We used end-point-dilution cultures to measure the level of infectious human immunodeficiency virus type 1 (HIV-1) in peripheral-blood mononuclear cells (PBMC) and plasma of 54 infected patients who were not receiving antiviral chemotherapy. HIV-1 was recovered from the plasma and PBMC of every seropositive patient, but from none of 22 seronegative control subjects. The mean titers in plasma were 30, 3500, and 3200 tissue-culture-infective doses (TCID) per milliliter for patients with asymptomatic infection, the acquired immunodeficiency syndrome (AIDS), and the AIDS-related complex, respectively. In PBMC, the mean titers were significantly higher for symptomatic patients (AIDS, 2200, and AIDS-related complex, 2700 TCID per 10(6) PBMC) than asymptomatic patients (20 TCID per 10(6) PBMC). The values for the symptomatic patients were considered to indicate that at least 1 in 400 circulating mononuclear cells harbored HIV-1. The HIV-1 titers of seven patients with AIDS or AIDS-related complex treated with zidovudine for four weeks decreased significantly in plasma but not in PBMC. In addition, the mean titer in the plasma of 20 patients receiving long-term zidovudine treatment (130 TCID per milliliter) was 25-fold lower than the mean for comparable untreated patients with AIDS or AIDS-related complex. We conclude that the levels of HIV-1 in plasma and PBMC are much higher than previous estimates. This high degree of HIV-1 viremia raises the possibility that the direct cytopathic effect of this retrovirus alone may be sufficient to explain much of the pathogenesis of AIDS.

Prospective observation of infants in an urban slum demonstrated that diarrheal diseases were associated with the development of malnutrition that was in turn linked to intestinal barrier disruption and that diarrhea was more severe in already malnourished children. The enteric protozoa were unexpectedly important causes of diarrhea in this setting. This study demonstrates the complex interrelationship of malnutrition and diarrhea in infants in low-income settings and points to the potential for infectious disease interventions in the prevention and treatment of malnutrition.

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.

Abstract. Oral vaccines appear less effective in children in the developing world. Proposed biologic reasons include concurrent enteric infections, malnutrition, breast milk interference, and environmental enteropathy (EE). Rigorous study design and careful data management are essential to begin to understand this complex problem while assuring research subject safety. Herein, we describe the methodology and lessons learned in the PROVIDE study (Dhaka, Bangladesh). A randomized clinical trial platform evaluated the efficacy of delayed-dose oral rotavirus vaccine as well as the benefit of an injectable polio vaccine replacing one dose of oral polio vaccine. This rigorous infrastructure supported the additional examination of hypotheses of vaccine underperformance. Primary and secondary efficacy and immunogenicity measures for rotavirus and polio vaccines were measured, as well as the impact of EE and additional exploratory variables. Methods for the enrollment and 2-year follow-up of a 700 child birth cohort are described, including core laboratory, safety, regulatory, and data management practices. Intense efforts to standardize clinical, laboratory, and data management procedures in a developing world setting provide clinical trials rigor to all outcomes. Although this study infrastructure requires extensive time and effort, it allows optimized safety and confidence in the validity of data gathered in complex, developing country settings. BACKGROUND AND RATIONALE

The observed in vitro and in vivo benefit of combination treatment with anti-human immunodeficiency virus (HIV) agents prompted us to examine the potential of resistance development when two protease inhibitors are used concurrently. Recombinant HIV-1 (NL4-3) proteases containing combined resistance mutations associated with BMS-186318 and A-77003 (or saquinavir) were either inactive or had impaired enzyme activity. Subsequent construction of HIV-1 (NL4-3) proviral clones containing the same mutations yielded viruses that were severely impaired in growth or nonviable, confirming that combination therapy may be advantageous. However, passage of BMS-186318-resistant HIV-1 (RF) in the presence of either saquinavir or SC52151, which represented sequential drug treatment, produced viable viruses resistant to both BMS-186318 and the second compound. The predominant breakthrough virus contained the G48V/A71T/V82A protease mutations. The clone-purified RF (G48V/A71T/V82A) virus, unlike the corresponding defective NL4-3 triple mutant, grew well and displayed cross-resistance to four distinct protease inhibitors. Chimeric virus and in vitro mutagenesis studies indicated that the RF-specific protease sequence, specifically the Ile at residue 10, enabled the NL4-3 strain with the triple mutant to grow. Our results clearly indicate that viral genetic background will play a key role in determining whether cross-resistance variants will arise.

Recent studies suggest small intestine bacterial overgrowth (SIBO) is common among developing world children. SIBO’s pathogenesis and effect in the developing world are unclear. Our objective was to determine the prevalence of SIBO in Bangladeshi children and its association with malnutrition. Secondary objectives included determination of SIBO’s association with sanitation, diarrheal disease, and environmental enteropathy. We performed a cross-sectional analysis of 90 Bangladeshi 2-year-olds monitored since birth from an impoverished neighborhood. SIBO was diagnosed via glucose hydrogen breath testing, with a cutoff of a 12-ppm increase over baseline used for SIBO positivity. Multivariable logistic regression was performed to investigate SIBO predictors. Differences in concomitant inflammation and permeability between SIBO-positive and -negative children were compared with multiple comparison adjustment. A total of 16.7% (15/90) of the children had SIBO. The strongest predictors of SIBO were decreased length-for-age Z score since birth (odds ratio [OR], 0.13; 95% confidence interval [CI], 0.03 to 0.60) and an open sewer outside the home (OR, 4.78; 95% CI, 1.06 to 21.62). Recent or frequent diarrheal disease did not predict SIBO. The markers of intestinal inflammation fecal Reg 1β (116.8 versus 65.6 µg/ml; P = 0.02) and fecal calprotectin (1,834.6 versus 766.7 µg/g; P = 0.004) were elevated in SIBO-positive children. Measures of intestinal permeability and systemic inflammation did not differ between the groups. These findings suggest linear growth faltering and poor sanitation are associated with SIBO independently of recent or frequent diarrheal disease. SIBO is associated with intestinal inflammation but not increased permeability or systemic inflammation.

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.

BackgroundLewis and secretor histo–blood group antigens (HBGAs) have been associated with decreased susceptibility to P[8] genotype rotavirus (RV) infections. Efficacy of vaccines containing attenuated P[8] strains is decreased in low-income countries. Host phenotype might impact vaccine efficacy (VE) by altering susceptibility to vaccination or RV diarrhea (RVD). We performed a substudy in a monovalent RV vaccine (RV1) efficacy trial in Bangladesh to determine the impact of Lewis and secretor status on risk of RVD and VE.MethodsIn infants randomized to receive RV1 or no RV1 at 10 and 17 weeks with 1 year of complete active diarrheal surveillance, we performed Lewis and secretor phenotyping and genotyped the infecting strain of each episode of RVD.ResultsA vaccine containing P[8] RV protected secretors and nonsecretors similarly. However, unvaccinated nonsecretors had a reduced risk of RVD (relative risk, 0.53 [95% confidence interval, .36–.79]) mediated by complete protection from P[4] but not P[8] RVs. This effect reduced VE in nonsecretors to 31.7%, compared to 56.2% among secretors, and decreased VE for the overall cohort.ConclusionsHost HBGA status may impact VE estimates by altering susceptibility to RV in unvaccinated children; future trials should therefore account for HBGA status.Clinical Trials RegistrationNCT01375647.