The American Journal of Tropical Medicine and Hygiene - Current Issue

Medication adherence is critical to the effectiveness of benznidazole (BZ) therapy for the treatment of Chagas disease. Assessing BZ adherence using traditional plasma sampling methods presents numerous challenges in resource-limited settings. Dried blood spot (DBS) sampling of BZ can be used to overcome logistical barriers and provides a less invasive method for assessing BZ levels. A BZ DBS assay using liquid chromatography-tandem mass spectrometry was developed and applied to a clinical study of infants and children being treated with BZ for Trypanosoma cruzi infection in Argentina. The assay was validated over a concentration range of 9.8–5,000 ng/mL. Inter-assay and intra-assay measures ranged from −2.9% to 2.7% and 0.5% to 8.3% for accuracy and from 3.5% to 12% and 1.6% to 13.6% for precision, respectively. The mean recovery of BZ was greater than 91%. Partitioning ratios for DBSs/plasma ranged from 0.95 to 1.02. A cohort of 10 infants and six children with T. cruzi infection being treated with BZ had median BZ concentrations of 1.2 (IQR 0.29, 2.14) µg/mL with seven of 65 (11%) samples above the BZ treatment goal of 3 μg/mL for adults. The reported DBS assay is a simple and accurate method for the quantitative measurement of BZ that can be applied to facilitate urgently needed clinical studies of BZ for the treatment of Chagas disease and assess BZ adherence in resource-limited settings.

We report the case of a 64-year-old woman found to have urban-acquired Trypanosoma brucei (T.b.) gambiense human African trypanosomiasis (HAT) as the cause of sustained fever starting 9 months after returning to Canada from Democratic Republic of the Congo, in the context of concomitant multiple myeloma and HIV-1 coinfection. Approaches for the management of both clinical stages of T.b. gambiense HAT are well defined for endemic settings using current diagnostics and treatments. However, few data inform the diagnosis and management of patients with bone marrow suppression from active malignancy, recent anticancer therapy, or HIV coinfection. We discuss the implications of immunosuppression for diagnosis and management of T.b. gambiense HAT.

Malaria transmission is dependent on the density and distribution of mosquito vectors, but drivers of vector abundance have not been adequately studied across a range of transmission settings. To inform intervention strategies for high-burden areas, further investigation is needed to identify predictors of vector abundance. Active household (HH) surveillance was conducted in Nchelenge district, Luapula Province, northern Zambia, a high-transmission setting with limited impact of malaria control. Between April 2012 and July 2017, mosquitoes were collected indoors during HH visits using CDC light traps. Demographic, environmental, and climatological correlates of vector abundance were identified using log-binomial regression models with robust standard errors. The primary malaria vectors in this setting were Anopheles funestus sensu stricto (s.s.) and Anopheles gambiae s.s. Anopheles funestus predominated in both seasons, with a peak in the dry season. Anopheles gambiae peaked at lower numbers in the rainy season. Environmental, climatic, and demographic factors were correlated with HH vector abundance. Higher vector counts were found in rural areas with low population density and among HHs close to roads and small streams. Vector counts were lower with increasing elevation and slope. Anopheles funestus was negatively associated with rainfall at lags of 2–6 weeks, and An. gambiae was positively associated with rainfall at lags of 3–10 weeks. Both vectors had varying relationships with temperature. These results suggest that malaria vector control in Nchelenge district should occur throughout the year, with an increased focus on dry-season transmission and rural areas.

Test positivity rate (TPR)—confirmed cases per 100 suspected cases tested, and test-confirmed malaria case rate (IR)—cases per 1,000 population, are common indicators used routinely for malaria surveillance. However, few studies have explored relationships between these indicators over time and space. We studied the relationship between these indicators in children aged < 11 years presenting with suspected malaria to the outpatient departments of level IV health centers in Nagongera, Kihihi, and Walukuba in Uganda from October 2011 to June 2016. We evaluated trends in indicators over time and space, and explored associations using multivariable regression models. Overall, 65,710 participants visited the three clinics. Pairwise comparisons of TPR and IR by month showed similar trends, particularly for TPRs < 50% and during low-transmission seasons, but by village, the relationship was complex. Village mean annual TPRs remained constant, whereas IRs drastically declined with increasing distance from the health center. Villages that were furthest away from the health centers (fourth quartile for distance) had significantly lower IRs than nearby villages (first quartile), with an incidence rate ratio of 0.40 in Nagongera (95% CI: 0.23–0.63; P = 0.001), 0.55 in Kihihi (0.40–0.75; P < 0.001), and 0.25 in Walukuba (0.12–0.51; P < 0.001). Regression analysis results emphasized a nonlinear (cubic) relationship between TPR and IR, after accounting for month, village, season, and demographic factors. Results show that the two indicators are highly relevant for monitoring malaria burden. However, interpretation differs with TPR primarily indicating demand for malaria treatment resources and IR indicating malaria risk among health facility catchment populations.

