Sitara Swarna Rao, A (2009) Molecular Epidemiology of Cryptosporidiosis and characterization of the Human Immune Response to Cryptosporidium Spp. in Southern India. Doctoral thesis, The Tamilnadu Dr. M.G.R. Medical University, Chennai.
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Abstract
Cryptosporidiosis affects young children and HIV infected adults in the developing world. Among children it is a common cause of diarrhea in under-fives, especially in the malnourished. Infections with Cryptosporidium spp., along with Giardia and enteroaggregative E. coli, also result in long term effects on growth causing stunting and may also result in deficits of cognitive function. Among the HIV infected, Cryptosporidium spp is the most common cause of opportunistic diarrhea in less-developed countries with a clear correlation with worsening cell mediated immunity. Although the pathogen is endemic in these countries, its ability to survive in the environment for a prolonged duration can lead to diarrheal disease epidemics affecting children in the entire community. Prior studies in India identified this parasite in children and in the HIV infected in various parts of the country but little information was available on the circulating species and subgenotypes or immunity to this parasite. In this study, we have documented the prevalent species among children with diarrhea in the community in Vellore and in the hospital setting in Delhi, Trichy and Vellore. In children both in the community and among those coming to the hospital, C. hominis was the most common species with a few infections due to zoonotic species like C. parvum, C. felis and C. meleagridis. Most of the children with cryptosporidial diarrhea both in the community and in the hospital setting were under the age of 2 years. In the community, C. hominis-infected children had a significantly greater severity of diarrhea. Our findings indicate that hospitalization due to cryptosporidial diarrhea is mainly due to C. hominis, probably attributable to greater severity of disease with anthroponotic than zoonotic species. Predominance of a single subgenotype Cpgp40/15 Ia was seen in the community indicating a common source of infection, probably water borne. A spatial analysis of children with cryptosporidial diarrhea was carried out to study the transmission dynamics of cryptosporidiosis in this community using GIS technology and two space time clusters were revealed. All of the C. parvum isolates in the community were the anthroponotic subgenotype Ic/IIc. Among children in the hospital setting, a more diverse set of subgenotypes was seen. Subgenotypes Ie, Ia, Ib and Id were common in all centers and 2 novel subgenotypes of C. parvum were seen in the south Indian centers of Trichy and Vellore. Using real time PCR, when six children from the Vellore hospital based study were followed up for approximately 3 weeks, oocyst shedding decreased to low or undetectable levels within 12 days of admission in 3 cases, while in 3 cases, even at 16, 18 and 24 days post diarrhea, oocyst levels remained high long after the disappearance of any symptoms. This evidence that oocysts were shed for greater than 3 weeks following an episode of diarrhea may contribute to the prolonged effects of cryptosporidiosis, including stunting and growth retardation. In order to identify the prevalence and impact of repeated infections, 20 children with cryptosporidial diarrhea in the community were screened. Multiple episodes of cryptosporidiosis occurred in 8/20 of the children. A total of 36 episodes were seen and most were symptomatic. In most cases, there was a decrease in duration and severity of diarrhea or occurrence of asymptomatic infections after the initial episode. When children with a single and multiple episodes were compared, significant differences in WAZ (weight for age) and HAZ (height for age) Z scores between the 2 groups at 24 months was seen and the difference in WAZ persisted at 36 months. This study illustrates the importance of using molecular tools to estimate the true frequency of cryptosporidial infections and the extended duration of oocyst shedding during early childhood cryptosporidial diarrhea. We also investigated maternal, longitudinal and post-diarrheal antibody responses to specific cryptosporidial antigens, gp15 and gp40 in the children from the community in Vellore. Mothers of children who had no cryptosporidial diarrhea were seropositive more often than the mothers of children with subsequent cryptosporidial diarrhea. The commonest transitional pathway was for the children of seropositive mothers who were seronegative at 3.5 months then seroconverted at 9 months and stayed seropositive at 24 months. Children of seropositive women seroconverted earlier (N/P ratio, 0.58 at 9 months) than children of seronegative women (1.0). In serum collected before and after an episode of cryptosporidial diarrhea, there was a statistically significant rise in titer to both antigens (gp15 and gp40) tested. Pre existing antibodies to gp15 were associated with a shorter duration of diarrhea. These findings on the immune response to gp40 and gp15 antigens has important implications in evaluating these glycoprotein antigens as potential vaccine candidates for cryptosporidiosis. Cryptosporidiosis in HIV infected adults was also characterized in a hospital based study at Vellore. The prevalence of cryptosporidial diarrhea was 15% which was lower than a previous study that documented 25%. This decrease in prevalence could be attributed to several causes, including the increasing accessibility of HAART for Indian patients. Nearly 30% of cases enrolled in this study had some history of antiretroviral therapy. When CD4 counts were examined, a greater number of patients with diarrhea and cryptosporidial infections had counts below 200 cells per cu. mm and more than 90% of cases identified to have cryptosporidial diarrhea had CD4 counts below 200. Molecular typing data from this study indicated that although C. hominis was the most common species identified, around 40% of cases with diarrhea were infected with other zoonotic species. Several studies indicate that immunocompromised individuals are susceptible to a wider range of species and genotypes. A wide diversity in Cpgp40/15 subgenotypes was also seen. This is the first study to quantitate cryptosporidiosis in HIV infected patients. However, there was no difference in median Ct values between cases with and without diarrhea. Immune response to 2 immunodominant antigens, gp15 and Cp23 was characterized in this HIV infected population. More than 90% of patients were found to be seropositive for both antigens, a much greater prevalence rate than that seen in studies from the West. Cases with cryptosporidiosis had higher median levels of antibody than patients without cryptosporidiosis. When antibody levels in patients with and without cryptosporidial diarrhea were analyzed, no protective effect could be determined for either antigen tested in this study. In this study, CD4 counts did not have any impact on the seropositive status of the patient. This lack of association between antibody response and CD4 count has also been documented in previous studies and could possibly be due to the fact that CD4 counts or T cells probably play a more important role in immunity from this parasite while the antibody response may act as a marker of infection or prior exposure. This study has used multiple tools to characterize cryptosporidiosis in children HIV infected adults in India. These data will help in understanding the transmission and natural history of cryptosporidial diarrhea in India and will help formulate more rational approaches to the control of this disease, especially in the context of developing countries.
| Item Type: | Thesis (Doctoral) |
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| Uncontrolled Keywords: | Molecular Epidemiology, Cryptosporidiosis, characterization, human immune response, cryptosporidium spp., India. |
| Subjects: | Respiratory Medicine > Gastroenterology > Respiratory Medicine > Gastroenterology |
| Depositing User: | Devi S |
| Date Deposited: | 28 Jun 2017 07:44 |
| Last Modified: | 11 Sep 2022 06:38 |
| URI: | http://repository-tnmgrmu.ac.in/id/eprint/522 |
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