Source Tracing of Leishmania donovani in Emerging Foci of Visceral Leishmaniasis, Western Nepal.

We sequenced Leishmania donovani genomes in blood samples collected in emerging foci of visceral leishmaniasis in western Nepal. We detected lineages very different from the preelimination main parasite population, including a new lineage and a rare one previously reported in eastern Nepal. Our findings underscore the need for genomic surveillance.

Governing the Access to COVID-19 Tools Accelerator: towards greater participation, transparency, and accountability.

The Access to COVID-19 Tools Accelerator (ACT-A) is a multistakeholder initiative quickly constructed in the early months of the COVID-19 pandemic to respond to a catastrophic breakdown in global cooperation. ACT-A is now the largest international effort to achieve equitable access to COVID-19 health technologies, and its governance is a matter of broad public importance. We traced the evolution of ACT-A's governance through publicly available documents and analysed it against three principles embedded in the founding mission statement of ACT-A: participation, transparency, and accountability. We found three challenges to realising these principles. First, the roles of the various organisations in ACT-A decision making are unclear, obscuring who might be accountable to whom and for what. Second, the absence of a clearly defined decision making body; ACT-A instead has multiple centres of legally binding decision making and uneven arrangements for information transparency, inhibiting meaningful participation. Third, the nearly indiscernible role of governments in ACT-A, raising key questions about political legitimacy and channels for public accountability. With global public health and billions in public funding at stake, short-term improvements to governance arrangements can and should now be made. Efforts to strengthen pandemic preparedness for the future require attention to ethical, legitimate arrangements for governance.

Adapting international clinical trials during COVID-19 and beyond.

BACKGROUND: The COVID-19 pandemic and resulting restrictions, particularly travel restrictions, have had significant impact on the conduct of global clinical trials. Our clinical trials programme, which relied on in-person visits for training, monitoring and capacity building across nine low- and middle-income countries, had to adapt to those unprecedented operational challenges. We report the adaptation of our working model with a focus on the operational areas of training, monitoring and cross-site collaboration. THE NEW WORKING MODEL: Adaptations include changing training strategies from in-person site visits with three or four team members to a multi-pronged virtual approach, with generic online training for good clinical practice, the development of a library of study-specific training videos, and interactive virtual training sessions, including practical laboratory-focused training sessions. We also report changes from in-person monitoring to remote monitoring as well as the development of a more localized network of clinical trial monitors to support hybrid models with in-person and remote monitoring depending on identified risks at each site. We established a virtual network across different trial and study sites with the objective to further build capacity for good clinical practice-compliant antimalarial trials and foster cross-country and cross-study site collaboration. CONCLUSION: The forced adaptation of these new strategies has come with advantages that we did not envisage initially. This includes improved, more frequent engagement through the established network with opportunities for increased south-to-south support and a substantially reduced carbon footprint and budget savings. Our new approach is challenging for study sites with limited prior experience but this can be overcome with hybrid models. Capacity building for laboratory-based work remains difficult using a virtual environment. The changes to our working model are likely to last, even after the end of the pandemic, providing a more sustainable and equitable approach to our research.

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.

Field evaluation of the diagnostic performance of EasyScan GO: a digital malaria microscopy device based on machine-learning.

BACKGROUND: Microscopic examination of Giemsa-stained blood films remains the reference standard for malaria parasite detection and quantification, but is undermined by difficulties in ensuring high-quality manual reading and inter-reader reliability. Automated parasite detection and quantification may address this issue. METHODS: A multi-centre, observational study was conducted during 2018 and 2019 at 11 sites to assess the performance of the EasyScan Go, a microscopy device employing machine-learning-based image analysis. Sensitivity, specificity, accuracy of species detection and parasite density estimation were assessed with expert microscopy as the reference. Intra- and inter-device reliability of the device was also evaluated by comparing results from repeat reads on the same and two different devices. This study has been reported in accordance with the Standards for Reporting Diagnostic accuracy studies (STARD) checklist. RESULTS: In total, 2250 Giemsa-stained blood films were prepared and read independently by expert microscopists and the EasyScan Go device. The diagnostic sensitivity of EasyScan Go was 91.1% (95% CI 88.9-92.7), and specificity 75.6% (95% CI 73.1-78.0). With good quality slides sensitivity was similar (89.1%, 95%CI 86.2-91.5), but specificity increased to 85.1% (95%CI 82.6-87.4). Sensitivity increased with parasitaemia rising from 57% at < 200 parasite/µL, to ≥ 90% at > 200-200,000 parasite/µL. Species were identified accurately in 93% of Plasmodium falciparum samples (kappa = 0.76, 95% CI 0.69-0.83), and in 92% of Plasmodium vivax samples (kappa = 0.73, 95% CI 0.66-0.80). Parasite density estimates by the EasyScan Go were within ± 25% of the microscopic reference counts in 23% of slides. CONCLUSIONS: The performance of the EasyScan Go in parasite detection and species identification accuracy fulfil WHO-TDR Research Malaria Microscopy competence level 2 criteria. In terms of parasite quantification and false positive rate, it meets the level 4 WHO-TDR Research Malaria Microscopy criteria. All performance parameters were significantly affected by slide quality. Further software improvement is required to improve sensitivity at low parasitaemia and parasite density estimations. Trial registration ClinicalTrials.gov number NCT03512678.