Data on fever and malaria cases reported by health facilities are used for tracking incidence and quantification of malaria commodity needs in Guinea. Periodic assessments of the quality of malaria case management and routine data are a critical activity for the malaria program. In May–June 2018, survey teams visited 126 health facilities in six health districts purposefully selected to represent a spectrum (Stratum 1—high, Stratum 2—intermediate, and Stratum 3—low) of perceived quality of case management and reporting, as assessed from an a priori analysis of routine data. Surveyors performed exit interviews with 939 outpatients and compared results with registry data for interviewed patients. Availability of rapid diagnostic tests (RDTs) and artemisinin-based combination therapies (ACTs) was 100% in Strata 1 and 2, compared with 82% (95% CI: 63–92%) for RDTs and 86% (68–95%) for any formulation of ACT in Stratum 3. Correct case management for suspect malaria cases was 85% in both Stratum 1 (95% CI: 78–90%) and Stratum 2 (79–89%), but only 52% (37–67%) in Stratum 3. Concordance between exit interviews and registry entries for key malaria indicators was significantly higher in Strata 1 and 2 than in Stratum 3. Both adherence to national guidelines for testing and treatment and data quality were high in Strata 1 and 2, but substandard in Stratum 3. The survey results reflected the trends seen in the routine data, suggesting that analysis of routine data can identify areas requiring more attention to improve malaria case management and reporting.

Intrahepatic pseudoaneurysm (IHPA) is generally iatrogenic, and it warrants urgent management because of its reportedly significant risk of rupture leading to considerable mortality. Intrahepatic pseudoaneurysm caused by amebic liver abscess (ALA) is exceedingly rare with fewer than seven published reports. All reported symptomatic cases have been treated with hepatic artery embolization; therefore, little is known about its natural course and effect of abscess drainage on its outcome. We report the first case of symptomatic intracavitary IHPA caused by ALA in which ultrasound-guided percutaneous catheter drainage of the abscess resulted in the spontaneous resolution of the IHPA.

Cryptosporidium is a parasite that causes watery diarrhea among both children and adults. However, because many physicians do not routinely ask for Cryptosporidium diagnostic test, cryptosporidiosis prevalence is likely underestimated. The current study investigated the prevalence of cryptosporidiosis among children admitted to the hospital with gastrointestinal symptoms. Stool sample was collected from each child and subjected to routine microbiological culture. Cryptosporidium presence was tested by three different methods: real-time PCR (RT-PCR), quick antigen, and microscopic examination with acid-fast staining. Each positive specimen was further tested with BioFire FilmArray Multiplex PCR (bioMérieux SA, Marcy-l'Etoile, France) to determine that Cryptosporidium is the only pathogen in the sample. Demographic and epidemiological data were collected from the patients’ medical records. Of 291 patient stool samples, nine were positive for Cryptosporidium hominis or Cryptosporidium parvum. The average age of the nine Cryptosporidium-positive cases was 2.3 years, lower than the average age of the study population (4.2 years). Of the positive cases, 66.7% were Arabs and 33.3% were Jews. The main complaint of children with Cryptosporidium was diarrhea. Regarding the laboratory methods for Cryptosporidium identification, of the positive samples, 100% (9/9) were identified by RT-PCR, 88% (8/9) were positive by antigen test, and only 67% (6/9) were positive by microscopic examination with acid-fast staining. Because of the low incidence of Cryptosporidium among patients in our facility, the prevalence of cryptosporidiosis still cannot be established. Nevertheless, among the other pathogens found in stool samples, Cryptosporidium is the second leading cause of hospitalization due to gastrointestinal disease in children in our area.

