Contribution of Schistosoma mansoni to systemic inflammation and microbial translocation among people with HIV in Zambia.

BACKGROUND: Schistosoma mansoni is hyperendemic in many rural areas of Zambia where up to 77% of people are positive for infection via serologic evaluation. Zambia also has a high prevalence of HIV infection. Individually, S. mansoni and HIV infection impair gastrointestinal barrier integrity and induce inflammation, but the effects of coinfection are not well understood. We set out to test the hypothesis that HIV would exacerbate intestinal barrier failure in patients with S. mansoni infection. METHODS: Adults attending medical outpatient clinics in Kaoma, Western Province, Zambia, were enrolled in a case-control study to determine the relative contributions of schistosomiasis and HIV to microbial translocation (measured as soluble CD14 [sCD14] and lipopolysaccharide binding protein [LBP]) and inflammation (measured as CRP). RESULTS: Among 152 adults evaluated, 74 (49%) were HIV-seropositive, 45 (29%) were shedding schistosome ova (Kato-Katz), 120 (81%) were seropositive for schistosome antibodies (i.e. prior or current infection, with or without egg shedding) and 16 (11%) were HIV/schistosome coinfected (defined by Kato-Katz). HIV infection was associated with higher circulating sCD14 concentrations (p=0.003 by Kruskal-Wallis test), but schistosomiasis was not. HIV infection was associated with greater exposure to schistosomes assessed serologically (OR=2.48, 95% CI 1.05 to 5.86; p=0.03), but reduced likelihood of egg shedding (OR 0.47, 95% CI 0.21 to 1.01; p=0.03). CONCLUSIONS: There was no evidence for a compounding or synergistic effect of coinfection on microbial translocation that appeared to be correlated with HIV infection. Further studies are needed to understand how the increase in LBP secondary to HIV infection may decrease schistosome egg excretion in coinfected individuals.

Molecular characterization of Cryptosporidium spp. from patients with diarrhoea in Lusaka, Zambia.

Cryptosporidium is a major etiological agent of diarrhoeal diseases among children and immune-compromised individuals in sub-Saharan African countries. We conducted a study to determine the prevalence and genetic characteristics of Cryptosporidium spp. in stool samples from patients with diarrhoea who presented at the University Teaching Hospital in Lusaka, Zambia. Cryptosporidium species and subtypes from 71 microscopically confirmed cryptosporidiosis stool samples collected between 2017 and 2019 were determined by polymerase chain reaction followed by partial sequencing of the small subunit rRNA and 60-kDa glycoprotein (gp60) gene. Additionally, data for the period between 2014 and 2019 were reviewed and analysed for cryptosporidiosis seasonal and age distribution. Cryptosporidium was more prevalent in the rainy season. The highest number of cases was reported among the 1-4 year age group. By sequence analysis of the 71 positive isolates, Cryptosporidium hominis (n = 42; 59.2%), C. parvum (n = 27; 38%), C. felis (n = 1; 1.4%), and C. meleagridis (n = 1; 1.4%) were identified. Four C. hominis subtype families (Ia, Ib, Id, and Ie) and three C. parvum subtype families (IIc, IIe, and IIs) were identified. The most frequent subtypes were IeA11G3T3 (n = 20; 28.2%), IIcA5G3 (n = 12; 16.9%), IIeA12G1 (n = 11; 15.5%) and IaA30R3 (n = 10; 14.1%). The observed species/subtypes of C. hominis and C. parvum indicated that the infection was mainly transmitted through the anthroponotic route. The identification of C. felis and C. meleagridis suggests that an atypical zoonotic transmission cycle also exists.