Multidrug-resistant bacteria with ESBL genes: a growing threat among people living with HIV/AIDS in Nepal.

BACKGROUND: Bacterial opportunistic infections are common in people living with HIV/AIDS (PLHA). Besides HIV-TB co-infection, lower respiratory tract infections (LRTIs) due to multidrug-resistant (MDR) bacteria cause significant morbidity and mortality among PLHA. This study identified bacterial co-infection of the lower respiratory tract and detected plasmid-mediated blaTEM and blaCTX-M genes among Extended-Spectrum ß-Lactamase (ESBL) producing isolates from sputum samples in PLHA. METHODS: A total of 263 PLHA with LRTIs were enrolled in this study, out of which, 50 were smokers, 70 had previous pulmonary tuberculosis, and 21 had CD4 count < 200 cells/µl. Sputum samples collected from PLHA were processed with standard microbiological methods to identify the possible bacterial pathogens. The identified bacterial isolates were assessed for antibiotic susceptibility pattern using modified Kirby Bauer disk diffusion method following Clinical Laboratory Standard Institute (CLSI) guidelines. In addition, plasmid DNA was extracted from MDR and ESBL producers for screening of ESBL genes; blaCTX-M and blaTEM by conventional PCR method using specific primers. RESULTS: Of 263 sputum samples, 67 (25.48%) showed bacterial growth. Among different bacterial pathogens, Klebsiella pneumoniae, (17; 25.37%) was the most predominant, followed by Haemophillus influenzae, (14; 20.90%) and Escherichia coli, (12; 17.91%). A higher infection rate (4/8; 50%) was observed among people aged 61-70 years, whereas no infection was observed below 20 years. About 30.0% (15/50) of smokers, 32.86% (23/70) cases with previous pulmonary tuberculosis, and 52.38% (11/21) with CD4 count < 200 cells/µl had bacterial LRTIs. Among 53 bacterial isolates excluding H. influenzae, 28 isolates were MDR and 23 were ESBL producers. All ESBL producers were sensitive to colistin and polymyxin B. Among ESBL producers, 47.83% (11/23) possessed blaCTX-M, 8.6% (2/23) were positive for blaTEM gene, and 43.48% (10/23) possessed both ESBL genes. CONCLUSION: The increasing rate of MDR bacterial infections, mainly ESBL producers of LRTIs causes difficulty in disease management, leading to high morbidity and mortality of PLHA. Hence, it is crucial to know the antibiogram pattern of the isolates to recommend effective antimicrobial therapy to treat LRTIs in PLHA.

Detection of NDM-1 and VIM Genes in Carbapenem-Resistant Klebsiella pneumoniae Isolates from a Tertiary Health-Care Center in Kathmandu, Nepal.