The CYD-TDV vaccine is licensed in multiple endemic countries based on vaccine efficacy (VE) against symptomatic, virologically confirmed dengue demonstrated in two phase 3 trials (CYD14, 2- to 14-year-olds, Asia; CYD15, 9- to 16-year-olds, Latin America). 50% plaque reduction neutralization test (PRNT50) titers at baseline and month 13 (post-vaccination) were associated with VE and may enable bridging VE to adults. Two phase 2 trials of CYD-TDV measured baseline and month 13 PRNT50 titers: CYD22 (9- to 45-year-olds, Vietnam) and CYD47 (18- to 45-year-olds, India). 50% plaque reduction neutralization test distributions were compared between age cohorts, and four versions of an epidemiological bridging method were used to estimate VE against any serotype (dengue virus [DENV]-Any) and against each serotype over 25 months post first vaccination in a hypothetical CYD14 + CYD15 18- to 45-year-old cohort (bridging population 1) and in the actual CYD47 18- to 45-year-old cohort (bridging population 2). Baseline and month 13 geometric mean PRNT50 titers to each serotype were significantly greater in 18- to 45-year-olds than 9- to 16-year-olds for all comparisons. The four methods estimated VE against DENV-Any at 75.3–86.0% (95% CIs spanning 52.5–100%) for bridging population 1 and 68.4–77.5% (95% CIs spanning 42.3–88.5%) for bridging population 2. The vaccine efficacy against serotype 1, 2, 3, and 4 was estimated at 56.9–76.9%, 68.3–85.8%, 91.4–95.0%, and 93.2–100% (bridging population 1) and 44.5–66.9%, 53.2–69.2%, 79.8–92.0%, and 90.6–95.0% (bridging population 2), respectively; thus, CYD-TDV would likely confer improved efficacy in adults than 9- to 16-year-olds. Using the same methods, we predicted VE against hospitalized DENV-Any over 72 months of follow-up, with estimates 59.1–73.5% (95% CIs spanning 40.9–92.2%) for bridging population 1 and 50.9–65.9% (95% CIs spanning 38.1–82.1%) for bridging population 2.

Faced with the reemergence of yellow fever (YF) in the metropolitan region of São Paulo, Brazil, we developed a retrospective study to describe the cases of YF attended at the Institute of Infectology Emilio Ribas from January to March 2018 and analyze the factors associated with death, from the information obtained in the hospital epidemiological investigation. A total of 72 cases of sylvatic YF were confirmed, with 21 deaths (29.2% lethality rate). Cases were concentrated in males (80.6%) and in the age group of 30 to 59 years (56.9%). Two logistic regression models were performed, with continuous variables adjusted for the time between onset of symptoms and hospitalization. The first model indicated age (odds ratiosadjusted [ORadj]: 1.038; CI 95%: 1.008–1.212), aspartate aminotransferase (AST) (ORadj: 1.038; CI 95%: 1.005–1.072), and creatinine (ORadj: 2.343; CI 95%: 1.205–4.553) were independent factors associated with mortality. The second model indicated age (ORadj: 1.136; CI 95%: 1.013–1.275), alanine aminotransferase (ALT) (ORadj: 1.118; CI 95%: 1.018–1.228), and creatinine (ORadj: 2.835; CI 95%: 1.352–5,941). The risk of death in the model with continuous variables was calculated from the increase of 1 year (age), 1 mg/dL (creatinine), and 100 U/L for AST and ALT. Another logistic regression analysis with dichotomous variables indicated AST > 1,841 IU/L (ORadj: 12.92; CI 95%: 1.50–111.37) and creatinine > 1.2 mg/dL (ORadj: 81.47; CI 95%: 11.33–585.71) as independent factors associated with death. These results may contribute to the appropriate clinical management of patients with YF in health-care services and improve the response to outbreaks and public health emergencies.

Tick-borne encephalitis (TBE) is a viral infectious disease and has become a reemerging public health threat in recent years in northeastern China. However, no studies has characterized the epidemiologic features and explored the spatial dynamics and environmental factors of TBE cases in Jilin Province. In this study, we have described the epidemiological features of 846 reported human TBE cases from 2006 to 2016 in Jilin Province. There was an obvious single peak pattern of TBE cases from May to July in Jilin Province. More than 60% of TBE cases occurred in farmers, and the people in 50- to 59-year-old group had the high incidence of the disease. The results of Getis-Ord Gi* statistics demonstrated that the human TBE cases were more clustered in the northeastern border including Dunhua and Yanji cities and Antu and Wangqing counties, and southern areas including Huinan, Jingyu, Jiangyuan, and Liuhe counties in Jilin Province. We demonstrated that the temporal dynamics of TBE in Jilin was significantly associated with the dynamics of meteorological factors especially after 2009. The results from the auto-logistic regression analysis showed that the percentage coverage of forest, temperature, and autoregressive term were significantly associated with the occurrence of human TBE cases in Jilin Province. Our findings will provide a scientific evidence for the targeted prevention and control programs.

