ox (n = 10)] and sandflies samples (3976 from 142 households) were collected from the villages of these 4 districts. Human blood samples were tested for VL antibodies using the rK39 rapid diagnostic test (InBios International, Seattle, WA). kDNA of L.donovani was amplified by PCR from DNA extracted from human blood, animal blood and sandfly samples. RESULTS: Out of 331 screened across 4 districts,32 were positive on rK39 serology and 16 were positive by PCR amplification of kDNA from L. donovani. The majority of the positive serology and PCR tests were from the Ishworpur village in the Sarlahi district where there was an outbreak of 18 cases of VL. This study also revealed the presence of L. donovani DNA in female P. argentipes sandflies collected from the Ishworpur village of Sarlahi, 6 villages in the Saptari,10 villages in the Palpa, and from 9 villages in the Morang. Blood samples from domestic animals in the same villages were negative for kDNA detection by PCR. CONCLUSIONS: The results of human and sandfly findings strongly point towards local transmission of visceral leishmaniasis in these 4 districts of Nepal. Notably, there is a significant level of transmission in the Ishworpur village in the Sarlahi district. The observations from this study suggest that domestic animals are not a reservoir host for L. donovani in these districts in Nepal. Ongoing surveillance is needed to identify new outbreaks such as in the Sarlahi district.

Crossing Vessel in Pelvi Ureteric Junction Obstruction: A Histopathological Analysis.

OBJECTIVE: The aim of the study is to identify whether crossing vessel is a cause or an associated finding in Pelvi Ureteric Junction Obstruction. MATERIAL AND METHODS: This is a prospective study of a total of 128 patients who underwent laparoscopic pyeloplasty from January 2016 to June 2020. All patients who underwent laparoscopic pyeloplasty and pelvi ureteric junction segments were sent for histopathological examination. The presence of crossing vessels is documented intraoperative and patients were divided into two groups, group 1 having pelvi ureteric junction obstruction with crossing vessel, and group 2, pelvi ureteric junction obstruction without crossing vessels. Histopathological examination findings of pelvi ureteric junction segment including inflammation, fibrosis, muscle hypertrophy, muscle disarray, and synaptophysin were recorded. Unpaired Student t-test was used for comparing differences between continuous normally distributed data from 2 samples and non-parametric tests were applied for continuous data. RESULTS: Of the total 128 patients, crossing vessels were identified in 42 (32.8%), and 86 (67.2%) were without crossing vessels. The demographic profile of patients between the 2 groups was comparable. On histopathological examination, moderate-to-severe chronic inflammation was seen in 23.8% and 44.2% (P > .05) in group 1 and group 2, respectively; fibrosis and muscular hypertrophy were higher in group 2 but statistically insignificant (P > .05), and muscle disarray was higher in group 1 but statistically insignificant (P > .05). Synaptophysin was positive in 4.8% and 4.7% in group 1 and group 2, respectively. CONCLUSION: The differences in histopathological examination between the 2 groups were not statistically significant. However, in patients with crossing vessels, there was a higher degree of inflammation, which may lead to early pelvi ureteric junction obstruction.

High level of persister frequency in clinical staphylococcal isolates.

BACKGROUND: Staphylococcus aureus is a notorious human pathogen that causes often lethal systemic conditions that are mostly medical device associated biofilm infections. Similarly, coagulase negative staphylococci are emerging as leading pathogen for nosocomial infections owing to their ability to form biofilm on implanted medical equipment. Chronic in nature, these infections are difficult to treat. Such recalcitrance of these infections is caused mainly due to the presence of persister cells, which exhibit transient yet extreme tolerance to antibiotics. Despite tremendous clinical significance, there is lack of studies on persister cells formation among clinical bacterial isolates. Considering the importance of factors influencing persister formation, in this study, we evaluate the association of antibiotic tolerance with biofilm production, antibiotic stress, growth phase, specimen type, and dependency on staphylococcal species. Biofilm formation was detected among 375 clinical staphylococcal isolates by quantitative tissue culture plate method (TCP) and icaAD genes by genotypic method. The antibiotic susceptibility was determined by Kirby Bauer disc diffusion method while minimum inhibitory concentration values were obtained by agar dilution method. Persister cells were measured in the susceptible staphylococcal isolates in the presence of clinically relevant antibiotics. RESULTS: In the study, 161 (43%) S. aureus and 214 (57%) coagulase negative staphylococci (CNS) were isolated from different clinical samples. TCP method detected biofilm production in 84 (52.2%) S. aureus and 90 (42.1%) CNS isolates. The genotypic method detected icaAD genes in 86 (22.9%) isolates. Majority (> 90%) of both the biofilm producers and non-producers were sensitive to chloramphenicol and tetracycline but were resistant to penicillin. Interestingly, all isolates were sensitive to vancomycin irrespective of biofilm production. While high persister frequency was observed among all staphylococci isolates in the stationary growth phase, the persister frequency in exponential growth phase was statistically high among isolates possessing icaAD genes compared to icaAD negative isolates. CONCLUSION: The research findings provide strong evidence that the clinical staphylococcal isolates exhibit extreme antibiotic tolerance suggesting their causal link with treatment failures. Understanding the factors influencing the formation and maintenance of persister cells are of utmost important aspect to design therapeutics and control recalcitrant bacterial infections.