BACKGROUND: Klebsiella pneumoniae is one of the leading causes of nosocomial infections. Carbapenems are used as the last resort for the treatment of multidrug resistant Gram-negative bacterial infections. In recent years, resistance to these lifesaving drugs has been increasingly reported due to the production of carbapenemase. The main objective of this study was to detect the carbapenem-resistant genes blaNDM-1 and blaVIM in K. pneumoniae isolated from different clinical specimens. METHODS: A total of 585 clinical specimens (urine, pus, sputum, blood, catheter tips, and others) from human subjects attended at Annapurna Neurological Institute and Allied Sciences, Kathmandu were obtained in the period between July 2018 and January 2019. The specimens were isolated and identified for K. pneumoniae. All K. pneumoniae isolates were processed for antimicrobial susceptibility testing (AST) using the disk diffusion method. The isolates were further phenotypically confirmed for carbapenemase production by the modified Hodge test (MHT) using imipenem (10 µg) and meropenem (10 µg) discs. Thus, confirmed carbapenemase-producing isolates were further screened for the production of blaNDM-1 and blaVIM using conventional polymerase chain reaction (PCR). RESULTS: Among the clinical isolates tested, culture positivity was 38.29% (224/585), and the prevalence of K. pneumoniae was 25.89% (58/224). On AST, K. pneumoniae exhibited resistance toward carbapenems including ertapenem, meropenem, and imipenem, while it showed the highest susceptibility rate against to tigecycline (93.1%; 54/58). Overall, AST detected 60.34% (35/58) carbapenem-resistant isolates, while the MHT phenotypically confirmed 51.72% (30/58) isolates as carbapenemase-producers and 48.28% (28/58) as carbapenemase nonproducers. On subsequent screening for resistant genes among carbapenemase-producers by PCR assay, 80% (24/30) and 3.33% (1/30) isolates were found to be positive for blaNDM-1 and blaVIM, respectively. In the same assay among 28 carbapenem nonproducing isolates, 9 (32.14%) isolates were positive for blaNDM-1 gene while none of them were tested positive for blaVIM gene. CONCLUSIONS: Molecular detection of resistant genes provides greater specificity and sensitivity than those with conventional techniques, thus aiding in accurate identification of antimicrobial resistance and clinical management of the disease.

Efficacy of Primaquine in Preventing Short- and Long-Latency Plasmodium vivax Relapses in Nepal.

BACKGROUND: Plasmodium vivax is the main cause of malaria in Nepal. Relapse patterns have not been characterized previously. METHODS: Patients with P. vivax malaria were randomized to receive chloroquine (CQ; 25 mg base/kg given over 3 days) alone or together with primaquine (PQ; 0.25 mg base/kg/day for 14 days) and followed intensively for 1 month, then at 1- to 2-month intervals for 1 year. Parasite isolates were genotyped. RESULTS: One hundred and one (49%) patients received CQ and 105 (51%) received CQ + PQ. In the CQ + PQ arm, there were 3 (4.1%) recurrences in the 73 patients who completed 1 year of follow-up compared with 22 of 78 (28.2%) in the CQ-only arm (risk ratio, 0.146 [95% confidence interval, .046-.467]; P < .0001). Microsatellite genotyping showed relatively high P. vivax genetic diversity (mean heterozygosity, 0.843 [range 0.570-0.989] with low multiplicity of infection (mean, 1.05) reflecting a low transmission preelimination setting. Of the 12 genetically homologous relapses, 5 (42%) occurred in a cluster after 9 months, indicating long latency. CONCLUSIONS: Although there may be emerging CQ resistance, the combination of CQ and the standard-dose 14-day PQ regimen is highly efficacious in providing radical cure of short- and long-latency P. vivax malaria in Nepal.

Designing malaria surveillance strategies for mobile and migrant populations in Nepal: a mixed-methods study.

BACKGROUND: As malaria cases have declined throughout Nepal, imported cases comprise an increasing share of the remaining malaria caseload, yet how to effectively target mobile and migrant populations (MMPs) at greatest risk is not well understood. This formative research aimed to confirm the link between imported and indigenous cases, characterize high-risk MMPs, and identify opportunities to adapt surveillance and intervention strategies to them. METHODS: The study used a mixed-methods approach in three districts in far and mid-western Nepal, including (i) a retrospective analysis of passive surveillance data, (ii) a quantitative health facility-based survey of imported cases and their MMP social contacts recruited by peer-referral, and (iii) focus group (FG) discussions and key informant interviews (KIIs) with a subset of survey participants. Retrospective case data were summarised and the association between monthly indigenous case counts and importation rates in the previous month was investigated using Bayesian spatio-temporal regression models. Quantitative data from structured interviews were summarised to develop profiles of imported cases and MMP contacts, including travel characteristics and malaria knowledge, attitudes and practice. Descriptive statistics of the size of cases' MMP social networks are presented as a measure of potential programme reach. To explore opportunities and barriers for targeted malaria surveillance, data from FGs and KIIs were formally analysed using a thematic content analysis approach. RESULTS: More than half (54.1%) of malaria cases between 2013 and 2016 were classified as imported and there was a positive association between monthly indigenous cases (incidence rate ratio (IRR) 1.02 95% CI 1.01-1.03) and the previous month's case importation rate. High-risk MMPs were identified as predominantly adult male labourers, who travel to malaria endemic areas of India, often lack a basic understanding of malaria transmission and prevention, rarely use ITNs while travelling and tend not to seek treatment when ill or prefer informal private providers. Important obstacles were identified to accessing Nepali MMPs at border crossings and at workplaces within India. However, strong social connectivity during travel and while in India, as well as return to Nepal for large seasonal festivals, provide opportunities for peer-referral-based and venue-based surveillance and intervention approaches, respectively. CONCLUSIONS: Population mobility and imported malaria cases from India may help to drive local transmission in border areas of far and mid-western Nepal. Enhanced surveillance targeting high-risk MMP subgroups would improve early malaria diagnosis and treatment, as well as provide a platform for education and intervention campaigns. A combination of community-based approaches is likely necessary to achieve malaria elimination in Nepal.