TITLE: Caractérisation moléculaire de Cryptosporidium spp. de patients souffrant de diarrhée à Lusaka, Zambie. ABSTRACT: Cryptosporidium est un agent étiologique majeur des maladies diarrhéiques chez les enfants et les personnes immunodéprimées dans les pays d'Afrique subsaharienne. Nous avons mené une étude pour déterminer la prévalence et les caractéristiques génétiques de Cryptosporidium spp. dans des échantillons de selles de patients souffrant de diarrhée qui se sont présentés à l'hôpital universitaire de Lusaka, en Zambie. Les espèces et sous-types de Cryptosporidium provenant de 71 échantillons de selles de cryptosporidiose, confirmés au microscope et prélevés entre 2017 et 2019, ont été déterminés par réaction en chaîne par polymérase suivie d'un séquençage partiel de la petite sous-unité de l'ARNr et du gène de la glycoprotéine de 60 kDa (gp60). De plus, les données pour la période entre 2014 et 2019 ont été examinées et analysées pour la distribution saisonnière et par âge de la cryptosporidiose. Cryptosporidium était plus répandu pendant la saison des pluies. Le plus grand nombre de cas a été signalé dans le groupe d'âge de 1 à 4 ans. Par analyse séquentielle des 71 isolats positifs, Cryptosporidium hominis (n = 42 ; 59,2 %), C. parvum (n = 27 ; 38 %), C. felis (n = 1 ; 1,4 %) et C. meleagridis (n = 1 ; 1,4 %) ont été identifiés. Quatre familles de sous-types de C. hominis (Ia, Ib, Id et Ie) et trois familles de sous-types de C. parvum (IIc, IIe et IIs) ont été identifiées. Les sous-types les plus fréquents étaient A11G3T3 (n = 20 ; 28,2 %), IIcA5G3 (n = 12 ; 16,9 %), IIeA12G1 (n = 11 ; 15,5 %) et IaA30R3 (n = 10 ; 14,1 %). Les espèces/sous-types observés de C. hominis et C. parvum indiquent que l'infection est principalement transmise par voie anthroponotique. L'identification de C. felis et C. meleagridis suggère qu'il existe également un cycle de transmission zoonotique atypique.

Retinoic acid elicits a coordinated expression of gut homing markers on T lymphocytes of Zambian men receiving oral Vivotif, but not Rotarix, Dukoral or OPVERO vaccines.

All-trans retinoic acid (ATRA) up-regulates, in laboratory animals, the expression of the gut homing markers α4ß7 integrin and CCR9 on lymphocytes, increasing their gut tropism. Here, we show that, in healthy adult volunteers, ATRA induced an increase of these gut homing markers on T cells in vivo in a time dependent manner. The coordinated increase of α4ß7 and CCR9 by ATRA was seen in 57% (12/21) of volunteers and only when given together with an oral Vivotif vaccine. When this coordinated response to ATRA and Vivotif vaccine was present, it was strongly correlated with the gut immunoglobulin A (IgA) specific response to vaccine LPS (ρ = 0.82; P = 0.02). Using RNA-Seq analysis of whole blood transcription, patients receiving ATRA and Vivotif in conjunction showed transcriptomic changes in immune-related pathways, particularly including interferon α/ß signaling pathway, membrane-ECM interactions and immune hubs. These results suggest that exogenous ATRA can be used to manipulate responses to a subclass of oral vaccines, so far limited to a live attenuated Vivotif vaccine.

High Schistosoma mansoni disease burden in a rural district of western Zambia.

Schistosoma mansoni disease is endemic in most parts of rural Zambia, and associated complications are common. We conducted a cross-sectional study among 754 people in rural communities of Kaoma District, western Zambia to determine the burden of S. mansoni infection and associated morbidity. Parasitology and ultrasonography assessments were conducted on consenting participants. The overall prevalence of S. mansoni infection and geometric mean egg count (GMEC) were 42.4% (304) and 86.6 eggs per gram (95% confidence interval = 75.6-99.6), respectively. Prevalence was highest in the age group of 15-19 years old (adjusted prevalence ratio = 1.70, P = 0.017). S. mansoni-related portal fibrosis was detected in 26% of the participants screened. Participants above 39 years old were 2.93 times more likely to have fibrosis than the 7-9 years old age group (P = 0.004). The study highlights the high burden of S. mansoni disease in this area and calls for immediate interventions to avert complications associated with the disease.

Differential changes in expression of intestinal antimicrobial peptide genes during Ascaris lumbricoides infection in Zambian adults do not respond to helminth eradication.