Detection of differential expression of miRNAs in computerized tomography-guided lung biopsy.

Aims: Nonsmall-cell lung carcinoma comprises 85% of lung malignancies and is usually associated with a poor prognosis due to diagnosis at advanced stages. Molecular diagnosis of computerized tomography (CT)-guided biopsy has the potential to identify subtypes of lung carcinoma like adenocarcinoma (AC) and squamous cell carcinoma (SCC) along with its molecular stratification. This approach will help predict the genetic signature of lung cancer in individual patients. Subjects and Methods: Histopathologically proved a CT-guided biopsy sample of lung cancer cases was used to screen for the expression of microRNA (miRNA) earlier quantitated in blood plasma. Primers against hsa-miR2114, hsa-miR2115, hsa-miR2116, hsa-miR2117, hsa-miR449c, and hsa-miR548q with control RNU6 were used to screen 30 AC, 30 SCC, 5 nonspecific granulomatous inflammation, and 8 control samples. Reverse transcription polymerase chain reaction (RT-PCR) data revealed expression of hsa-miR2114 and hsa-miR548q in AC as well as SCC. Results: RT-PCR data revealed that the expression of hsa-miR2116 and hsa-miR449c was found upregulated in AC while hsa-miR2117 was expressed in SCC cases. Bioinformatic analysis revealed that genes, where these miRNAs are located, were also upregulated while targets of these miRNAs were downregulated. Conclusions: miRNAs expression pattern in the CT-guided biopsy samples can be used as a potential tool to differentially diagnose lung cancer subtypes. The expression pattern of miRNAs matches very well in blood plasma and tissue samples, albeit levels were very low in the earlier case than later. This approach can also be used for screening mutations and other molecular markers in a personalized manner for the management of lung cancer patients.

TMQuery: a database of precomputed template modeling scores for assessment of protein structural similarity.

SUMMARY: Comparisons of protein structures are critical for developing novel protein designs, annotating protein functions and predicting protein structure. The template modeling score (TM-score) is a widely used but computationally expensive measure of protein similarity that is applicable to a wide variety of structural biology problems. We introduce TMQuery-a continuously updated database containing over eight billion pre-computed TM-score values for every pair of proteins in the Protein Data Bank, allowing researchers to quickly query and download TM-scores via a web interface. AVAILABILITY AND IMPLEMENTATION: Publicly available at https://tmquery.gsk.com/.

Serum Metabolic Disturbances in Lung Cancer Investigated through an Elaborative NMR-Based Serum Metabolomics Approach.

Detection of metabolic disturbances in lung cancer (LC) has the potential to aid early diagnosis/prognosis and hence improve disease management strategies through reliable grading, staging, and determination of neoadjuvant status in LC. However, a majority of previous metabolomics studies compare the normalized spectral features which not only provide ambiguous information but further limit the clinical translation of this information. Various such issues can be resolved by performing the concentration profiling of various metabolites with respect to formate as an internal reference using commercial software Chenomx. Continuing our efforts in this direction, the serum metabolic profiles were measured on 39 LC patients and 42 normal controls (NCs, comparable in age/sex) using high-field 800 MHz NMR spectroscopy and compared using multivariate statistical analysis tools to identify metabolic disturbances and metabolites of diagnostic potential. Partial least-squares discriminant analysis (PLS-DA) model revealed a distinct separation between LC and NC groups and resulted in excellent discriminatory ability with the area under the receiver-operating characteristic (AUROC) = 0.97 [95% CI = 0.89-1.00]. The metabolic features contributing to the differentiation of LC from NC samples were identified first using variable importance in projection (VIP) score analysis and then checked for their statistical significance (with p-value < 0.05) and diagnostic potential using the ROC curve analysis. The analysis revealed relevant metabolic disturbances associated with LC. Among various circulatory metabolites, six metabolites, including histidine, glutamine, glycine, threonine, alanine, and valine, were found to be of apposite diagnostic potential for clinical implications. These metabolic alterations indicated altered glucose metabolism, aberrant fatty acid synthesis, and augmented utilization of various amino acids including active glutaminolysis in LC.

