International links between Streptococcus pneumoniae vaccine serotype 4 sequence type (ST) 801 in Northern European shipyard outbreaks of invasive pneumococcal disease.

BACKGROUND: Pneumococcal disease outbreaks of vaccine preventable serotype 4 sequence type (ST)801 in shipyards have been reported in several countries. We aimed to use genomics to establish any international links between them. METHODS: Sequence data from ST801-related outbreak isolates from Norway (n = 17), Finland (n = 11) and Northern Ireland (n = 2) were combined with invasive pneumococcal disease surveillance from the respective countries, and ST801-related genomes from an international collection (n = 41 of > 40,000), totalling 106 genomes. Raw data were mapped and recombination excluded before phylogenetic dating. RESULTS: Outbreak isolates were relatively diverse, with up to 100 SNPs (single nucleotide polymorphisms) and a common ancestor estimated around the year 2000. However, 19 Norwegian and Finnish isolates were nearly indistinguishable (0-2 SNPs) with the common ancestor dated around 2017. CONCLUSION: The total diversity of ST801 within the outbreaks could not be explained by recent transmission alone, suggesting that harsh environmental and associated living conditions reported in the shipyards may facilitate invasion of colonising pneumococci. However, near identical strains in the Norwegian and Finnish outbreaks does suggest that transmission between international shipyards also contributed to those outbreaks. This indicates the need for improved preventative measures in this working population including pneumococcal vaccination.

Using genomics to examine the persistence of Streptococcus pneumoniae serotype 19A in Ireland and the emergence of a sub-clade associated with vaccine failures.

BACKGROUND: Streptococcus pneumoniae serotype 19A remains a significant cause of invasive pneumococcal disease (IPD) in Ireland despite the successful introduction of a 13-valent pneumococcal conjugate vaccine (PCV13) in 2010 which reduced the overall incidence of IPD in children. METHODS: Invasive Streptococcus pneumoniae serotype 19A isolates from the Irish reference laboratory between 2007-08 and 2017-18 were analysed using whole genome sequencing (WGS) to investigate the persistence of this vaccine-preventable serotype. We compared the entire national 19A collection to other international collections using a standardised nomenclature of Global Pneumococcal Sequencing Clusters (GPSC). RESULTS: Expansion of GPSCs and clonal complexes (CCs) may have been associated with vaccine introduction and antimicrobial prescribing policies. A sub-clade of GPSC1-CC320 (n = 25) unique to Ireland, included five of the ten vaccine failures/breakthrough cases identified (p = 0.0086). This sub-clade was not observed in a global GPSC1-CC320 collection. All isolates within the sub-clade (n = 25) contained a galE gene variant rarely observed in a global pneumococcal collection (n = 37/13454, p < 0.001) nor within GPSC1-CC320 (n = 19/227) (p < 0.001). The sub-clade was estimated to have emerged at the start of the PCV-vaccine era (ancestral origin 2000, range 1995-2004) and expanded in Ireland, with most isolated after PCV13 introduction (n = 24/25). CONCLUSIONS: The identification of a sub-clade/variant of serotype 19A highlights the benefit of using WGS to analyse genotypes associated with persistence of a preventable serotype of S. pneumoniae. Particularly as this sub-clade identified was more likely to be associated with IPD in vaccinated children than other 19A genotypes. It is possible that changes to the galE gene, which is involved in capsule production but outside of the capsular polysaccharide biosynthesis locus, may affect bacterial persistence within the population. Discrete changes associated with vaccine-serotype persistence should be further investigated and may inform vaccine strategies.

Factors associated with typical enteropathogenic Escherichia coli infection among children <5 years old with moderate-to-severe diarrhoea in rural western Kenya, 2008-2012.

Typical enteropathogenic Escherichia coli (tEPEC) infection is a major cause of diarrhoea and contributor to mortality in children <5 years old in developing countries. Data were analysed from the Global Enteric Multicenter Study examining children <5 years old seeking care for moderate-to-severe diarrhoea (MSD) in Kenya. Stool specimens were tested for enteric pathogens, including by multiplex polymerase chain reaction for gene targets of tEPEC. Demographic, clinical and anthropometric data were collected at enrolment and ~60-days later; multivariable logistic regressions were constructed. Of 1778 MSD cases enrolled from 2008 to 2012, 135 (7.6%) children tested positive for tEPEC. In a case-to-case comparison among MSD cases, tEPEC was independently associated with presentation at enrolment with a loss of skin turgor (adjusted odds ratio (aOR) 2.08, 95% confidence interval (CI) 1.37-3.17), and convulsions (aOR 2.83, 95% CI 1.12-7.14). At follow-up, infants with tEPEC compared to those without were associated with being underweight (OR 2.2, 95% CI 1.3-3.6) and wasted (OR 2.5, 95% CI 1.3-4.6). Among MSD cases, tEPEC was associated with mortality (aOR 2.85, 95% CI 1.47-5.55). This study suggests that tEPEC contributes to morbidity and mortality in children. Interventions aimed at defining and reducing the burden of tEPEC and its sequelae should be urgently investigated, prioritised and implemented.

