Detection of Anti- Leptospira

Leptospirosis is a globally distributed zoonosis with varied clinical outcomes and multiorgan involvement in humans. In this study conducted from July 2011 to December 2011, 178 serum samples from patients suspected of leptospirosis were tested by Panbio IgM ELISA at National Public Health Laboratory, Kathmandu, out of which 51 (28.65%) were positive for anti-Leptospira IgM antibody. Leptospirosis was more common in people in their 2nd and 3rd decades of their life which together comprised 56.86% of the total positive cases. Most of those tested positive were farmers followed by students and housewives. Both animal contact and water contact seemed to play significant roles in disease transmission. Symptoms were vague with the most common being fever, headache, myalgia, abdominal pain, vomiting, jaundice, and diarrhoea. Life style heavily dominated by agronomical and farming activities in Nepal is conducive to leptospirosis transmission. Leptospirosis seems to be a significant public health problem in Nepal but is underestimated. In resource poor countries like Nepal where laboratories performing MAT or maintaining cultures are rarely available, serological test like ELISA could well depict the scenario of the disease prevalence

Cryptosporidium infection among the school children of Kathmandu Valley

Introduction: Cryptosporidium, a coccidian protozoan genus has emerged as an important cause of parasitic diarrhea among children living in developing countries. The present study aimed to determine the infection of Cryptosporidium and other intestinal parasites among the school children of Kathmandu Valley. Methods: Ethical approval for this research was obtained from Institutional review board, Institute of Medicine. A total of five hundred and seven stool samples from children between the age group 3-14 years, studying in 13 different schools of Kathmandu were collected during the study period (May- November, 2014) and processed in the Public Health Research Laboratory, Institute of Medicine, Kathmandu, Nepal. A modified Acid Fast staining technique (Kinyoun’s method) was used to detect oocysts of Cryptosporidium from the formal-ether concentrated stool samples. Result: The overall, prevalence of intestinal parasites among children was found to be 22.68% (115/507)with Giardia lamblia being the most predominant parasite showing the incidence of 10.45% (53/507). Cryptosporidium oocysts were detected in 0.79% (4/507) of the students investigated with the highest occurrence during the month of July i.e., 2 followed by single positive case for each, June and September. Conclusion: The detection of Cryptosporidium oocysts and observance of diarrheal symptoms, together with the pattern of age-specific occurrence, livestock presence at home, consumption of untreated drinking water and raw vegetables/fruits consumption habit among infected children suggest that in low-income Kathmandu communities, cryptosporidiosis coupled with poor sanitary practice is a public-health issue causing potentially serious consequencesBhandari D, Tandukar S, Sherchand S, Thapa P, Shah P

Climate change and infectious disease research in Nepal: Are the available prerequisites supportive enough to researchers?

Available online 10 January 2020Although Nepal has been identified as a country highly vulnerable to adverse health and socioeconomic impacts arising from climate change, extant research on climate sensitive infectious diseases has yet to develop the evidence base to adequately address these threats. In this opinion paper we identify and characterise basic requirements that are hindering the progress of climate change and infectious disease research in Nepal. Our opinion is that immediate attention should be given to strengthening Nepal's public health surveillance system, promoting inter-sectoral collaboration, improving public health capacity, and enhancing community engagement in disease surveillance. Moreover, we advocate for greater technical support of public health researchers, and data sharing among data custodians and epidemiologists/researchers, to generate salient evidence to guide relevant public health policy formulation aimed at addressing the impacts of climate change on human health in Nepal. International studies on climate variability and infectious diseases have clearly demonstrated that climate sensitive diseases, namely vector-borne and food/water-borne diseases, are sensitive to climate variation and climate change. This research has driven the development and implementation of climate-based early warning systems for preventing potential outbreaks of climate-sensitive infectious diseases across many European and African countries. Similarly, we postulate that Nepal would greatly benefit from a climate-based early warning system, which would assist in identification or prediction of conditions suitable for disease emergence and facilitate a timely response to reduce mortality and morbidity during epidemics.Dinesh Bhandari, Peng Bi, Jeevan Bahadur Sherchand, Meghnath Dhimal, Scott Hanson-Ease

