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We developed a real-time-PCR assay utilizing a molecular-beacon probe for the detection of Entamoeba histolytica and compared its sensitivity to stool antigen detection and traditional PCR. A total of 205 stool and liver abscess pus specimens from patients and controls were used for this purpose, 101 (49%) of which were positive by the TechLab E. histolytica-specific antigen detection test, while the other 104 (51%) stool and liver abscess pus specimens were negative by the antigen detection test. DNA was extracted from the stool and liver abscess pus specimens by the QIAGEN method and the small-subunit rRNA gene of E. histolytica and then amplified by traditional and real-time PCR. Out of these 205 stool and liver abscess pus specimens, 124 were positive by the real-time-PCR assay and 90 were positive by the traditional-PCR test. Compared to the real-time-PCR assay, the antigen detection test was 79% sensitive and 96% specific. When the traditional-PCR test results were compared to the real-time-PCR assay, the sensitivity of traditional PCR was 72% and the specificity was 99%. In conclusion, all three methods for the detection of E. histolytica were highly specific, with real-time PCR being the most sensitive.The World Health Organization has recommended that Entamoeba histolytica "should be specifically identified and if present should be treated" (27). Classic microscopic examination of the parasite E. histolytica in stool cannot differentiate it from the nonpathogenic but identically appearing parasites Entamoeba dispar and Entamoeba moshkovskii. While E. histolytica trophozoites are more likely than E. dispar and E. moshkovskii to contain ingested erythrocytes, most often E. histolytica trophozoites in patient stools lack ingested red cells (7, 10, 11). Not only is microscopy unable to differentiate E. histolytica from E. dispar or E. moshkovskii, it is at best only 10 to 60% sensitive and confounded with false-positive results due to misidentification of macrophages and nonpathogenic species of Entamoeba (12,13,18,23). Culture along with isoenzyme (zymodeme) analysis enables differentiation of E. histolytica from E. dispar or E. moshkovskii and was considered the gold standard for diagnosing amebic infection in the last 2 decades. However, amebic cultures and isoenzyme analysis require a week to complete and are negative with many microscopy-positive stool samples, in some cases due to delay in sample processing or due to the institution of antiamebic therapy prior to stool collection (13,14,23).New approaches to the detection of E. histolytica are based on the detection of an E. histolytica-specific antigen and DNA. Several groups have reported the detection of amebic antigen in stool samples, serum, liver abscess pus samples, and saliva using enzyme-linked immunosorbent assay methods (1,2,4,5,9,12,13,14,15,16,19,20,21,22,23). The TechLab (Blacksburg, Virginia) Entamoeba histolytica II kit is the only Food and Drug Administration-approved test that is specific and sensitive for the detection of E. histoly...

ObjectivesTo support the Bangladesh National Kala-azar Elimination Programme (NKEP), we investigated the feasibility of using trained village volunteers for detecting post-kala-azar dermal leishmaniasis (PKDL) cases, using polymerase chain reaction (PCR) for confirmation of diagnosis and treatment compliance by PKDL patients in Kanthal union of Trishal sub-district, Mymensingh, Bangladesh.MethodsIn this cross-sectional study, Field Research Assistants (FRAs) conducted census in the study area, and the research team trained village volunteers on how to look for PKDL suspects. The trained village volunteers (TVVs) visited each household in the study area for PKDL suspects and referred the suspected PKDL cases to the study clinic. The suspected cases underwent physical examinations by a qualified doctor and rK39 strip testing by the FRAs and, if positive, slit skin examination (SSE), culture, and PCR of skin specimens and peripheral buffy coat were done. Those with evidence of Leishmania donovani (LD) were referred for treatment. All the cases were followed for one year.ResultsThe total population of the study area was 29,226 from 6,566 households. The TVVs referred 52 PKDL suspects. Probable PKDL was diagnosed in 18 of the 52 PKDL suspect cases, and PKDL was confirmed in 9 of the 18 probable PKDL cases. The prevalence of probable PKDL was 6.2 per 10,000 people in the study area. Thirteen PKDL suspects self-reported from outside the study area, and probable and confirmed PKDL was diagnosed in 10 of the 13 suspects and in 5 of 10 probable PKDL cases respectively. All probable PKDL cases had hypopigmented macules. The median time for PKDL development was 36 months (IQR, 24–48). Evidence of the LD parasite was documented by SSE and PCR in 3.6% and 64.3% of the cases, respectively. PCR positivity was associated with gender and severity of disease. Those who were untreated had an increased risk (odds ratio = 3.33, 95%CI 1.29–8.59) of having persistent skin lesions compared to those who were treated. Patients' treatment-seeking behavior and treatment compliance were poor.ConclusionImproved detection of PKDL cases by TVVs is feasible and useful. The NKEP should promote PCR for the diagnosis of PKDL and should find ways for improving treatment compliance by patients.

This birth cohort study describes the early childhood burden of cryptosporidiosis in rural and urban Bangladesh. Important findings include an association of repeated and subclinical cryptosporidiosis with growth faltering, and disparate predominant Cryptosporidium species in the urban vs rural site.

Background. An estimated 1 million children die each year before their fifth birthday from diarrhea. Previous population-based surveys of pediatric diarrheal diseases have identified the protozoan parasite Entamoeba histolytica, the etiological agent of amebiasis, as one of the causes of moderate-to-severe diarrhea in sub-Saharan Africa and South Asia.Methods. We prospectively studied the natural history of E. histolytica colonization and diarrhea among infants in an urban slum of Dhaka, Bangladesh.Results. Approximately 80% of children were infected with E. histolytica by the age of 2 years. Fecal anti-galactose/N-acetylgalactosamine lectin immunoglobulin A was associated with protection from reinfection, while a high parasite burden and expansion of the Prevotella copri level was associated with diarrhea.Conclusions. E. histolytica infection was prevalent in this population, with most infections asymptomatic and diarrhea associated with both the amount of parasite and the composition of the microbiota.