Zoology
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Item Antibody specificities of children living in a malaria endemic area to inhibitory and blocking epitopes on MSP-119 of Plasmodium falciparum(Elsevier, 2009) Omosun, Y. O.; Adoro, S.; Anumudu, C. I.; Odaibo, A. B.; Uthiapibull, C.; Holder, A. A.; Nwagwu, M.; Nwuba, R. I.Merozoite surface protein-119 (MSP-119) specific antibodies which include processing inhibitory, blocking and neutral antibodies have been identified in individuals exposed to Plasmodium falciparum. Here we intend to look at the effect of single and multiple amino acid substitutions of MSP-119 on the recognition by polyclonal antibodies from children living in Igbo-Ora, Nigeria. This would provide us with information on the possibility of eliciting mainly processing inhibitory antibodies with a recombinant MSP-119 vaccine. Blood was collected from children in the rainy season and binding of anti-MSP-119 antibodies to modified mutants of MSP-119 was analysed by ELISA. The MSP-119 mutant proteins with single substitutions at positions 22 (Leu→Arg), 43 (Glu→Leu) and 53 (Asn→Arg) and the MSP-119 mutant protein with multiple substitutions at positions 27 + 31 + 34 + 43 (Glu→Tyr, Leu→Arg, Tyr→Ser, Glu→Leu); which had inhibitory epitopes; had the highest recognition. Children recognised both sets of mutants with different age groups having different recognition levels. The percentage of malaria positive individuals (32–80%) with antibodies that bound to the mutants MSP-119 containing epitopes that recognise only processing inhibitory and not blocking antibodies, were significantly different from those with antibodies that did not bind to these mutants (21–28%). The amino acid substitutions that abolished the binding of blocking antibodies without affecting the binding of inhibitory antibodies are of particular interest in the design of MSP-119 based malaria vaccines. Although these MSP-119 mutants have not been found in natural population, their recognition by polyclonal antibodies from humans naturally infected with malaria is very promising for the future use of MSP-119 mutants in the design of a malaria vaccine.Item Cellular responses to modified Plasmodium falciparum MSP 119 antigens in individuals previously exposed to natural malaria infection(Springer, 2009) Okafor, C. M. F.; Anumudu, C. I.; Omosun, Y. O.; Uthaipibull, C.; Ayede, I.; Awobode, H. O.; Odaibo, A. B.; Langhorne, J.; Holder, A. A.; Nwuba, R. I.; Troye-Blomberg, M.Background: MSP1 processing-inhibitory antibodies bind to epitopes on the 19 kDa C-terminal region of the Plasmodium falciparum merozoite surface protein 1 (MSP119), inhibiting erythrocyte invasion. Blocking antibodies also bind to this antigen but prevent inhibitory antibodies binding, allowing invasion to proceed. Recombinant MSP119 had been modified previously to allow inhibitory but not blocking antibodies to continue to bind. Immunization with these modified proteins, therefore, has the potential to induce more effective protective antibodies. However, it was unclear whether the modification of MSP119 would affect critical T-cell responses to epitopes in this antigen. Methods: The cellular responses to wild-type MSP119 and a panel of modified MSP119 antigens were measured using an in-vitro assay for two groups of individuals: the first were malaria-naïve and the second had been naturally exposed to Plasmodium falciparum infection. The cellular responses to the modified proteins were examined using cells from malaria-exposed infants and adults. Results: Interestingly, stimulation indices (SI) for responses induced by some of the modified proteins were at least two-fold higher than those elicited by the wild-type MSP119. A protein with four amino acid substitutions (Glu27→Tyr, Leu31→Arg, Tyr34→Ser and Glu43→Leu) had the highest stimulation index (SI up to 360) and induced large responses in 64% of the samples that had significant cellular responses to the modified proteins.Item Optimization of the T-cell proliferation assay in fascioliasis using a non-radioactive method, the Alamar Blue Assay(2009) Anumudu, C. I.; Molehin, A. J.; Nwuba, R. I.T-cell proliferation studies are traditionally carried out with radioactive reagents or fluorescent reagents that require measurement with advanced technology instrumentation. We attempted to calibrate the optimal conditions suitable for the use of a non-radioactive assay for the measurement of a T-cell proliferation assay in bovine fascioliasis, but applicable to the study of