Filoviruses, which include ebolaviruses and marburgvirus, can cause outbreaks of highly lethal hemorrhagic fever. This disease causes significant morbidity and mortality in humans and non-human primates, with human fatality rates reaching 90% during some outbreaks. Currently, there is lack of licensed vaccines or antivirals for these viruses. Since early symptoms of filovirus infection mimic more common diseases, there is a strong unmet public health and biodefense need for broad-spectrum filovirus rapid diagnostics. We have generated a panel of mouse single-chain Fv-antibodies (scFvs) to filovirus glycoproteins (GPs) using cell-free ribosome display and determined their cross-reactivity profiles to all known filovirus species. Two scFvs (4-2 and 22-1) were able to detect all known Ebolavirus and Marburgvirus species. This is the first report on ribosome display scFvs that can detect a broad set of filovirus GPs, which demonstrates the potential for use in diagnostics.

rVSVΔG-ZEBOV-GP vaccine is a live recombinant (r) vesicular stomatitis virus (VSV), where the VSV G protein is replaced with the Zaire Ebola virus (ZEBOV) glycoprotein (GP). For vaccine immunogenicity testing, clinical trial sera collected during an active ZEBOV outbreak underwent gamma irradiation (GI) before testing in biosafety level 2 laboratories to inactivate possible wild-type ZEBOV. Before irradiating pivotal trial samples, two independent studies evaluated the impact of GI (50 kGy) on binding ZEBOV-GP (ELISA) antibodies against rVSVΔG-ZEBOV-GP, using sera from a North American phase 1 study. Gamma irradiation was associated with slightly higher antibody concentrations in pre-vaccination samples and slightly lower concentrations postvaccination. Results indicate that GI is a viable method for treating samples from regions where filoviruses are endemic, with minor effects on antibody titers. The impact of GI on immunogenicity analyses should be considered when interpreting data from irradiated specimens.

In 2016, the World Health Assembly endorsed the Global Health Sector Strategy on Viral Hepatitis, which calls for elimination of hepatitis B virus (HBV) by 2030 (definition: ≤ 0.1% hepatitis B surface antigen [HBsAg] prevalence among children aged 5 years). The burden of chronic HBV infection among children in Haiti is unknown. We conducted a nationally representative cross-sectional serological survey among 5- to 7-year-old children based on a two-stage cluster design with two strata: West (includes metropolitan Port-au-Prince) and non-West (all other departments). We collected demographic, socioeconomic, and vaccination history data and tested for HBsAg using a rapid point-of-care test. We estimated HBsAg prevalence and evaluated the association of HBV infection with vaccination history, demographics, and socioeconomic characteristics. Of the 1,152 children, seven (0.5%, 95% CI: 0.2–1.2) were HBsAg positive. The HBsAg prevalence varied by region (West: 0.1%, 95% CI: 0.01–0.9; non-West: 0.7%, 95% CI: 0.2–1.9) (P = 0.1), gender (males: 0.7%, 95% CI: 0.2–2.4; females: 0.2%, 95% CI: 0.05–1.1) (P = 0.3), and caregiver’s education level (none: 0.8%, 95% CI: 0.2–3.1; some or completed primary: 0.5%, 95% CI: 0.1–1.8; some secondary: 0.4%, 95% CI: 0.1–1.8; secondary and higher: 0.0%, 95% CI: 0–0), although the differences were not statistically significant. None of the HBsAg-positive children had documented vaccination with hepatitis B vaccine (HepB). Haiti’s chronic HBV infection prevalence among children is low; however, it is above the elimination target. To reach elimination, Haiti needs to achieve high coverage with the three HepB doses and introduce a HepB birth dose.