Non-linear effect of temperature variation on childhood rotavirus infection: A time series study from Kathmandu, Nepal

Available online 30 July 2020Introduction: This study aimed to investigate the effects of temperature variability on rotavirus infections among children under 5 years of age in Kathmandu, Nepal. Findings may informinfection control planning, especially in relation to the role of environmental factors in the transmission of rotavirus infection. Methods: Generalized linear Poisson regression equationswith distributed lag non-linearmodelwere fitted to estimate the effect of temperature (maximum,mean and minimum) variation onweekly counts of rotavirus infections among children under 5 years of age living in Kathmandu, Nepal, over the study period (2013 to 2016). Seasonality and long-term effects were adjusted in the model using Fourier terms up to the seventh harmonic and a time function, respectively. We further adjusted the model for the confounding effects of rainfall and relative humidity. Results: During the study period, a total of 733 cases of rotavirus infection were recorded, with amean of 3 cases per week. We detected an inverse non-linear association between rotavirus infection and average weekly mean temperature, with increased risk (RR: 1.52; 95% CI: 1.08–2.15) at the lower quantile (10th percentile) and decreased risk (RR: 0.64; 95% CI: 0.43–0.95) at the higher quantile (75th percentile). Similarly, we detected an increased risk [(RR: 1.93; 95% CI: 1.40–2.65) and (RR: 1.42; 95% CI: 1.04–1.95)] of rotavirus infection for both maximum and minimum temperature at their lower quantile (10th percentile). We estimated that 344 (47.01%) cases of rotavirus diarrhoea among the children under 5 years of age were attributable to minimum temperature. The significant effect of temperature on rotavirus infectionwas not observed beyond lag zero week. Conclusion: An inverse non-linear association was estimated between rotavirus incidence and all three indices of temperature, indicating a higher risk of infection during the cooler times of the year, and suggesting that transmission of rotavirus in Kathmandu, Nepal may be influenced by temperature.Dinesh Bhandari, Peng Bi, Meghnath Dhimal, Jeevan Bahadur Sherchand, Scott Hanson-Ease

Prevalence of Arcobacter and other pathogenic bacteria in river water in Nepal

Published: 10 July 2019This study aims to determine the diversity of pathogenic bacteria in the Bagmati River, Nepal, during a one-year period. A total of 18 river water samples were collected from three sites (n = 6 per site) along the river. Bacterial DNA, which were extracted from the water samples, were analyzed for bacterial 16S rRNA genes by next-generation sequencing for 13 of 18 samples, and by quantitative PCR targeting Arcobacter for all 18 samples. The 16S rRNA sequencing identified an average of 97,412 ± 35,909 sequences/sample, which were then categorized into 28 phyla, 61 classes, and 709 bacterial genera. Eighteen (16%) genera of 111 potential pathogenic bacteria were detected with abundance ratios of >1%; Arcobacter, Acinetobacter, and Prevotella were the dominant genera. The Arcobacter abundance ratios were 28.6% (n = 1), 31.3 ± 15.8% (n = 6), and 31.8 ± 17.2% (n = 6) at the upstream, midstream, and downstream sites, respectively. Arcobacter was detected in 14 (78%) of 18 samples tested, with concentrations ranging from 6.7 to 10.7 log10 copies/100 mL, based on quantitative PCR. Our results demonstrate the poor bacterial quality of the Bagmati River water, suggesting a need for implementing more measures to reduce fecal contamination in the river water.Rajani Ghaju Shrestha, Sarmila Tandukar, Dinesh Bhandari, Samendra P. Sherchan, Yasuhiro Tanaka, Jeevan B. Sherchand and Eiji Haramot

Delimitation of lymphatic filariasis transmission risk areas: a geo-environmental approach