other infectious diseases in our developing 'country setting, Crude antigen extract was prepared from 15 adult Fasciola gigantica flukes: Cellular responses were detected by the proliferatjon of peripheral blood mononuclear cells (PBMC) in response to stimulation by serial dilutions of the crude antigen extract. The results showed that the antigen dilution 1:1,600 gave the highest PBMC proliferative response (Stimulation Index, S.I = 1.10± 0:2). Percentage reduced Alamar Blue was 27.3-71.6%. This suggests that the cell-mediated immune response in bovine immunity to Fasciola infection may be reliably measured in our setting with the Alamar Blue Assay.Item Cytokine profiles and antibody responses to Plasmodium falciparum malaria infection in individuals living in Ibadan, southwest Nigeria(Faculty of Medicine, Makerere University, 2009) Iriemenam, N. C.; Okafor, C. M.; Balogun, H. A.; Ayede, I.; Omosun, Y.; Persson, J. O.; Hagstedt, M.; Anumudu, C. I.; Nwuba, R. I.; Troye-Blomberg, M.; Berzins, K.Background: The ability of the host immune system to efficiently clear Plasmodium falciparum parasites during a malaria infection depends on the type of immune response mounted by the host. Study design: In a cross-sectional study, we investigated the cellular-and antibody responses in individuals with P. falciparum infection, in an attempt to identify immunological signs indicative of the development of natural immunity against malaria in Ibadan, Nigeria. Levels of IL-10, IL-12(p70), IFN-γ, and IgM, IgG and IgG1-4 subclasses in the serum of 36 symptomatic children with microscopically confirmed malaria parasitaemia and 54 asymptomatic controls were analysed by ELISA. Results: IFN-γ and IL-10 were significantly higher in the symptomatic children (p=0.009, p=0.025 respectively) than in the asymptomatic controls but no differences were seen for IL-12(p70). Estimated higher ratios of IFN-γ/IL-10 and IFN-γ/IL-12 were also observed in the symptomatic children while the asymptomatic controls had higher IL-12/IL-10 ratio. The mean concentration levels of anti-P. falciparum IgG1, IgG2, IgG3 antibodies were statistically significantly higher in the individuals >5 years of age than <5 years while anti-P. falciparum IgG3 antibodies were notably low in <5 years category. Children <5 years had higher IgM antibodies than IgG and the expression of IgG subclasses increased with age. Conclusion: Taken together, malaria infection is on a delicate balance of pro- and anti-inflammatory cytokines. The higher levels of IFN-γ seen in the symptomatic children (<6months) may be instrumental in immune-protection against malaria by limiting parasite replication. The observed variations in immunoglobulin subclass levels were age-dependent and exposure-relatedItem Antibody specificities of children living in a malaria endemic area to inhibitory and blocking epitopes on MSP-119 of Plasmodium falciparum(Elsevier, 2009) Omosun, Y. O.; Adoro, S.; Anumudu, C. I.; Odaibo, A. B.; Uthiapibull, C.; Holder, A. A.; Nwagwu, M.; Nwuba, R. I.Merozoite surface protein-119 (MSP-119) specific antibodies which include processing inhibitory, blocking and neutral antibodies have been identified in individuals exposed to Plasmodium falciparum. Here we intend to look at the effect of single and multiple amino acid substitutions of MSP-119 on the recognition by polyclonal antibodies from children living in Igbo-Ora, Nigeria. This would provide us with information on the possibility of eliciting mainly processing inhibitory antibodies with a recombinant MSP-119 vaccine. Blood was collected from children in the rainy season and binding of anti-MSP-119 antibodies to modified mutants of MSP-119 was analysed by ELISA. The MSP-119 mutant proteins with single substitutions at positions 22 (Leu→Arg), 43 (Glu→Leu) and 53 (Asn→Arg) and the MSP-119 mutant protein with multiple substitutions at positions 27 + 31 + 34 + 43 (Glu→Tyr, Leu→Arg, Tyr→Ser, Glu→Leu); which had inhibitory epitopes; had the highest recognition. Children recognised both sets of mutants with different age groups having different recognition levels. The percentage of malaria positive individuals (32–80%) with antibodies that bound to the mutants MSP-119 containing epitopes that recognise only processing inhibitory and not blocking antibodies, were significantly different from those with antibodies that did not bind to these mutants (21–28%). The amino acid substitutions that abolished the binding of blocking antibodies without affecting the binding of inhibitory