The albumin–bilirubin (ALBI) score was originally established to stratify prognosis in patients with cirrhosis. The diagnostic accuracy of ALBI score in liver fibrosis staging in patients with chronic hepatitis B remains to be investigated. The present retrospective study, therefore, aimed to evaluate the ability of this score to stage liver fibrosis in these patients. Briefly, consecutive patients with hepatitis B virus (HBV) infection who underwent liver biopsy examinations in Kagawa University Hospital were enrolled. Liver fibrosis stage was assessed using a modified Meta-Analysis of Histological Data in Viral Hepatitis score. Albumin–bilirubin scores were calculated according to the following equation: (log10 total bilirubin [T-Bil] × 0.66) + (albumin [Alb] × −0.085). A total of 91 patients were enrolled in this study. Albumin–bilirubin score was able to differentiate stage 4 from stage 3 fibrosis (P < 0.05). When an ALBI score of −2.190 was adopted as the cutoff value for differentiating stage 4 from stages 1–3, the sensitivity, specificity, and positive likelihood ratio were 85.7%, 74.0%, and 3.300, respectively. Kaplan–Meier analysis showed that baseline ALBI scores < −2.190 correlated with better hepatocellular carcinoma (HCC)–free survival (P < 0.05). In conclusion, ALBI score can be used for liver fibrosis staging in Japanese chronic hepatitis B patients and can help distinguish cirrhotic from non-cirrhotic status. Furthermore, ALBI score was useful as a prognosis biomarker in our patients, with smaller ALBI scores predicting better HCC-free survival. Because calculating ALBI score is easy using serum T-Bil and Alb alone, ALBI score will help clinicians with decision-making in management of HBV-infected patients.

Hepatitis B virus (HBV) is endemic throughout Africa, but its prevalence in the Democratic Republic of the Congo (DRC) is incompletely understood. We used dried blood spot (DBS) samples from the 2013 to 2014 Demographic and Health Survey in the DRC to measure the prevalence of HBV using the Abbott ARCHITECT HBV surface antigen (HBsAg) qualitative assay. We then attempted to sequence and genotype HBsAg-positive samples. The weighted national prevalence of HBV was 3.3% (95% CI: 1.8–4.7%), with a prevalence of 2.2% (95% CI: 0.3–4.1%) among children. Hepatitis B virus cases occurred countrywide and across age strata. Genotype E predominated (60%), and we found a unique cluster of genotype A isolates (30%). In conclusion, DBS-based HBsAg testing from a nationally representative survey found that HBV is common and widely distributed among Congolese adults and children. The distribution of cases across ages suggests ongoing transmission and underscores the need for additional interventions to prevent HBV infection.

Hepatitis E virus (HEV) infection is associated with a high fatality rate among pregnant women, and gestational complications have been reported among pregnant women infected with hepatitis A virus (HAV). The aim of this study was to determine the seroprevalence of HAV and HEV infections among pregnant women in Haiti. We stratified the population (n = 1,307) between West and non-West regions. Specimens were tested for total HAV antibody (anti-HAV), and IgM and IgG HEV antibody (anti-HEV). Overall, 96.8% pregnant women were positive for total anti-HAV, 10.3% for IgG anti-HEV, and 0.3% for IgM anti-HEV. The prevalence of IgG anti-HEV in the non-West region (12.3%) was significantly greater than that in the West region (5.3%) (P < 0.0001). Most pregnant women in Haiti had evidence of past exposure and immunity to HAV. The non-West region had a higher prevalence of HEV. Improvement in water and sanitation will help in the prevention and control of these infections in Haiti.

Japanese encephalitis virus (JEV) is the most common cause of viral encephalitis in Asia, and it is increasingly a global public health concern because of its recent geographic expansion. Although commercial vaccines are available and used in some endemic countries, JEV continues to cause illness, with more than 60,000 cases reported annually. To develop a reproducible positive control antibody useable in diagnosis of JEV infections, murine hybridomas were developed from mice inoculated with a combination of IXIARO JEV vaccine and JEV domain III of the envelope protein (E-DIII). Monoclonal antibodies (MAbs) were characterized for their ability to neutralize virus in vitro. Monoclonal antibody 17BD3-2 was found to be JEV specific and highly neutralizing, with a plaque reduction neutralization test (PRNT)90 endpoint titer of 1.25 μg/mL. The functional epitopes were mapped using virus neutralization escape variants to amino acid residues S309, K312, and G333 in E-DIII. This MAb may be substituted for human immune sera used as a positive control in PRNT for distribution to public health laboratories worldwide in potential future outbreaks of JEV.