Initiation and feasibility of a multi-specialty minimally invasive surgical programme using a novel robotic system: A case series.

INTRODUCTION: There are a number of small case series examining new robotic surgical systems, but this is the first large case series assessing the feasibility of the Versius® system from CMR Surgical (1 Evolution Business Park, Cambridge, UK) in a multi-specialty setting. MATERIALS AND METHODS: All patients undergoing Versius®-assisted surgery in a previously robot-naïve centre were consented for collection of data on demographics, pre-, intra-, and postoperative outcomes. Data collection was performed prospectively from the start of the robotic surgical programme. RESULTS: 160 operations were performed over a 19-month period, including 68 colorectal, 60 gynaecology, and 32 general surgery cases. The conversion rate to open surgery was 4.4% for colorectal, and 0% for gynaecology and general surgery. Median length of stay was 6 days for colorectal, 1 day for gynaecology, and 0 days for general surgery. Other outcomes were comparable to existing literature for robotic assisted surgery. CONCLUSION: The Versius® system is safe and feasible for use in a multi-specialty minimally invasive surgery programme, including colorectal, general surgical & gynaecological cases, and operative volume can be safely and easily scaled up in a district general hospital setting without prior robotic surgical experience.

CCR6 Deficiency Increases Infarct Size after Murine Acute Myocardial Infarction.

Ischemia-reperfusion injury after the reopening of an occluded coronary artery is a major cause of cardiac damage and inflammation after acute myocardial infarction. The chemokine axis CCL20-CCR6 is a key player in various inflammatory processes, including atherosclerosis; however, its role in ischemia-reperfusion injury has remained elusive. Therefore, to gain more insight into the role of the CCR6 in acute myocardial infarction, we have studied cardiac injury after transient ligation of the left anterior descending coronary artery followed by reperfusion in Ccr6-/- mice and their respective C57Bl/6 wild-type controls. Surprisingly, Ccr6-/- mice demonstrated significantly reduced cardiac function and increased infarct sizes after ischemia/reperfusion. This coincided with a significant increase in cardiac inflammation, characterized by an accumulation of neutrophils and inflammatory macrophage accumulation. Chimeras with a bone marrow deficiency of CCR6 mirrored this adverse Ccr6-/- phenotype, while cardiac injury was unchanged in chimeras with stromal CCR6 deficiency. This study demonstrates that CCR6-dependent (bone marrow) cells exert a protective role in myocardial infarction and subsequent ischemia-reperfusion injury, supporting the notion that augmenting CCR6-dependent immune mechanisms represents an interesting therapeutic target.

Meningococcal Carriage among Household Contacts of Patients with Invasive Meningococcal Disease in Kathmandu, Nepal: A Longitudinal Study.

Because asymptomatic carriers are key source of transmission, information on meningococcal carriage in the community provides a scientific basis for appropriate preventive/control strategies. This longitudinal study (January 2017-December 2019) aimed to estimate carriage rate of meningococci among household contacts of meningococcal meningitis cases within Kathmandu Valley, Nepal. Throat swab samples were collected at first visit from each person in households, twice a month for up to 2 months and subsequently on a monthly basis for a further 4 months. Altogether, 1125 throat samples were processed by conventional culture for the identification of meningococci. To the best of our knowledge, this is the first longitudinal study on meningococcal carriage in Nepal. The meningococcal carriage rate among household contacts was 15%. All carriers were aged 19 years or older. There was no statistically significant gender difference. The duration of carriage was 60 days. Twenty of 36 isolates belonged to serogroup A, and 16 were non-serogroupable (NG). Serogroups isolated from the same individuals did not change within the follow-up period. All meningococcal isolates over the past 38 years in Nepal that have been reported in previous studies have belonged to serogroup A. The detection of NG meningococcal isolates in apparently healthy household contacts clearly indicates the importance of vigilance through surveillance and periodic in-depth studies.

Phenotypic and genotypic characterization of biofilm producing clinical coagulase negative staphylococci from Nepal and their antibiotic susceptibility pattern.