antibodies are of particular interest in the design of MSP-119 based malaria vaccines. Although these MSP-119 mutants have not been found in natural population, their recognition by polyclonal antibodies from humans naturally infected with malaria is very promising for the future use of MSP-119 mutants in the design of a malaria vaccine.Item Epidemiology of malaria in children living at Igbo-Ora, South Western Nigeria(Faculty of Science Obafemi Awolowo University, Ile-Ife, Nigeria, 2008) Nwuba, R. I.; Omosun, Y. O.; Anumudu, C. I.; Adoro, S.; Odaibo, A. B.; Nwagwu, M.Malaria transmission is seasonal with higher transmission occurring in the rainy season. The burden of malaria falls mainly on children and causes anaemia and fever. Children of school going age are affected and this leads to absence from school. Blood samples were collected from children aged 10 days to 15 years in dry and rainy seasons. Parasite densities were determined by microscopy. Malaria prevalence was higher in the rainy season than in the dry season. In the dry season, 42.4% of the children studied were positive for P. falciparum. While at the end of the rainy season 48.4% of the children were malaria positive. The parasite prevalence was not significantly different between males and females. Parasite densities varied from 18 to 52174 parasites per 111 of blood. The most abundant group ranged from 1-100 (59%). There was a significant correlation between parasite density and age with the mean parasite density decreasing with age group. The study shows that malaria is more prevalent in the rainy season, and children in rural areas have high prevalence of asymptomatic parasitemia which might lead to symptomatic malaria. The results show that malaria immunity increases with age in both seasons.Item Specificities of antibodies to Plasmodium falciparum merozoite surface protein (MSP)-1 19(2002) Nwuba, R. I.; Adoro, S. A.; Anumudu, C. I.; Odaibo, A. B.; Omosun, Y.; Holder, A. A.; Nwagwu, M.In a survey of children living in South Western Nigeria, plasma levels of anti-MSP119 antibodies were not associated with patent parasitemia. Anti-MSP119 antibody titres correlated positively with age, indicating that development of antibodies to MSP119 may depend on long-term exposure to parasites. Using competitive ELISA, 82% of the samples inhibited the binding of processing-inhibitory monoclonal antibodies (mAb) 12.8 and 12.10 to immobilized recombinant MSP119. The binding of mAb 12.8 in the presence of 18% of these samples .was reduced to less than 10%. This suggest that these samples contain polyc1onal antibodies that have a similar binding specificity to that of mAb 12..S, which recognizes an epitope located in the first epidermal growth factor domain of MSP119. Our data provide useful leads for the design of an MSP119-based vaccine.Item Epidemiological factors that promote the development of severe malaria anaemia in children in Ibadan(Faculty of Medicine, Makerere University, 2007) Anumudu, C. I.; Okafor, C. M. F.; Ngwumohaike, V.; Afolabi, K. A.; Nwuba, R. I.; Nwagwu, M.Background: Effective control and management of severe malaria cases depends on a clear understanding of the local epidemiological factors and specific clinical manifestations of the disease in the different endemic regions. Objectives: To determine the prevalence of severe malaria and epidemiological factors that affect the development of malaria anaemia. Methods: A cross-sectional survey was carried out among children below 5 years of age, at the Adeoyo State Maternity Hospital, Ibadan, Nigeria. Questionnaires and case histories were taken from patients clinically diagnosed of malaria. Thus, 372 volunteers were recruited into the study from the 3131 paediatric cases that reported over the10-week period to the out-patient department (OPD) of the hospital. 229 (61.6%) of the recruited volunteers presented with fever (>37.5 oC) at consultation. These had malaria parasite and PCV tests done. Results: Clinical diagnosis was confirmed microscopically in 78% (290/372) for Plasmodium infection using thick film slides. Anaemia (PCV <28%) prevalence was 28.2%. Factors that contributed to the rapid progression of uncomplicated malaria to severe status included: age of the child, level of parasitaemia, careless response and attitude of parents or guardians to fever in the children; parents’ preoccupation with their jobs or other healthy children and unwillingness to use available health facilities. Conclusion: The study underscores the need for community