Endemic transmission of measles has been reestablished in Venezuela, and outbreaks of diphtheria remain ongoing across Latin America (LA). Hence, a large cross-sectional population-based serosurveillance study was conducted on Bonaire, one of the Dutch Leeward Antilles, to assess specific age and population groups at risk. Participants (aged 0–90 years) donated a blood sample and completed a questionnaire (n = 1,129). Antibodies against measles and diphtheria were tested using bead-based multiplex immunoassays. Our data revealed that immunity against measles is suboptimal, especially for those aged less than 5 years from Suriname, Aruba, and former Dutch Antilles (SADA), and adolescents from LA; and against diphtheria for persons aged more than 30 years, particularly among females and residents from SADA and LA. As refugees arrive persistently, health authorities on the Dutch Leeward Antilles should be on alert to detect early cases and prevent subsequent transmission. Ultimately, there is an urgent need for serosurveillance studies in the Caribbean region.

Anemia is known to impact a child’s growth and development, but not all anemias are caused by iron deficiency, and the CDC and WHO have emphasized investigating other contributors to anemia. This cross-sectional sub-study of a 2012–2016 maternal-child cohort in coastal Kenya evaluated 244 children and found 185 (76%) to have been anemic on at least one time point since birth. At the time of assessment in 2016, evaluation included a complete blood count, nutritional assessment, and testing for parasitic infections, focusing on the primary outcome of anemia, defined as hemoglobin (Hb) < 11 g/dL. The average age at assessment was 20.5 ± 7 months. Ninety-five percent had a lifetime average Hb in the anemic range. Adjusting for age and gender, prior or current malaria infection (prior: Hb β = −0.99, 95% CI: −1.49 to −0.49, P = 0.01), or having any current infection with hookworm, Trichuris, Strongyloides, Ascaris, and/or malaria (β = −0.84, 95% CI: −1.36 to −0.33, P = 0.01) was associated with decreased current Hb. Nutritional evaluation revealed that children with a declining Hb ate fewer vitamin-A–rich vegetables per week (P = 0.01) or eggs (P = 0.01), drank more milk (P = 0.07), and ate more bread (P = 0.01), and were more likely to live in a household that experienced food shortage (P = 0.05). The high prevalence of anemia, polyparasitism, and dietary insufficiency among children in rural coastal Kenya suggests that remedial interventions will need to address both diet and parasitic infections to effectively combat this significant health threat.

The etiology of infections of the central nervous system (CNS) in Nepal often remains unrecognized because of underdeveloped laboratory facilities. The aim of this study was to investigate the etiology of CNS infections in a rural area of Nepal using molecular methods. From November 2014 to February 2016, cerebrospinal fluid (CSF) was collected from 176 consecutive patients presenting at United Mission Hospital in Tansen, Nepal, with symptoms of possible CNS infection. After the CSF samples were stored and transported frozen, polymerase chain reaction (PCR) was performed in Sweden, targeting a total of 26 pathogens using the FilmArray® ME panel (BioFire, bioMerieux, Salt Lake City, UT), the MeningoFinder® 2SMART (PathoFinder, Maastricht, The Netherlands), and an in-house PCR test for dengue virus (DENV), Japanese encephalitis virus (JEV), and Nipah virus (NiV). The etiology could be determined in 23%. The bacteria detected were Haemophilus influenzae (n = 5), Streptococcus pneumoniae (n = 4), and Neisseria meningitidis (n = 1). The most common virus was enterovirus detected in eight samples, all during the monsoon season. Other viruses detected were cytomegalovirus (n = 6), varicella zoster virus (n = 5), Epstein–Barr virus (n = 3), herpes simplex virus (HSV) type 1 (HSV-1) (n = 3), HSV-2 (n = 3), human herpes virus (HHV) type 6 (HHV-6) (n = 3), and HHV-7 (n = 2). Cryptococcus neoformans/gatti was found in four samples. None of the samples were positive for DENV, JEV, or NiV. Of the patients, 67% had been exposed to antibiotics before lumbar puncture. In conclusion, the etiology could not be found in 77% of the samples, indicating that the commercial PCR panels used are not suitable in this setting. Future studies on the etiology of CNS infections in Nepal could include metagenomic techniques.