Quantifying primaquine effectiveness and improving adherence: a round table discussion of the APMEN Vivax Working Group.

The goal to eliminate malaria from the Asia-Pacific by 2030 will require the safe and widespread delivery of effective radical cure of malaria. In October 2017, the Asia Pacific Malaria Elimination Network Vivax Working Group met to discuss the impediments to primaquine (PQ) radical cure, how these can be overcome and the methodological difficulties in assessing clinical effectiveness of radical cure. The salient discussions of this meeting which involved 110 representatives from 18 partner countries and 21 institutional partner organizations are reported. Context specific strategies to improve adherence are needed to increase understanding and awareness of PQ within affected communities; these must include education and health promotion programs. Lessons learned from other disease programs highlight that a package of approaches has the greatest potential to change patient and prescriber habits, however optimizing the components of this approach and quantifying their effectiveness is challenging. In a trial setting, the reactivity of participants results in patients altering their behaviour and creates inherent bias. Although bias can be reduced by integrating data collection into the routine health care and surveillance systems, this comes at a cost of decreasing the detection of clinical outcomes. Measuring adherence and the factors that relate to it, also requires an in-depth understanding of the context and the underlying sociocultural logic that supports it. Reaching the elimination goal will require innovative approaches to improve radical cure for vivax malaria, as well as the methods to evaluate its effectiveness.

Glucose-6-phosphate dehydrogenase deficiency in people living in malaria endemic districts of Nepal.

BACKGROUND: Glucose-6-phosphate dehydrogenase (G6PD) is a rate limiting enzyme of the pentose phosphate pathway and is closely associated with the haemolytic disorders among patients receiving anti-malarial drugs, such as primaquine. G6PD deficiency (G6PDd) is an impending factor for radical treatment of malaria which affects the clearance of gametocytes from the blood and subsequent delay in the achievement of malaria elimination. The main objective of this study was to assess the prevalence of G6PD deficiency in six malaria endemic districts in Southern Nepal. METHODS: A cross-sectional population based prevalence survey was conducted in six malaria endemic districts of Nepal, during April-Dec 2013. A total of 1341 blood samples were tested for G6PDd using two different rapid diagnostic test kits (Binax-Now® and Care Start™). Equal proportions of participants from each district (n ≥ 200) were enrolled considering ethnic and demographic representation of the population groups. RESULTS: Out of total 1341 blood specimens collected from six districts, the overall prevalence of G6PDd was 97/1341; 7.23% on Binax Now and 81/1341; 6.0% on Care Start test. Higher prevalence was observed in male than females [Binax Now: male 10.2%; 53/521 versus female 5.4%; 44/820 (p = 0.003) and Care Start: male 8.4%; 44/521 versus female 4.5%; 37/820 (p = 0.003)]. G6PDd was higher in ethnic groups Rajbanshi (11.7%; 19/162) and Tharu (5.6%; 56/1005) (p = 0.006), major inhabitant of the endemic districts. Higher prevalence of G6PDd was found in Jhapa (22/224; 9.8%) and Morang districts (18/225; 8%) (p = 0.031). In a multivariate analysis, male were found at more risk for G6PDd than females, on Binax test (aOR = 1.97; CI 1.28-3.03; p = 0.002) and Care Start test (aOR = 1.86; CI 1.16-2.97; p = 0.009). CONCLUSIONS: The higher prevalence of G6PDd in certain ethnic group, gender and geographical region clearly demonstrates clustering of the cases and ascertained the risk groups within the population. This is the first study in Nepal which identified the vulnerable population groups for G6PDd in malaria endemic districts. The finding of this study warrants the need for G6PDd testing in vulnerable population groups in endemic districts, and also facilitates use of primaquine in mass supporting timely progress for malaria elimination.