BACKGROUND: Coagulase-negative staphylococci (CNS) survive as commensals of skin, anterior nares and external canals of human and were regarded as non-infectious pathogens. However, they are emerging as a major cause of nosocomial infectious due to their ability to form biofilms and high resistance to several classes of antibiotics. This study examines the biofilm forming abilities of 214 clinical CNS isolates using phenotypic and genotypic methods, and determines their antibiotic susceptibility patterns. METHODS: A total of 214 clinical isolates collected from different clinical samples were identified as CNS and their antibiotic susceptibility determined by CLSI guidelines. The biofilm forming ability of all isolates was determined by three phenotypic methods; Congo red agar (CRA) method, tube adherence method (TM) and tissue culture plate (TCP) method and by genotypic method for the detection of icaAD genes. RESULTS: Among all the isolates, S. epidermidis (57.5%) was found the most frequently, followed by S. saprophyticus (18.7%), S. haemolyticus (11.2%), S. hominis (7%), and S. capitis (5.6%). Antibiotic susceptibility pattern demonstrated 91.6% isolates were resistant to penicillin and 66.8% to cefoxitin while 91.1% isolates were susceptible to chloramphenicol. Constitutive and inducible clindamycin resistant phenotype as measured by D-test was seen among 28% and 14.5% of isolates respectively. Tissue culture plate method detected biofilm production in 42.1% isolate followed by 31.8% through tube method while 20.1% isolates were found to produce slime in Congo red agar method. The genotypic assay revealed presence of icaA and icaD genes in 19.2% isolates. CONCLUSION: The study shows a high prevalence of biofilm formation and inducible clindamycin resistance in CNS isolates, indicating the importance of in-vitro biofilm production test and D-test in routine laboratory diagnostics. Implementation of efficient diagnostic techniques for detection of biofilm production in clinical samples can help manage staphylococcal infections and minimize risks of treatment failures in hospitals.

Detection of Streptococcus pneumoniae, Neisseria meningitidis and Haemophilus influenzae in Culture Negative Cerebrospinal Fluid Samples from Meningitis Patients Using a Multiplex Polymerase Chain Reaction in Nepal.

The rapid identification of bacteria causing meningitis is crucial as delays in the treatment increase mortality rate. Though considered as the gold standard for the laboratory diagnosis of bacterial meningitis, culture might give false negative results in a case of patients under antibiotics prior to lumbar puncture. This study aimed to detect Streptococcus pneumoniae, Neisseria meningitidis and Haemophilus influenzae by a multiplex polymerase chain reaction (PCR) in culture-negative cerebrospinal fluid samples collected from clinically suspected meningitis cases attending different hospitals in Kathmandu, Nepal from January 2017 to December 2019. S. pneumoniae, N. meningitidis and H. influenzae were detected in 8.59% (33/384) of the specimens by PCR and 7.55% (29/384) of the specimens by culture. Correlation between culture and PCR of the same sample was good (Spearman's rho correlation coefficient = 0.932). However, the difference in positivity between culture and PCR was statistically not significant (p value > 0.05). In four specimens, culture could not detect any of the targeted bacteria whereas PCR could detect presence of H. influenzae. PCR increases the diagnostic yield for bacterial meningitis. PCR may be considered as an adjunctive test for establishing the cause of infection in culture negative clinically suspected meningitis cases.

Seropositivity of Visceral leishmaniasis on people of VL endemic three districts of Nepal.

Visceral leishmaniasis (VL) is a life-threatening vector borne disease caused by the Leishmania donovani species complex. In Nepal, it is transmitted to humans by L donovani infected Phlebotomus argentipes sand flies [12]. The pathogenesis of VL is complex, and the clinical presentation ranges from asymptomatic infection to severe and fatal disease. Asymptomatic infection may act as potential reservoirs for sustained transmission of VL in endemic areas. We investigated the sero-prevalence of symptomatic and asymptomatic infection of VL in people of three endemic districts of Nepal by serology targeting family members and neighbors of VL patients. Sero-survey was conducted among 189 people of villages endemic to VL from Palpa, Sarlahi and Saptari districts during 2016 to 2018 using the rK39 rapid diagnostic test (InBios International, Seattle, WA) to detect anti-Leishmania antibodies. Sero-positivity was 35.7% (10/28) in people tested from Sarlahi districts, 6% (3/50) in Saptari district and 1.7% (1/59) from the Palpa district. In Sarlahi, sero-positivity was found to be highest among the age group below 15 years (44.5%). All family members of diagnosed VL cases in Saptari and Palpa districts were found to be rK39 test negative. In Sarlahi district, among the ten sero-positive cases, nine were febrile and became symptomatic VL cases after few days and one case remained asymptomatic during the six month follow up. Asymptomatic cases in VL endemic districts of Nepal were found to be sero-positive, screening of people in VL endemic districts would be important for prevention of VL transmission.