Meeting Report: WHO Workshop on modelling global mortality and aetiology estimates of enteric pathogens in children under five. Cape Town, 28-29th November 2018.

Investment in vaccine product development should be guided by up-to-date and transparent global burden of disease estimates, which are also fundamental to policy recommendation and vaccine introduction decisions. For low- and middle-income countries (LMICs), vaccine prioritization is primarily driven by the number of deaths caused by different pathogens. Enteric diseases are known to be a major cause of death in LMICs. The two main modelling groups providing mortality estimates for enteric diseases are the Institute for Health Metrics and Evaluation (IHME) at the University of Washington, Seattle and the Maternal Child Epidemiology Estimation (MCEE) group, led by Johns Hopkins Bloomberg School of Public Health. Whilst previous global diarrhoea mortality estimates for under five-year-olds from these two groups were closely aligned, more recent estimates for 2016 have diverged, particularly with respect to numbers of deaths attributable to different enteric pathogens. This has impacted prioritization and investment decisions for vaccines in the development pipeline. The mission of the Product Development for Vaccines Advisory Committee (PDVAC) at the World Health Organisation (WHO) is to accelerate product development of vaccines and technologies that are urgently needed and ensure they are appropriately targeted for use in LMICs. At their 2018 meeting, PDVAC recommended the formation of an independent working group of subject matter experts to explore the reasons for the difference between the IHME and MCEE estimates, and to assess the respective strengths and limitations of the estimation approaches adopted, including a review of the data on which the estimates are based. Here, we report on the proceedings and recommendations from a consultation with the working group of experts, the IHME and MCEE modelling groups, and other key stakeholders. We briefly review the methodological approaches of both groups and provide a series of proposals for investigating the drivers for the differences in enteric disease burden estimates.

Diarrhoea, enteric pathogen detection and nutritional indicators among controls in the Global Enteric Multicenter Study, Kenya site: an opportunity to understand reference populations in case-control studies of diarrhoea.

Given the challenges in accurately identifying unexposed controls in case-control studies of diarrhoea, we examined diarrhoea incidence, subclinical enteric infections and growth stunting within a reference population in the Global Enteric Multicenter Study, Kenya site. Within 'control' children (0-59 months old without diarrhoea in the 7 days before enrolment, n = 2384), we examined surveys at enrolment and 60-day follow-up, stool at enrolment and a 14-day post-enrolment memory aid for diarrhoea incidence. At enrolment, 19% of controls had ⩾1 enteric pathogen associated with moderate-to-severe diarrhoea ('MSD pathogens') in stool; following enrolment, many reported diarrhoea (27% in 7 days, 39% in 14 days). Controls with and without reported diarrhoea had similar carriage of MSD pathogens at enrolment; however, controls reporting diarrhoea were more likely to report visiting a health facility for diarrhoea (27% vs. 7%) or fever (23% vs. 16%) at follow-up than controls without diarrhoea. Odds of stunting differed by both MSD and 'any' (including non-MSD pathogens) enteric pathogen carriage, but not diarrhoea, suggesting control classification may warrant modification when assessing long-term outcomes. High diarrhoea incidence following enrolment and prevalent carriage of enteric pathogens have implications for sequelae associated with subclinical enteric infections and for design and interpretation of case-control studies examining diarrhoea.

Associations between presence of handwashing stations and soap in the home and diarrhoea and respiratory illness, in children less than five years old in rural western Kenya.