Productivity and profitability of black rice as affected by transplanting methods and crop geometry.

Black rice is a highly nutritious cereal that has been introduced to Nepal recently. Due to its late introduction, only a few agronomic research have been conducted so far. Hence, farmers are not aware about the appropriate transplanting methods and cropping geometry for profitable black rice cultivation. To fulfill the research gap and to establish a basic benchmark for further studies, the research focuses on responses of two black rice genotypes at different transplanting methods and cropping geometry. The profitability analysis with respect to transplanting methods and cropping geometry revealed, transplanting 21 days old seedlings with any geometrical pattern would yield and profit more as compared to SRI. Similarly, farmers can get a highest net revenue of 9379.3 $ at the B/C ratio of 12.07 from fine black rice as compared to coarse black rice that has a net revenue of 4485.7 $ at the B/C of 7.38. The highest productivity (2.70 t ha-1), net revenue (6018.5 $), and B/C ratio (13.7) were observed at the crop geometry of 20 cm × 15 cm for coarse black rice. Whereas, the highest yield (4.60 t ha-1), net revenue (10889.8 $), and B/C ratio (19.5) was observed in 20 cm × 10 cm for fine black rice. The higher net revenue and B/C ratio of premium black rice genotypes was due to their higher market price. The correlation analysis suggested tillering index (Ti) and net biomass accumulated up to 60 days after transplanting (DAT) had the highest positive correlation with yield of both black rice genotypes. Hence, the authors recommend researchers to work on additional agronomic practices that enhance the tillering index and net biomass production up to 60 DAT considering transplanting methods yield more as compared to SRI and crop geometry of 20 cm × 15 cm and 20 cm × 10 cm are the most productive and profitable cropping geometry for coarse and fine black rice genotypes, respectively.

Extended Spectrum Beta Lactamase Escherichia Coli in Bagmati River, Kathmandu Valley.

BACKGROUND: Antimicrobial resistance organisms in the peripheral communities of an environment can be predicted by the presence of extended-spectrum beta-lactamase Escherichia coli in that environment. The close connectivity between humans and water sources can facilitate the entry of antimicrobial resistant organisms into the human ecosystem. The aim of this study was to assess beta lactamase producing Escherichia coli from Bagmati river within Kathmandu valley. METHODS: In the year 2020, a cross-sectional study was conducted on water samples collected from 66 locations along the Bagmati River. Coliforms were isolated by five tubes dilution method and identified by cultural and biochemical tests. Further Escherichia coli was isolated in eosin methylene blue agar at 44.5 °C. Antibiotic susceptibility test was performed by Kirby Bauer disk diffusion methods. Beta lactamase gene types were detected by using conventional multiplex polymerase chain reaction. RESULTS: A total of 615 bacterial isolates were identified among which 39 % (n=241) were Escherichia coli. Extended spectrum beta lactamase producing Escherichia coli was confirmed in 16.6 % (40/241) of total Escherichia coli isolates. Among 66 sites this isolate was detected in 26 (40 %) sampling sites excluding upstream regions. All the Escherichia coli isolates were multidrug resistance showing higher percentage (>99 %) of resistant for penicillin, tetracycline and erythromycin antibiotics. There were significant differences in resistance rate for cefotaxime and ceftazidime by extended spectrum beta lactamase producing and non-producing Escherichia coli (p<0.05). CONCLUSIONS: Presence of multidrug resistance extended spectrum beta lactamase producing Escherichia coli in river streams suggests the chances of circulating within river system and hence transmitting in human community. KEY WORDS: Bagmati river; drug resistance; escherichia coli; human.

Efficacy of Native Bacillus thuringiensis against Mosquito Vector.