involved partnership for malaria control especially through health education for the home management of malaria, especially among those experiencing some form of inequity in access to healthcare.Item Variation in the relationship between anti-MSP-119 antibody response and age in children infected with Plasmodium falciparum during the dry and rainy seasons(Elsevier, 2005) Omosun, Y. O.; Anumudu, C. I.; Adoro, S.; Odaibo, A. B.; Sodeinde, O.; Holder, A. A.; Nwagwu, M.; Nwuba, R. I.Malaria remains a major parasitic disease in Africa, with 300–500 million new infections each year. There is therefore an urgent need for the development of new effective measures, including vaccines. Plasmodium falciparum merozoite surface protein-119 (MSP-119) is a prime candidate for a blood-stage malaria vaccine. Blood samples were collected from children aged 10 days to 15 years in the months of January–March (N=351) and October–November (N=369) corresponding to the dry and rainy seasons, respectively. P. falciparum infection was determined by microscopy and enzyme linked immunosorbent assay (ELISA) was used to determine the total IgG and IgG subclasses. There was a significant increase in the mean anti-MSP-119 antibody titre in the dry season (p < 0.05), compared to the rainy season. A significantly positive correlation between the anti-MSP-119 antibody titre and parasite density (p < 0.01, r = 0.138) was observed. In the rainy season, unlike in the dry season, P. falciparum positive children had higher anti-MSP-119 antibody titres than P. falciparum negative children and this difference was significant (p < 0.05). When all individuals were grouped together, the anti-MSP-119 antibody titre increased with age in both seasons (r = 0.186 and 0.002), this increase was more apparent in the dry season. However, when the study population was divided into P. falciparum positive and negative groups, it was observed that in the rainy season, there was a negative correlation between anti-MSP-119 titre and age in P. falciparum positive individuals, while those who were P. falciparum negative had a positive correlation between anti-MSP-119 titre and age. Analysis of anti-MSP-119 IgG subclass showed that IgG1 and IgG3 mean titres were highest in both the dry and rainy seasons with an increase in the mean antibody titres for IgG1, IgG2 and IgG3 in the rainy season. In the dry season there was a positive correlation between IgG1, IgG2, and IgG3 titres with age, while IgG4 was negative, whereas in the rainy season there was a positive correlation between IgG2 and IgG4 (non-cytophilic antibodies) with age and a negative correlation for IgG1 and IgG3 (cytophilic antibodies) with age. Seasonal differences in the level of MSP-119 IgG subclass titres were observed for P. falciparum negative and positive individuals. Only samples, which were positive for IgG2 and IgG4, showed positive correlation between parasitemia and total IgG. The incidence of P. falciparum infection, which increases during the rainy season, might be an important determinant of anti-MSP-119 antibody levels in children living in Igbo-Ora and the results point to the fact that non-cytophilic antibodies to MSP-119 in children might be associated with an increase in total IgG and parasitemia.Item Clinical malaria diagnosis in adults: the value of signs symptoms and antibodies(College of Medical Sciences, University of Benin, 2004-12) Anumudu, C. .; Ukoha, U.; Nwuba, R. I.; Nwagwu, M.In the absence of microscopic examination, the high prevalence of asymptomatic malaria infections and the non-specific symptoms of the disease make clinical diagnosis difficult in highly endemic areas. Data from daily medical records 0f 111 adult volunteers obtained in a 13-month longitudinal survey were analysed using Pearson's correlation to investigate the relationship between parasitaernia and clinical symptoms and to determine the predictive strength of various clinical symptoms for malaria. Forty three percent of the subjects were blood smear positive at one or more times in the study. Parasite prevalence and clinical symptoms followed a seasonal distribution, being higher and occurring more often in the high transmission periods. High antibody responders to the circumsporozoite protein (CSP) showed lower parasite prevalence and fewer symptoms compared to the other responders. Malaria parasitaemia was significantly correlated with fever (p < 0.01). Fever, joint pain and headaches could be useful in endemic areas as symptom indicators of malaria for adults.