OBJECTIVE: We tested whether soap presence in the home or a designated handwashing station was associated with diarrhoea and respiratory illness in Kenya. METHODS: In April 2009, we observed presence of a handwashing station and soap in households participating in a longitudinal health surveillance system in rural Kenya. Diarrhoea and acute respiratory illness (ARI) in children < 5 years old were identified using parent-reported syndromic surveillance collected January-April 2009. We used multivariate generalised linear regression to estimate differences in prevalence of illness between households with and without the presence of soap in the home and a handwashing station. RESULTS: Among 2547 children, prevalence of diarrhoea and ARI was 2.3 and 11.4 days per 100 child-days, respectively. Soap was observed in 97% of households. Children in households with soap had 1.3 fewer days of diarrhoea/100 child-days (95% CI -2.6, -0.1) than children in households without soap. ARI prevalence was not associated with presence of soap. A handwashing station was identified in 1.4% of households and was not associated with a difference in diarrhoea or ARI prevalence. CONCLUSIONS: Soap presence in the home was significantly associated with reduced diarrhoea, but not ARI, in children in rural western Kenya. Whereas most households had soap in the home, almost none had a designated handwashing station, which may prevent handwashing at key times of hand contamination.

What are the most sensitive and specific sign and symptom combinations for influenza in patients hospitalized with acute respiratory illness? Results from western Kenya, January 2007-July 2010.

Influenza causes severe illness and deaths, and global surveillance systems use different clinical case definitions to identify patients for diagnostic testing. We used data collected during January 2007-July 2010 at hospital-based influenza surveillance sites in western Kenya to calculate sensitivity, specificity, positive predictive value, and negative predictive value for eight clinical sign/symptom combinations in hospitalized patients with acute respiratory illnesses, including severe acute respiratory illness (SARI) (persons aged 2-59 months: cough or difficulty breathing with an elevated respiratory rate or a danger sign; persons aged ≥5 years: temperature ≥38 °C, difficulty breathing, and cough or sore throat) and influenza-like illness (ILI) (all ages: temperature ≥38 °C and cough or sore throat). Overall, 4800 persons aged ≥2 months were tested for influenza; 416 (9%) had laboratory-confirmed influenza infections. The symptom combination of cough with fever (subjective or measured ≥38 °C) had high sensitivity [87·0%, 95% confidence interval (CI) 83·3-88·9], and ILI had high specificity (70·0%, 95% CI 68·6-71·3). The case definition combining cough and any fever is a simple, sensitive case definition for influenza in hospitalized persons of all age groups, whereas the ILI case definition is the most specific. The SARI case definition did not maximize sensitivity or specificity.

Community perceptions of bloody diarrhoea in an urban slum in South Asia: implications for introduction of a Shigella vaccine.

Understanding local perceptions of disease causation could help public health officials improve strategies to prevent bloody diarrhoea. A cross-sectional survey was conducted in Dhaka, Bangladesh to elicit community beliefs about the causes of and prevention strategies for bloody diarrhoea. Between March and June 2003, we interviewed 541 randomly selected respondents. Overall, 507 (93%) respondents perceived that a vaccine could prevent bloody diarrhoea. If a vaccine provided lifetime protection, 445 (83%) respondents stated that they would opt to get the vaccine and would pay a median of $0·05 (range U.S.$0·01-0·15) for it, equivalent to <1% of their median weekly income. There was almost universal perception that an effective vaccine to prevent bloody diarrhoea was highly beneficial and acceptable. While respondents valued a vaccine for prevention of bloody diarrhoea, they were only willing to pay minimally for it. Therefore, achieving a high rate of Shigella vaccine coverage may require subsidy of vaccine purchase.

Rift Valley fever in Kenya: history of epizootics and identification of vulnerable districts.

Since Kenya first reported Rift Valley fever (RVF)-like disease in livestock in 1912, the country has reported the most frequent epizootics of RVF disease. To determine the pattern of disease spread across the country after its introduction in 1912, and to identify regions vulnerable to the periodic epizootics, annual livestock disease records at the Department of Veterinary Services from 1910 to 2007 were analysed in order to document the number and location of RVF-infected livestock herds. A total of 38/69 (55%) administrative districts in the country had reported RVF epizootics by the end of 2007. During the 1912-1950 period, the disease was confined to a district in Rift Valley province that is prone to flooding and where livestock were raised in proximity with wildlife. Between 1951 and 2007, 11 national RVF epizootics were recorded with an average inter-epizootic period of 3·6 years (range 1-7 years); in addition, all epizootics occurred in years when the average annual rainfall increased by more than 50% in the affected districts. Whereas the first two national epizootics in 1951 and 1955 were confined to eight districts in the Rift Valley province, there was a sustained epizootic between 1961 and 1964 that spread the virus to over 30% of the districts across six out of eight provinces. The Western and Nyanza provinces, located on the southwestern region of the country, had never reported RVF infections by 2007. The probability of a district being involved in a national epizootic was fivefold higher (62%) in districts that had previously reported disease compared to districts that had no prior disease activity (11%). These findings suggests that once introduced into certain permissive ecologies, the RVF virus becomes enzootic, making the region vulnerable to periodic epizootics that were probably precipitated by amplification of resident virus associated with heavy rainfall and flooding.