BACKGROUND: Intestinal helminthiasis modulates immune responses to vaccines and environmental allergens. To explore the impact on intestinal host defense, we assessed expression of antimicrobial peptide genes, together with T cell subset markers and cytokines, in patients with ascariasis before and after treatment. METHODS: Case patients (n = 27) and control subjects (n = 44) underwent enteroscopy for collection of jejunal biopsy specimens, which were used in quantitative, real-time reverse-transcription polymerase chain reaction for a range of host defense genes; blood samples were also analyzed simultaneously. RESULTS: The level of gene expression (mRNA) of HD5, hBD1, and LL-37 was lower in case patients than in control subjects, and the level of expression of HD6 was increased. However, after successful eradication, there was no trend to values seen in control subjects. Helminthiasis was associated with increased intestinal expression of the Th1 genes T-bet and interferon-γ. In peripheral blood mononuclear cells (PBMCs), a mixed profile of T cell markers and cytokines was increased. Ascaris-induced down-regulation of HD5 was observed in individuals with higher RORγt expression in PBMCs, but we found no evidence that this was mediated by circulating interleukin-22. CONCLUSIONS: Human ascariasis was associated with changes in antimicrobial peptide gene expression and immunological markers. Such changes may have implications for susceptibility to infectious disease and responsiveness to oral vaccines in tropical populations.

High dose prolonged treatment with nitazoxanide is not effective for cryptosporidiosis in HIV positive Zambian children: a randomised controlled trial.

BACKGROUND: Treatment of cryptosporidiosis in HIV infected children has proved difficult and unsatisfactory with no drugs having demonstrable efficacy in controlled trials except nitazoxanide. We hypothesised that a prolonged course of treatment with high dose nitazoxanide would be effective in treating cryptosporidiosis in HIV positive Zambian children. METHODS: We performed a double-blind, randomised, placebo controlled trial in paediatric patients in the UTH in Lusaka. The study included HIV positive children between one and eleven years of age if 2 out of 3 stool samples were positive for oocysts of Cryptosporidium spp. Children were given nitazoxanide suspension in a dose of 200 mg twice daily (bid) for 28 days (if 1-3 years old) or 400 mg bid for 28 days (if 4-11 years old), or matching placebo. RESULTS: Sixty children were randomised and 52 were fully evaluated. Only five children were 4 years of age or over and received the higher dose. In the primary efficacy analysis, 11 out of 26 (42%) in the active treatment group achieved a 'Well' clinical response compared to 8 out of 26 (35%) in the placebo group. Parasitological response was declared as 'Eradicated' in 27% in the active group and 35% in the placebo group. Mortality (16/52, 31%) did not differ by treatment allocation. CONCLUSION: We found no significant benefit in children with cryptosporidiosis despite high dose and longer treatment duration. This is the second randomised controlled trial to suggest that in Zambian children with HIV-related immunosuppression nitazoxanide does not eradicate this infection nor provide clinical symptom reduction. TRIAL REGISTRATION: The trial was registered as ISRCTN41089957.

Susceptibility to intestinal infection and diarrhoea in Zambian adults in relation to HIV status and CD4 count.

BACKGROUND: The HIV epidemic in sub-Saharan Africa has had a major impact on infectious disease, and there is currently great interest in the impact of HIV on intestinal barrier function. A three year longitudinal cohort study in a shanty compound in Lusaka, Zambia, carried out before anti-retroviral therapy was widely available, was used to assess the impact of HIV on susceptibility to intestinal infectious disease. We measured the incidence and seasonality of intestinal infection and diarrhoea, aggregation of disease in susceptible individuals, clustering by co-habitation and genetic relatedness, and the disease-to-infection ratio. METHODS: Adults living in a small section of Misisi, Lusaka, were interviewed every two weeks to ascertain the incidence of diarrhoea. Monthly stool samples were analysed for selected pathogens. HIV status and CD4 count were determined annually. RESULTS: HIV seroprevalence was 31% and the prevalence of immunosuppression (CD4 count 200 cells/microL or less) was 10%. Diarrhoea incidence was 1.1 episodes per year and the Incidence Rate Ratio for HIV infection was 2.4 (95%CI 1.7-3.3; p < 0.001). The disease-to-infection ratio was increased at all stages of HIV infection. Aggregation of diarrhoea in susceptible individuals was observed irrespective of immunosuppression, but there was little evidence of clustering by co-habitation or genetic relatedness. There was no evidence of aggregation of asymptomatic infections. CONCLUSION: HIV has an impact on intestinal infection at all stages, with an increased disease-to-infection ratio. The aggregation of disease in susceptible individuals irrespective of CD4 count suggests that this phenomenon is not a function of cell mediated immunity.