BACKGROUND: Larval source management is an effective measure to control mosquito-borne diseases. Bacillus thuringiensis produces specific insecticidal crystal proteins toxic to mosquito larvae. In many parts of the South East Asian region, Bacillus thuringiensis is used for larval source management. In Nepal, larvicidal Bacillus thuringiensis is not available. The study aims to isolate larvicidal Bacillus thuringiensis from soil samples of Nepal to control mosquitoes. METHODS: Native Bacillus thuringiensis was obtained from soil samples by the acetate selection method. It was identified by observing crystal protein with Coomassie Brilliant Blue stain in a light microscope. The mosquito larvae were collected from different breeding habitats. A preliminary bioassay was performed by inoculating three loopful of 48 hours culture of spherical crystal protein producing Bacillus thuringiensis in a plastic cup containing 25 larvae and 100 ml of sterile distilled water. The cup was incubated at room temperature for 24 hours to observe the mortality of larvae. Further selective bioassay was performed with the isolate which showed 100% mortality, as described above in four replicates along with the negative and positive control. RESULTS: Out of 1385 Bacillus thuringiensis obtained from 454 soil samples, 766 (55.30%) were spherical crystal protein producers, among them, a single strain (14P2A) showed 100% mortality against mosquito larvae. The lethal concentration doses required to kill 50% and 90% of the larval population were 32.35 and 46.77 Parts per million respectively. CONCLUSIONS: The native Bacillus thuringiensis produces the crystal protein effective in killing mosquito larvae. The native Bacillus thuringiensis should be included as a tool to control mosquito-borne diseases in Nepal.

Metallo-ß-lactamase-producing Pseudomonas aeruginosa Isolates from Two Tertiary Care Centres in a District of Nepal: A Descriptive Cross-sectional Study.

INTRODUCTION: Pseudomonas aeruginosa isolates producing metallo-ß-lactamase have caused nosocomial outbreaks, severe infections, and ineffective carbapenem therapy worldwide since 1991. Due to their prevalence, hospital infection control techniques are difficult. This study aimed to find out the prevalence of metallo-ß-lactamase among P. aeruginosa isolates from two tertiary care hospitals in Kathmandu. METHODS: A descriptive cross-sectional study was conducted at the Department of Microbiology and Department of Pathology of two tertiary care centres in Kathmandu from 7 December 2021 to 6 April 2023, after receiving ethical approval from the Ethical Review Board. Isolated strains were identified and tested for antibiotic susceptibility by modified Kirby-Bauer Methods. Metallo-ß-lactamase presence was confirmed using an imipenem-imipenem/ ethylenediaminetetraacetic acid disc. A convenience sampling method was used. The point estimate was calculated at 95% Confidence Interval. RESULTS: Among 255, Pseudomanas aeruginosa isolates, the distribution of metallo-ß-lactamase-producing Pseudomanas aeruginosa was 103 (40.39%) (34.32-46.69 at 95% Confidence Interval). Multidrug resistance categories included multidrug resistance 74 (71.80%), extensively drug resistance 32 (31.10%), P. aeruginosa difficult-to-treat 16 (15.53%) and carbapenem-resistant P. aeruginosa was determined to be 82 (79.60%). CONCLUSIONS: The study found a high prevalence of metallo-ß-lactamase-producing Pseudomanas aeruginosa isolates, requiring early identification, infection control measures, and an all-inclusive antimicrobial therapy protocol to reduce their spread in medical settings.

Field assessment of the operating procedures of a semi-quantitative G6PD Biosensor to improve repeatability of routine testing.

