Department of Medical Microbiology
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Browsing Department of Medical Microbiology by Author "Gyan, B."
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Item Epidemiology and Molecular Characterization of Giardia Lamblia and Cryptosporidium Sp. Infections among Children In Accra, Ghana(University of Ghana, 2013-07) Anim-Baidoo, I.; Ayeh-Kumi, P.F.; Gyan, B.; Adjei, A.A.Giardiasis and cryptosporidiosis remain as part of the commonest gastroenteritis in Ghana. The diseases are caused by the protozoan parasites, Giardia lamblia and Cryptosporidium sp. respectively. Inadequate supply of treated water and poor sanitation are some of the key factors leading to the spread of these infections. Being zoonotic diseases, it is suspected that, a large proportion of human infections could come from infected domestic and farm animals. Though, use of molecular tools has helped to understand how the diseases spread in humans, animals, and the environment, very little information is available on the epidemiology and transmission routes of G. lamblia and Cryptosporidium sp. in Ghana, and a genetic characterization of the parasite has also not been thoroughly investigated. Information on clinical manifestations of Giardia infections and co-infections with other diarrhoeal causing agents particularly rotavirus remain scanty. In the present study, the epidemiology and molecular characterization of the two parasitic infections were investigated. The study, a prospective cross-sectional hospital and community-based, was conducted in Accra, Ghana. A total of 485 patients comprising of 365 diarrhoeic and 120 non-diarrhoeic children of age ≤ 5 years, were studied. Stool samples were tested microscopically, and by enzyme immunoassay kits. Positive samples were tested by the semi- nested polymerase chain reaction (PCR) and subsequently characterized into genotypes by PCR-RFLP, and nucleotide sequence analysis. Demographic and clinical data were obtained by a structured questionnaire. In the hospital-based study, prevalence rates of 5.8% and 22.0% were observed for G. lamblia and Cryptosporidium sp. infections respectively, and prevalence in diarrhoeic children was significantly higher than non-diarrhoeic children (P< 0.0001). Infection in day care centres was 10.1% for G. lamblia and 4.2% for Cryptosporidium. Neither gender nor breastfeeding habits, education level of mother, presence of domestic animals, source of children’s food, seasons (dry or rainy) was a risk factor for infections of the two parasites. However, age and source of drinking water were identified as associated risk factors for infection. G. lamblia genotype B and Cryptosporidium parvum were identified in the genotyping study. Although severity of rotaviral diarrhoea was reduced by Giardia co-infection, the results cannot be conclusive. Although both parasites were present in the studied population, cryptosporidial diarrhoea appears to be more common than giardial diarrhoea. The presence of infections among non-diarrhoeal children is of much concern, as they can spread infections unknowingly. The presence of genotype B as the only prevailing genotype of G. lamblia indicates that infections from animals will be uncommon, but Cryptosporidium parvum transmission could be either anthroponotic or zoonotic. The co- infection study had a limitatio n, and therefore demands further investigation. Several Cryptosporidium isolates that were successfully sequenced but whose identity were not clear, need further investigation as they could be new species emerging.Item Phenotypic Characterization of Host-Pathogen Interaction on Mycobacterium Africanum(University of Ghana, 2015-12) Tetteh, J.K.A.; Newman, M.J.; Yeboah-Manu, D.; Gyan, B.; University of Ghana, College of Health Sciences School of Biomedical and Allied Health Sciences Department of Medical MicrobiologyBackground: Mycobacterium africanum (MAF) and Mycobacterium tuberculosis sensu strico (MTBss) are two members of a closely related bacterial species of Mycobacterium tuberculosis complex (MTBC) that causes tuberculosis (TB) in humans. However MAF is known to cause up to 50% of human pulmonary TB in West Africa only. MAF has been sub-divided into MAF West African 1 (MAF1) (Lineage 5) and MAF West African 2 (MAF2) (Lineage 6), as two distinct phylogenetical lineages within MTBC. Subsequently the absence of Mycobacterium tuberculosis deletion gene (TbD1) strains in MTBss has been referred to as modern lineage whilst ancient lineage (MAF1 and MAF2) have the presence of TbD1. Ghana represents one of the few countries within Central West Africa known to have this unique genetic diversity of MAF1, MAF2 and MTBss that causes TB cases in significant proportions. While previously it was thought MAF is genetically very closely related to MTBss such that there are no important phenotypic differences between the two species, current advance in molecular biology indicate that substantial genetic difference exit between the two that can translate into significant phenotypic differences including immunogenicity and virulence. Aim: The aim of the study was to analyze the phenotypic features of host-pathogen interaction in Mycobacterium africanum and compared to Mycobacterium tuberculosis sensu strico strains. Methodology: The study was embedded in 2 different projects. Retrospective archived cryopreserved peripheral blood mononuclear cells (PBMCs) of MAF-infected and MTBss-infected patients were stimulated with growth medium (negative control), Staphylococcus enterotoxin B (SEB, positive control) and recombinant early secreted antigenic protein 6 kiloDalton/culture filtrate protein 10 kiloDalton fusion protein (rESAT-6/CFP-10), surface stained for T-subsets (CD4 and CD8) and intracellular cytokine, interferon gamma (IFN-), and acquired with FACS Calibur flow cytometer. The second study used characterized large sequence polymorphism (LSP) clinical isolates identified as MAF1, MAF2 and MTBss to determine intracellular growth assay in human monocyte–derived macrophages (MDM), mean doubling time and pro-inflammatory tumour necrosis factor–alpha (TNF-α), interleukin 6 (IL-6) and 12p70 cytokines by enzyme-linked immunosorbent assay (ELISA). Results: The percentage frequencies of CD4+IFN-+ and CD8+ IFN-+ T cells of MAF-infected patients did not differ from the percentage frequencies CD4+ IFN-+ and CD8+ IFN-+ T cells of MTB-infected patients in response to rESAT-6/CFP-10 fusion protein (p>0.05). Uptake of MAF1, MAF2 and MTBss representing modern and ancient strains respectively at 4hours was not significant (p>0.05). Mean intracellular growth index from 24hours to 72hours was significantly rapid for MTBss (modern) lineage compared to MAF1 and MAF2 (ancient) lineages (p<0.05). In contrast the mean doubling time of MTBss (modern) lineage was significantly lower compared to MAF1 and MAF2 (ancient) lineages (p<0.05). Levels of pro-inflammatory cytokines released into the supernatants by MTBss, MAF1 and MAF2 at 4hours was not statistically significant (p>0.05). However at 24hours to 72hours levels released by MAF1 and MAF2 (ancient) lineages was significantly higher than MTBss (modern) lineage (p<0.05). Conclusion: The study has shown that MAF-infected patients had similar T subset response to rESAT-6/CFP-10 fusion protein relative to MTBss-infected patients. Furthermore MAF had reduced uptake, low intracellular growth rate and a higher doubling time in MDM. Likewise MAF (ancient) lineages have hyper-inflammatory response thereby inducing a ‘slow growth’ phenotype highlighting the point that MAF indeed has lower virulence and longer latency leading to slower progression to active disease in the host.