Successful co-administration of a human rotavirus and oral poliovirus vaccines in Bangladeshi infants in a 2-dose schedule at 12 and 16 weeks of age.

Co-administration of oral live-attenuated human rotavirus vaccine RIX4414 (Rotarix) and oral polio vaccine (OPV) was assessed. Healthy infants were randomised to receive 2-doses of either: RIX4414 or placebo co-administered with OPV (12 and 16 weeks of age); or RIX4414 or placebo given 15 days after OPV. After vaccination, 56.5-66.7% of RIX4414 and 18.6% of placebo recipients had seroconverted for rotavirus IgA. No significant differences between RIX4414 groups with or without OPV co-administration were observed. No statistically significant differences were observed between groups for polio seroprotection rates. RIX4414 vaccine was immunogenic when co-administered with OPV and did not interfere with OPV seroprotection rates.

Tracking epidemic Chikungunya virus into the Indian Ocean from East Africa.

The largest documented outbreak of Chikungunya virus (CHIKV) disease occurred in the Indian Ocean islands and India during 2004-2007. The magnitude of this outbreak led to speculation that a new variant of the virus had emerged that was either more virulent or more easily transmitted by mosquito vectors. To study this assertion, it is important to know the origin of the virus and how the particular strain circulating during the outbreak is related to other known strains. This study genetically characterized isolates of CHIKV obtained from Mombasa and Lamu Island, Kenya, during 2004, as well as strains from the 2005 outbreak recorded in Comoros. The results of these analyses demonstrated that the virus responsible for the epidemic that spread through the Indian Ocean originated in coastal Kenya during 2004 and that the closest known ancestors are members of the Central/East African clade. Genetic elements that may be responsible for the scope of the outbreak were also identified.

Risk factors for Nipah virus encephalitis in Bangladesh.

Nipah virus (NiV) is a paramyxovirus that causes severe encephalitis in humans. During January 2004, twelve patients with NiV encephalitis (NiVE) were identified in west-central Bangladesh. A case-control study was conducted to identify factors associated with NiV infection. NiVE patients from the outbreak were enrolled in a matched case-control study. Exact odds ratios (ORs) and 95% confidence intervals (CIs) were calculated by using a matched analysis. Climbing trees (83% of cases vs. 51% of controls, OR 8.2, 95% CI 1.25-infinity) and contact with another NiVE patient (67% of cases vs. 9% of controls, OR 21.4, 95% CI 2.78-966.1) were associated with infection. We did not identify an increased risk for NiV infection among persons who had contact with a potential intermediate host. Although we cannot rule out person-to-person transmission, case-patients were likely infected from contact with fruit bats or their secretions.

Tuberculosis risk among staff of a large public hospital in Kenya.

SETTING: In sub-Saharan Africa, high rates of tuberculosis (TB) and human immunodeficiency virus (HIV) infection pose a serious threat for occupationally acquired TB among health care workers. OBJECTIVE: To identify factors associated with TB disease among staff of an 1800-bed hospital in Kenya. DESIGN: We calculated TB incidence among staff and conducted a case-control study where cases (n = 65) were staff diagnosed with TB and controls (n = 316) were randomly selected staff without recent TB. RESULTS: The annual incidence of TB from 2001 to 2005 ranged from 645 to 1115 per 100000 population. Factors associated with TB disease were additional daily hours spent in rooms with patients (adjusted odds ratio [aOR] 1.3, 95%CI 1.2-1.5), working in areas where TB patients received care (aOR 2.1, 95%CI 1.1-4.2), HIV infection (aOR 29.1, 95%CI 5.1-167) and living in a slum (aOR 4.7, 95%CI 1.8-12.5) or hospital-provided low-income housing (aOR 2.6, 95%CI 1.2-5.6). CONCLUSION: Hospital exposures were associated with TB disease among staff at this hospital regardless of their job designation, even after controlling for living conditions, suggesting transmission from patients. Health care facilities should improve infection control practices, provide quality occupational health services and encourage staff testing for HIV infection to address the TB burden in hospital staff.