In remote communities, diagnosis of G6PD deficiency is challenging. We assessed the impact of modified test procedures and delayed testing for the point-of-care diagnostic STANDARD G6PD (SDBiosensor, RoK), and evaluated recommended cut-offs. We tested capillary blood from fingerpricks (Standard Method) and a microtainer (BD, USA; Method 1), venous blood from a vacutainer (BD, USA; Method 2), varied sample application methods (Methods 3), and used micropipettes rather than the test's single-use pipette (Method 4). Repeatability was assessed by comparing median differences between paired measurements. All methods were tested 20 times under laboratory conditions on three volunteers. The Standard Method and the method with best repeatability were tested in Indonesia and Nepal. In Indonesia 60 participants were tested in duplicate by both methods, in Nepal 120 participants were tested in duplicate by either method. The adjusted male median (AMM) of the Biosensor Standard Method readings was defined as 100% activity. In Indonesia, the difference between paired readings of the Standard and modified methods was compared to assess the impact of delayed testing. In the pilot study repeatability didn't differ significantly (p = 0.381); Method 3 showed lowest variability. One Nepalese participant had <30% activity, one Indonesian and 10 Nepalese participants had intermediate activity (≥30% to <70% activity). Repeatability didn't differ significantly in Indonesia (Standard: 0.2U/gHb [IQR: 0.1-0.4]; Method 3: 0.3U/gHb [IQR: 0.1-0.5]; p = 0.425) or Nepal (Standard: 0.4U/gHb [IQR: 0.2-0.6]; Method 3: 0.3U/gHb [IQR: 0.1-0.6]; p = 0.330). Median G6PD measurements by Method 3 were 0.4U/gHb (IQR: -0.2 to 0.7, p = 0.005) higher after a 5-hour delay compared to the Standard Method. The definition of 100% activity by the Standard Method matched the manufacturer-recommended cut-off for 70% activity. We couldn't improve repeatability. Delays of up to 5 hours didn't result in a clinically relevant difference in measured G6PD activity. The manufacturer's recommended cut-off for intermediate deficiency is conservative.

The readiness of the Asian research ethics committees in responding to the COVID-19 pandemic: A multi-country survey.

Background: COVID-19 is a highly challenging infectious disease. Research ethics committees (RECs) have challenges reviewing research on this new pandemic disease under a tight timeline and public pressure. This study aimed to assess RECs' responses and review during the outbreak in seven Asian countries where the Strategic Initiative for Developing Capacity in Ethical Review (SIDCER) networks are active. Methods: The online survey was conducted in seven Asian countries from April to August 2021. Two sets of online questionnaires were developed, one set for the chairs/secretaries and another set for the REC members.The REC profiles obtained from the REC members are descriptive in nature. Data from the chairs/secretaries were compared between the RECs with external quality assessment (SIDCER-Recognized RECs, SR-RECs) and non-external quality assessment (Non-SIDCER-Recognized RECs, NSR-RECs) and analyzed using a Chi-squared test. Results: A total of 688 REC members and 197 REC chairs/secretaries participated in the survey. Most RECs have standard operating procedures (SOPs), and have experience in reviewing all types of protocols, but 18.1% had no experience reviewing COVID-19 protocols. Most REC members need specific training on reviewing COVID-19 protocols (93%). In response to the outbreak, RECs used online reviews, increased meeting frequency and single/central REC. All SR-RECs had a member composition as required by the World Health Organisation ethics guidelines, while some NSR-RECs lacked non-affiliated and/or layperson members. SR-RECs reviewed more COVID-related product development protocols and indicated challenges in reviewing risk/benefit and vulnerability (0.010), informed consent form (0.002), and privacy and confidentiality (P = 0.020) than NSR-RECs. Conclusions: Surveyed RECs had a general knowledge of REC operation and played a significant role in reviewing COVID-19-related product development protocols. Having active networks of RECs across regions to share updated information and resources could be one of the strategies to promote readiness for future public health emergencies.

Multi-drug Resistance, ß-Lactamases Production, and Coexistence of bla NDM-1 and mcr-1 in Escherichia coli Clinical Isolates From a Referral Hospital in Kathmandu, Nepal.

The ability of pathogenic Escherichia coli to produce carbapenemase enzymes is a characteristic that allows them to resist various antibiotics, including last-resort antibiotics like colistin and carbapenem. Our objectives were to identify rapidly developing antibiotic resistance (AR), assess ß-lactamases production, and detect mcr-1 and bla NDM-1 genes in the isolates. A prospective cross-sectional study was carried out in a referral hospital located in Kathmandu from November 2019 to December 2020 using standard laboratory and molecular protocols. Among 77 total E. coli isolates, 64 (83.1%) of them were categorized as MDR. Phenotypically 13 (20.3%) colistin-resistant, 30 (46.9%) ESBL and 8 (12.5%) AmpC producers, and 5 (7.8%) ESBL/AmpC co-producers were distributed among MDR-E. coli. Minimum inhibitory concentrations (MIC) against the majority of MDR isolates were exhibited at 1 g/L. Of these 77 E. coli isolates, 24 (31.2%) were carbapenem-resistant. Among these carbapenem-resistant bacteria, 11 (45.9%) isolates were reported to be colistin-resistant, while 15 (62.5%) and 2 (8.3%) were MBL and KPC producers, respectively. Out of 15 MBL producers, 6 (40%) harbored bla NDM-1, and 8 (61.5%) out of 13 colistin-resistant pathogens possessed mcr-1. The resistance by colistin- and carbapenem were statistically associated (P < .001). However, only 2 (18.2%) of the co-resistant bacteria were found to have both genes. Our study revealed the highly prevalent MDR and the carbapenem-resistant E. coli and emphasized that the pathogens possess a wide range of capabilities to synthesize ß-lactamases. These findings could assist to expand the understanding of AR in terms of enzyme production.