BACKGROUND: The Global Programme to Eliminate Lymphatic Filariasis (GPELF) depends upon Mass Drug Administration (MDA) to interrupt transmission. Therefore, delimitation of transmission risk areas is an important step, and hence we attempted to define a geo-environmental risk model (GERM) for determining the areas of potential transmission of lymphatic filariasis. METHODS: A range of geo-environmental variables has been selected, and customized on GIS platform to develop GERM for identifying the areas of filariasis transmission in terms of "risk" and "non-risk". The model was validated through a 'ground truth study' following standard procedure using GIS tools for sampling and Immuno-chromotographic Test (ICT) for screening the individuals. RESULTS: A map for filariasis transmission was created and stratified into different spatial entities, "risk' and "non-risk", depending on Filariasis Transmission Risk Index (FTRI). The model estimation corroborated well with the ground (observed) data. CONCLUSION: The geo-environmental risk model developed on GIS platform is useful for spatial delimitation purpose on a macro scale

Detection of Cryptosporidium, Giardia, fecal indicator bacteria, and total bacteria in commercial jar water in Kathmandu Valley, Nepal

Introduction: Jar water is a convenient and common source of drinking water in the Kathmandu Valley. However, studies including detailed microbial analyses of this source of potable water are lacking. In this study, jar water samples were examined for the occurrence of Cryptosporidium, Giardia, fecal indicator bacteria, and total bacteria. Methods: Thirty different brands of jars were collected in September 2014. Escherichia coli and total coliforms w ere determined using a Colilert reagent. Ten of the 30 brands w ere selected to test for Cryptosporidium, Giardia, and total bacteria. Bacterial DNA extraction from water samples w as performed using the Cica Geneus DNA Extraction Kit, follow ed by quantitative polymerase chain reaction (qPCR) targeting the 16S rRNA gene of bacterial DNA. Protozoan detection was accomplished by concentrating the samples using the electronegative membrane vortex method, followed by immunomagnetic separation and fluorescent staining. Results: E. coli w as detected in 10% of the samples, with a maximum concentration of 2 most probable number (MPN)/100 mL, whereas total coliforms were detected in 97% of the samples, with a maximum and mean concentration of 7.3 × 10 2 and 3.8 × 10 1 MPN/100 mL, respectively. Total coliforms concentrations in 40% of the samples ranged from 10 2 to 10 3 MPN/100 mL. Cryptosporidium and Giardia w ere not detected in any of the tested samples. Concentrations of total bacteria in the samples ranged from 10 4 to 10 6 cells/100 mL. Conclusions: Ninety-seven percent of the jar water brands were unsuitable for drinking without proper treatment based on the guideline values of the National Drinking Water Quality Standards (NDW Q S) of Nepal. There is no guideline value for total bacteria in NDW Q S however, high concentrations can be indicative of poor control on regrowth of bacteria and recontamination or inefficient water treatment methods.Malla B, Ghaju Shrestha R, Bhandari D, Tandukar S, Shrestha S, Yoshinaga H, Inoue D, Sei K, Nishida K, Tanaka Y, Sherchand JB, Haramoto