Aflatoxin levels in locally grown maize from Makueni District, Kenya.

OBJECTIVES: Investigations were carried out to determine aflatoxin levels in household maize in Makueni District and to correlate aflatoxin levels to maize drying and storage practices. Also, aflatoxin exposure in villages that reported aflatoxicosis cases in 2005 was compared with that in villages that did not report cases to assess whether aflatoxin exposure levels could be used to identify high-risk villages for targeted prevention interventions. DESIGN: A cross-sectional study. SETTING: Three divisions of Makueni district, Kibwezi, Makindu and Mtito Andei in Eastern Province, Kenya. SUBJECTS: Ninety six households were surveyed, and 104 maize samples were analysed for total aflatoxin levels from June to July 2005. The households were selected from high and low aflatoxicosis risk areas. RESULTS: Out of the 104 maize samples collected from 96 households, 37 (35.5%) had aflatoxin levels above the World Health Organisation (WHO) recommended maximum limit of 20 ppb. All of these samples were homegrown or purchased. Twenty one samples (20.1%) had levels above 100 ppb. Eleven (10.6%) had extremely high levels above 1000 ppb. No relief supply maize had aflatoxin levels above the WHO maximum limit. CONCLUSION: High levels of aflatoxin in homegrown and purchased maize suggested that aflatoxin exposure was widespread.

Prevalence of antibodies against Rift Valley fever virus in Kenyan wildlife.

Rift Valley fever virus (RVFV) is an arbovirus associated with periodic outbreaks, mostly on the African continent, of febrile disease accompanied by abortion in livestock, and a severe, fatal haemorrhagic syndrome in humans. However, the maintenance of the virus during the inter-epidemic period (IEP) when there is low or no disease activity detected in livestock or humans has not been determined. This study report prevalence of RVFV-neutralizing antibodies in sera (n=896) collected from 16 Kenyan wildlife species including at least 35% that were born during the 1999-2006 IEP. Specimens from seven species had detectable neutralizing antibodies against RVFV, including African buffalo, black rhino, lesser kudu, impala, African elephant, kongoni, and waterbuck. High RVFV antibody prevalence (>15%) was observed in black rhinos and ruminants (kudu, impala, buffalo, and waterbuck) with the highest titres (up to 1:1280) observed mostly in buffalo, including animals born during the IEP. All lions, giraffes, plains zebras, and warthogs tested were either negative or less than two animals in each species had low (or= 1:80. These data provide evidence that wild ruminants are infected by RVFV but further studies are required to determine whether these animals play a role in the virus maintenance between outbreaks and virus amplification prior to a noticeable outbreak.

Simplifying the exploration of volumetric images: development of a 3D user interface for the radiologist's workplace.

Volumetric imaging (computed tomography and magnetic resonance imaging) provides increased diagnostic detail but is associated with the problem of navigation through large amounts of data. In an attempt to overcome this problem, a novel 3D navigation tool has been designed and developed that is based on an alternative input device. A 3D mouse allows for simultaneous definition of position and orientation of orthogonal or oblique multiplanar reformatted images or slabs, which are presented within a virtual 3D scene together with the volume-rendered data set and additionally as 2D images. Slabs are visualized with maximum intensity projection, average intensity projection, or standard volume rendering technique. A prototype has been implemented based on PC technology that has been tested by several radiologists. It has shown to be easily understandable and usable after a very short learning phase. Our solution may help to fully exploit the diagnostic potential of volumetric imaging by allowing for a more efficient reading process compared to currently deployed solutions based on conventional mouse and keyboard.

Risk factors for typhoid fever in a slum in Dhaka, Bangladesh.

We systematically investigated risk factors for typhoid fever in Kamalapur, a poor urban area of Bangladesh, to inform targeted public health measures for its control. We interviewed patients with typhoid fever and two age-matched controls per case about exposures during the 14 days before the onset of illness. The municipal water supply was used by all 41 cases and 81of 82 controls. In multivariate analysis, drinking unboiled water at home was a significant risk factor [adjusted odds ratio (aOR) 12.1, 95% CI 2.2-65.6]. Twenty-three (56%) cases and 21 (26%) controls reported that water from the primary source was foul-smelling (aOR 7.4, 95% CI 2.1-25.4). Eating papaya was associated with illness (aOR 5.2, 95% CI 1.2-22.2). Using a latrine for defecation was significantly protective (aOR 0.1, 95% CI 0.02-0.9). Improved chlorination of the municipal water supply or disinfecting drinking water at the household level may dramatically reduce the risk of typhoid fever in Kamalapur. The protective effect of using latrines, particularly among young children, should be investigated further.