Genotypically Confirmed Vancomycin-Resistant Staphylococcus aureus With vanB Gene Among Clinical Isolates in Kathmandu.

Purpose: Methicillin-resistant Staphylococcus aureus, a common bacterial pathogen causes various infections. The acquisition of various antimicrobial-resistant genes in S. aureus has led to the transformation of this bacterium into a superbug. Vancomycin resistance among MRSA isolates is an emerging threat in empirical therapy of various infections. The study was hence aimed to find out the susceptibility status of S. aureus isolates toward vancomycin and detect mecA, vanA, and vanB genes among the isolates. Methods: A total of 1245 clinical samples from the participants attending a tertiary care hospital in Kathmandu were processed. S. aureus isolated from the samples were subjected to antibiotic susceptibility patterns using the modified Kirby-Bauer disk diffusion method. Agar dilution method was used to determine the minimum inhibitory concentration of vancomycin. The antibiotic-resistant genes such as mecA, vanA, and vanB among S. aureus isolates were screened by a conventional polymerase chain reaction. Results: Of 1245 samples, 80 S. aureus were identified. Out of which, 47.5% (38/80) were phenotypically confirmed MRSA isolates. mecA gene was detected in 84.2% (32/38) of MRSA isolates. 10.5% (4/38) were confirmed as vancomycin-intermediate S. aureus (VISA) by MIC determination. None of the isolates was positive for the vanA gene; however, 2 isolates were found to possess the vanB gene. The 2 isolates have vancomycin MIC breakpoints of 4 to 8 µg/mL. Conclusion: There might be a spreading of vancomycin resistance among S. aureus, creating serious public health problems. Therefore, measures to limit vancomycin resistance should be considered in healthcare facilities as immediately as possible.

Beta-Lactamase-Producing Gram-Negative Bacterial Isolates Among the Patients Attending a Tertiary Care Hospital, Kathmandu, Nepal.

Over the times, carbapenems have been the choice of drug for treating multidrug-resistant (MDR) and extended spectrum beta-lactamase (ESBL)-producing organisms. The current study aimed at determining the occurrence of metallo beta-lactamase (MBL) and AmpC beta-lactamase (ABL) in gram negative bacteria isolated from clinical samples. A cross-sectional study was conducted amongst the patients visiting Manmohan Memorial Medical College and Teaching Hospital (MMTH), Kathmandu, Nepal from August 2017 to January 2018. A total of 4351 samples including urine, pus, wound swab, endotracheal tip, catheter tip, and blood were collected from the patients and processed by standard conventional microbiological methods. Antibiotic susceptibility testing (AST) of the isolates was performed by Kirby-Bauer disk diffusion method. Double disc synergy test was performed on carbapenem resistant organisms to detect production of MBL and inhibitor-based test was used for the detection of ABL production. Of the 4351 samples, 421 bacterial isolates belonging to 16 different genera were recovered, of which 303 (71.97%) were Gram negative bacilli (GNB). E. coli (189/303) and S. aureus (80/118) were the most prevalent among gram negatives and gram positives, respectively. Bacterial incidence was found significantly associated with gender, specimen type, and the department where the patients were enrolled. Colistin-sulfate and polymycin-B were the most effective drug against GNB, whereas imipenem against gram positives. Prevalence of MDR and methicillin-resistant S. aureus (MRSA) was 35.15% and 60%, respectively. The prevalence of MBL and ABL-producing isolate was 11(3.6%) and 13(4.3%), respectively. Pseudomonas aeruginosa (5/11) and E. coli (9/13) were the major MBL and ABL producers, respectively. MBL and ABL production was found to be significantly associated with the age of the patient and the specimen type. A regular antibiotic surveillance activity with screening for MBL and ABL-producing bacterial isolates in the hospital settings to curb the incidence and transmission of such difficult-to-treat pathogens.