Gut microbiome transition across a lifestyle gradient in Himalaya

Published: November 15, 2018The composition of the gut microbiome in industrialized populations differs from those living traditional lifestyles. However, it has been difficult to separate the contributions of human genetic and geographic factors from lifestyle. Whether shifts away from the foraging lifestyle that characterize much of humanity's past influence the gut microbiome, and to what degree, remains unclear. Here, we characterize the stool bacterial composition of four Himalayan populations to investigate how the gut community changes in response to shifts in traditional human lifestyles. These groups led seminomadic hunting-gathering lifestyles until transitioning to varying levels of agricultural dependence upon farming. The Tharu began farming 250-300 years ago, the Raute and Raji transitioned 30-40 years ago, and the Chepang retain many aspects of a foraging lifestyle. We assess the contributions of dietary and environmental factors on their gut-associated microbes and find that differences in the lifestyles of Himalayan foragers and farmers are strongly correlated with microbial community variation. Furthermore, the gut microbiomes of all four traditional Himalayan populations are distinct from that of the Americans, indicating that industrialization may further exacerbate differences in the gut community. The Chepang foragers harbor an elevated abundance of taxa associated with foragers around the world. Conversely, the gut microbiomes of the populations that have transitioned to farming are more similar to those of Americans, with agricultural dependence and several associated lifestyle and environmental factors correlating with the extent of microbiome divergence from the foraging population. The gut microbiomes of Raute and Raji reveal an intermediate state between the Chepang and Tharu, indicating that divergence from a stereotypical foraging microbiome can occur within a single generation. Our results also show that environmental factors such as drinking water source and solid cooking fuel are significantly associated with the gut microbiome. Despite the pronounced differences in gut bacterial composition across populations, we found little differences in alpha diversity across lifestyles. These findings in genetically similar populations living in the same geographical region establish the key role of lifestyle in determining human gut microbiome composition and point to the next challenging steps of determining how large-scale gut microbiome reconfiguration impacts human biology.Aashish R. Jha, Emily R. Davenport, Yoshina Gautam, Dinesh Bhandari, Sarmila Tandukar, Katharine M. Ng, Gabriela K. Fragiadakis, Susan Holmes, Guru Prasad Gautam, Jeff Leach, Jeevan Bahadur Sherchand, Carlos D. Bustamante, Justin L. Sonnenbur

Onchocerca parasites and Wolbachia endosymbionts: evaluation of a spectrum of antibiotic types for activity against Onchocerca gutturosa in vitro

BACKGROUND: The filarial parasites of major importance in humans contain the symbiotic bacterium Wolbachia and recent studies have shown that targeting of these bacteria with antibiotics results in a reduction in worm viability, development, embryogenesis, and survival. Doxycycline has been effective in human trials, but there is a need to develop drugs that can be given for shorter periods and to pregnant women and children. The World Health Organisation-approved assay to screen for anti-filarial activity in vitro uses male Onchocerca gutturosa, with effects being determined by worm motility and viability as measured by reduction of MTT to MTT formazan. Here we have used this system to screen antibiotics for anti-filarial activity. In addition we have determined the contribution of Wolbachia depletion to the MTT reduction assay. METHODS: Adult male O. gutturosa were cultured on a monkey kidney cell (LLCMK 2) feeder layer in 24-well plates with antibiotics and antibiotic combinations (6 to 10 worms per group). The macrofilaricide CGP 6140 (Amocarzine) was used as a positive control. Worm viability was assessed by two methods, (i) motility levels and (ii) MTT/formazan colorimetry. Worm motility was scored on a scale of 0 (immotile) to 10 (maximum) every 5 days up to 40 days. On day 40 worm viability was evaluated by MTT/formazan colorimetry, and results were expressed as a mean percentage reduction compared with untreated control values at day 40. To determine the contribution of Wolbachia to the MTT assay, the MTT formazan formation of an insect cell-line (C6/36) with or without insect Wolbachia infection and treated or untreated with tetracycline was compared. RESULTS: Antibiotics with known anti-Wolbachia activity were efficacious in this system. Rifampicin (5 × 10(-5)M) was the most effective anti-mycobacterial agent; clofazimine (1.25 × 10(-5)M and 3.13 × 10(-6)M) produced a gradual reduction in motility and by 40 days had reduced worm viability. The other anti-mycobacterial drugs tested had limited or no activity. Doxycycline (5 × 10(-5)M) was filaricidal, but minocycline was more effective and at a lower concentration (5 × 10(-5)M and 1.25 × 10(-5)M). Inactive compounds included erythromycin, oxytetracycline, trimethoprim and sulphamethoxazole. The MTT assay on the insect cell-line showed that Wolbachia made a significant contribution to the metabolic activity within the cells, which could be reduced when they were exposed to tetracycline. CONCLUSION: The O. gutturosa adult male screen for anti-filarial drug activity is also valid for the screening of antibiotics for anti-Wolbachia activity. In agreement with previous findings, rifampicin and doxycycline were effective; however, the most active antibiotic was minocycline. Wolbachia contributed to the formation of MTT formazan in the MTT assay of viability and is therefore not exclusively a measure of worm viability and indicates that Wolbachia contributes directly to the metabolic activity of the nematode