Prevalence of sputum smear-positive tuberculosis in a rural area in Bangladesh.

The objective of the study was to determine the prevalence of smear-positive tuberculosis (TB) in a rural area in Bangladesh at Matlab. A TB surveillance system was established among 106,000 people in rural Bangladesh at Matlab. Trained field workers interviewed all persons aged > or = 15 years to detect suspected cases of TB (cough > 21 days) and sputum specimens of suspected cases were examined for acid-fast bacilli (AFB). Of 59,395 persons interviewed, 4235 (7.1%) had a cough for > 21 days. Sputum specimens were examined for AFB from 3834 persons, 52 (1.4%) of them were positive for AFB. The prevalence of chronic cough and sputum positivity were significantly higher among males compared to females (P < 0.001). The population-based prevalence rate of smear-positive TB cases was 95/100,000 among persons aged > or = 15 years. Cases of TB clustered geographically (relative risk 5.53, 95% CI 3.19-9.59). The high burden of TB among rural population warrants appropriate measures to control TB in Bangladesh. The higher prevalence of persistent cough and AFB-positive sputum among males need further exploration. Factors responsible for higher prevalence of TB in clusters should be investigated.

Emergence of multidrug-resistant Salmonella enterica serotype Typhi with decreased ciprofloxacin susceptibility in Bangladesh.

During 1989-2002, we studied the antimicrobial resistance of 3928 blood culture isolates of Salmonella enterica serotype Typhi (S. Typhi) in Dhaka, Bangladesh. Overall 32% (1270) of the strains were multidrug-resistant (MDR, resistant to chloramphenicol, ampicillin and trimethoprim-sulphamethoxazole); first detected in 1990 (rate of 8%), increased in 1994 (44%), declined in 1996 (22%, P<0.01 compared to 1994) and re-emerged in 2001 (36%) and 2002 (42%, P<0.01 compared to 1996). An increased MIC of ciprofloxacin (0.25 microg/ml) indicating decreased susceptibility to ciprofloxacin was detected in 24 (18.2%) out of 132 randomly selected strains during 1990-2002; more frequently in MDR than susceptible strains (46.3% vs. 5.5%, P<0.001), and the proportion of them rose to 47% in 2002 from 8% in 2000 (P<0.01). Ciprofloxacin (5 microg) disk diffusion zone diameters of < or =24 mm as break-point had 98% sensitivity and 100% specificity when compared with a ciprofloxacin MIC of 0.25 microg/ml as break-point for decreased susceptibility; being a useful and easy screen test. All strains were susceptible to ceftriaxone. The emergence of MDR S. Typhi with decreased ciprofloxacin susceptibility will further complicate the therapy of typhoid fever because of the lack of optimum treatment guidelines.

Application of an improved method for the recombinant k 39 enzyme-linked immunosorbent assay to detect visceral leishmaniasis disease and infection in Bangladesh.

Several serology-based immunoassays are used to diagnose visceral leishmaniasis (VL), a chronic protozoan parasitic disease caused by the Leishmania donovani complex. These tests are primarily designed to diagnose the most severe clinical form of VL, known as kala-azar. However, leishmanial infection is frequently asymptomatic and may manifest only as a positive serologic response or positive leishmanin skin test. We modified a previously described enzyme-linked immunosorbent assay (ELISA) that detects patient antibodies reactive with the recombinant Leishmania protein K39 (rK39) to confirm suspected kala-azar and to detect asymptomatic infection in a community study in Bangladesh. With the inclusion of a standard curve on each ELISA plate, the rK39 ELISA was more repeatable (kappa coefficient of agreement=0.970) and more reliable compared to the original method (kappa=0.587, P<0.001). The cutoff point for a positive antibody response was chosen based on the 99th percentile of the ELISA distribution for the negative-control sera. However, we found that sera from all patients with active kala-azar yielded values more than twice the magnitude of this cutoff. Using receiver-operator characteristic curves, we determined a second cutoff value predictive of kala-azar. Using these criteria, the sensitivity and specificity of the modified ELISA for kala-azar were 97.0% and 98.9%, respectively, for sera from our study population. We hypothesize that individuals with antibody levels greater than the 99th percentile of the negative controls but less than the cutoff point for kala-azar have asymptomatic leishmanial infections.