Molecular Studies on Microsporidia Species and their Prevalence in Lagos, Nigeria
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Date
2010-06
Authors
Ojuromi, O.T
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Abstract
Microsporidia cases due to Enterocytozoon bieneusi and Encephalitozoon intestinalis are emerging opportunistic infections associated with a wide range of clinical syndromes in humans. A cross-sectional study was conducted to determine the prevalence of microsporidia spores, Cryptosporidium oocyst and other enteric intestinal parasites from faeces of HIV-positive individuals (with and without diarrhoea) attending the HIV/AIDS Clinics and from HIV-negative individuals and animal samples from different sites in Lagos. Fresh faecal samples were collected from 770 HIV-positive patients {225(29.2%) (CI =26.1 to 32.6%) with diarrhoea and 545 (70.8%) (CI = 67.4 to 73.9%)} without diarrhoea of age range from 2-70 years (mean age 34.6 years); from 250 HIV- negative patients (age range 2-50 years; mean age = 25.0 years) and 250 animal samples. Microscopy was used for initial screening of all faecal samples using wet mount microscopy with 5% Lugol’s iodine, modified Weber green chromotrope stain (Trichrome stain) and Kinyoun carbol fuschin stain for the detection of enteric intestinal parasites, microsporidia spores and Cryptosporidium oocysts respectively. The prevalence rate of microsporidia infection was 51(6.6%). Of the 225 diarrhoeic faeces, 36 (16.0%) were positive for microsporidia spores, 41(80.4%) for Enterocytozoon bieneusi and 10 (19.6%) for Encephalitozoon species and 49 (6.4%) for Cryptosporidium oocysts. Of the 545, 15 (2.8%) and 13 (2.3%) patients were also positive for both organisms in non-diarrhoeic faeces respectively. Mixed infection was recorded in 14 (28.1%) between microsporidia spores and Cryptosporidium oocysts. Of the males, 21(6.3%) had microsporidia spores compared with Cryptosporidium oocysts 22 (6.6%) while the 30 (6.8%) of the females and 27(6.2%) males were positive for microsporidia spores and Cryptosporidium oocysts respectively (P value = 0.0154; P value = 0.0146). Similarly, among the diarrhoeal patients with CD4+ T-lymphocyte cell count < 200, 4.9% and 10.1% were positive for both microsporidia spores and Cryptosporidium oocysts.
Enteric parasites were identified among 89 (36.5%) individuals who were HIV-negative, of which Cryptosporidium accounted for 10 (11.2%). Other parasites found that were significantly associated with diarrhoea were Giardia lamblia (38.2%), Entamoeba histolytica/dispar (16.8%) and Trichuris trichiura (20.2%). Microsporidia spores were not detected among the HIV-negative infected patients. Microsporidia spores were detected more in rams (16.0%) and the least infection rate was found in cattle (12.0%). Cryptosporidium oocysts was detected in 24 (96%) of rabbit samples examined. Using polymerase chain reaction (PCR) and primers that amplified the conserved region of the small subunit rRNA (SSU-rRNA) of E.bieneusi, E.intestinalis. E.hellem and E.cuniculi. 15 (29.4%) were positive for E.bieneusi, 2 (3.92%) for Encephalitozoon intestinalis. Using Immunofluorescence antibody test (IFAT), all samples positive by specie-specific primers were also positive using monoclonal and polyclonal antibodies except for one sample that was positive for E.cuniculi. However, IFAT detected Enterocytozoon spores in 6 (24.0%) turkeys, 2 (8.0%) goats, 3 (12.0%) cattle and 4 (16.0%) rams. All E.bieneusi-positive samples genotyped for the ITS of the rRNA showed that 7 out of 15 specimens positive for E.bieneusi were similar or identical to the previously reported genotypes. Genotype K was found to be more predominant (n=4), Peru 6 or Genotype A (n=1), Peru 6 var (n=1) and Genotype B (n=1). Cryptosporidium parvum and C. hominis from HIV-infected patients was characterized by PCR-RFLP and sequencing analysis of the 60-Kda glycoprotein gene (GP60 gene). Two alleles were identified, each corresponding to IIeA11T3G3 and IIcA5G3 that are restricted to humans. The phylogenetic analysis of the genotypes of microsporidia showed close identity with those reported from birds, muskrats and humans. In our view, this is the first report of microsporidia detection in HIV-infected patients and animal samples and sequencing of Cryptosporidium from Lagos, Nigeria. These results highlight the inclusion of microsporidial diagnosis and other opportunistic infections in the management of HIV/AIDS infection in Nigeria.
Description
A Thesis Submitted to the School of Postgraduate Studies, University of Lagos
Keywords
Microsporidia , Encephalitozoon intestinalis , Enterocytozoon bieneusi , Clinical syndromes , Research Subject Categories::MEDICINE::Microbiology, immunology, infectious diseases
Citation
Ojuromi, O.T (2010). Molecular Studies on Microsporidia Species and their Prevalence in Lagos, Nigeria. A Thesis Submitted to University of Lagos School of Postgraduate Studies Phd Thesis and Dissertation, 177pp