Cytokine activation is predictive of mortality in Zambian patients with AIDS-related diarrhoea.

BACKGROUND: Mortality in Zambian AIDS patients is high, especially in patients with diarrhoea, and there is still unacceptably high mortality in Zambian patients just starting anti-retroviral therapy. We set out to determine if high concentrations of serum cytokines correlate with mortality. METHODS: Serum samples from 30 healthy controls (HIV seropositive and seronegative) and 50 patients with diarrhoea (20 of whom died within 6 weeks) were analysed. Concentrations of tumour necrosis factor receptor p55 (TNFR p55), macrophage migration inhibitory factor (MIF), interleukin (IL)-6, IL-12, interferon (IFN)-gamma and C-reactive protein (CRP) were measured by ELISA, and correlated with mortality after 6 weeks follow-up. RESULTS: Apart from IL-12, concentrations of all cytokines, TNFR p55 and CRP increased with worsening severity of disease, showing highly statistically significant trends. In a multivariable analysis high TNFR p55, IFN-gamma, CRP and low CD4 count (CD4 count <100) were predictive of mortality. Although nutritional status (assessed by body mass index, BMI) was predictive in univariate analysis, it was not an independent predictor in multivariate analysis. CONCLUSION: High serum concentrations of TNFR p55, IFN-gamma, CRP and low CD4 count correlated with disease severity and short-term mortality in HIV-infected Zambian adults with diarrhoea. These factors were better predictors of survival than BMI. Understanding the cause of TNFR p55, IFN-gamma and CRP elevation may be useful in development of interventions to reduce mortality in AIDS patients with chronic diarrhoea in Africa.

Effect of nitazoxanide on morbidity and mortality in Zambian children with cryptosporidiosis: a randomised controlled trial.

BACKGROUND: Cryptosporidiosis in children in developing countries causes persistent diarrhoea and malnutrition and is associated with increased mortality, but there is no effective treatment. We aimed to assess the effect of nitazoxanide-a new broad-spectrum antiparasitic drug-on morbidity and mortality in Zambian children with diarrhoea due to Cryptosporidium parvum. METHODS: Children with cryptosporidial diarrhoea who were admitted to the University Teaching Hospital, Lusaka, Zambia, between November, 2000, and July, 2001, and whose parents consented to their having an HIV test were randomly assigned nitazoxanide (100 mg twice daily orally for 3 days) or placebo. The primary endpoint was clinical response on day 7 after the start of treatment. Secondary endpoints included parasitological response by day 10 and mortality at day 8. Analysis was by intention to treat, with exclusion of patients subsequently found to be negative for C parvum or co-infected at baseline. The trial was stratified by HIV serology. FINDINGS: 50 HIV-seropositive and 50 HIV-seronegative children were recruited for the study, four of whom were subsequently excluded. In HIV-seronegative children, diarrhoea resolved in 14 (56%) of 25 receiving nitazoxanide and 5 (23%) of 22 receiving placebo (difference 33%, 95% CI 7-59; p=0.037). C parvum was eradicated from stool in 13 (52%) of 25 receiving nitazoxanide and three (14%) of 22 receiving placebo (38%, 95% CI 14-63; p=0.007). Four children (18%) of 22 in the placebo group had died by day 8, compared with none of 25 in the nitazoxanide group (-18%, -34 to 2; p=0.041). HIV-seropositive children did not benefit from nitazoxanide. Nitazoxanide was not significantly associated with adverse events in either stratum. INTERPRETATION: A 3-day course of nitazoxanide significantly improved the resolution of diarrhoea, parasitological eradication, and mortality in HIV-seronegative, but not HIV-seropositive, children.