With the discovery that the coccidian parasite Cryptosporidium sp. can cause severe symptoms in humans, many diagnostic techniques were quickly implemented such as microscopic visualization, immunofluorescence and PCR...With the discovery that the coccidian parasite Cryptosporidium sp. can cause severe symptoms in humans, many diagnostic techniques were quickly implemented such as microscopic visualization, immunofluorescence and PCR. Currently, there is no effective drug treatment and none of the current diagnostic methods is 100% accurate. In this study, a BALB/C mouse was subcutaneously immunized with Cryptosporidium parvum oocysts extract. The spleen was removed and the splenocytes were fused with SP2/0 myeloma cells in order to obtain hybridoma cells secreting antibodies specific to C. parvum antigens. Human and cattle fecal samples previously characterized by microscopy [Ziehl-Neelsen staining (ZN) and Lugol] and PCR for the presence of C. parvum and Giardia duodenalis, were analyzed by indirect immunofluorescence, using the developed hybridomas supernatants. The study shows that the selected hybridomas supernatants identify C. parvum oocysts in fecal samples in correlation with C. parvum oocysts identified using ZN/PCR. No false positive results were obtained and the two best supernatants gave 20% -30% of false negative results. No cross reaction with G. duodenalis was observed. By comparing our results with those obtained with an immunofluorescence commercial kit, it suggests the potential use of the monoclonal antibodies present in two of the hybridomas supernatants as a detection tool of C. parvum. With a reliability of 80.8% and 73.1% versus ZN and PCR methods for IFI, compared with a reliability of 76.9% and 92.3% versus ZN and PCR for commercial DIF kit, the supernatant 4.1D5 seems to be the most promising subject to further study its usefulness for C. parvum detection.展开更多
Cryptosporidium spp. infection is one of the causes of diarrhea in people living with HIV/AIDS. The objective of this study is to compare the sensitivity of microscopy and molecular biology to determine the prevalence...Cryptosporidium spp. infection is one of the causes of diarrhea in people living with HIV/AIDS. The objective of this study is to compare the sensitivity of microscopy and molecular biology to determine the prevalence of Cryptosporidium spp. in Patients Living With HIV (PLWH). This is a descriptive cross-sectional study conducted in three care centers for people living with HIV/AIDS in Abidjan. It took place from November 2018 to March 2020. Sociodemographic data were obtained via a questionnaire. Stool and blood samples were collected and analyzed for microscopy and Nested PCR detection of Cryptosporidium spp. Blood samples were analyzed for CD4+ count. A total of 363 stool samples were collected from the three sites. Individuals aged 40 - 50 years (36.52%) were most likely to participate in the study. HIV Type 1 accounted for 86.22% of the study population. The samples collected consisted of 47.65% diarrheal stool. Microscopic examination of the stool yielded a prevalence of 3.86% for Cryptosporidium spp. while the prevalence was 3.96% with molecular identification. No statistically significant difference was observed between these two prevalences (χ<sup>2</sup> = 0.26;p = 0.609). CD4+ count was the factor associated with Cryptosporidium spp. infection for both microscopy (OR = 0.887, p = 0.001) and PCR (OR = 0.896, p = 0.001). This study demonstrated that Nested PCR improves the detection of Cryptosporidium spp. in patient diagnosis.展开更多
文摘With the discovery that the coccidian parasite Cryptosporidium sp. can cause severe symptoms in humans, many diagnostic techniques were quickly implemented such as microscopic visualization, immunofluorescence and PCR. Currently, there is no effective drug treatment and none of the current diagnostic methods is 100% accurate. In this study, a BALB/C mouse was subcutaneously immunized with Cryptosporidium parvum oocysts extract. The spleen was removed and the splenocytes were fused with SP2/0 myeloma cells in order to obtain hybridoma cells secreting antibodies specific to C. parvum antigens. Human and cattle fecal samples previously characterized by microscopy [Ziehl-Neelsen staining (ZN) and Lugol] and PCR for the presence of C. parvum and Giardia duodenalis, were analyzed by indirect immunofluorescence, using the developed hybridomas supernatants. The study shows that the selected hybridomas supernatants identify C. parvum oocysts in fecal samples in correlation with C. parvum oocysts identified using ZN/PCR. No false positive results were obtained and the two best supernatants gave 20% -30% of false negative results. No cross reaction with G. duodenalis was observed. By comparing our results with those obtained with an immunofluorescence commercial kit, it suggests the potential use of the monoclonal antibodies present in two of the hybridomas supernatants as a detection tool of C. parvum. With a reliability of 80.8% and 73.1% versus ZN and PCR methods for IFI, compared with a reliability of 76.9% and 92.3% versus ZN and PCR for commercial DIF kit, the supernatant 4.1D5 seems to be the most promising subject to further study its usefulness for C. parvum detection.
文摘Cryptosporidium spp. infection is one of the causes of diarrhea in people living with HIV/AIDS. The objective of this study is to compare the sensitivity of microscopy and molecular biology to determine the prevalence of Cryptosporidium spp. in Patients Living With HIV (PLWH). This is a descriptive cross-sectional study conducted in three care centers for people living with HIV/AIDS in Abidjan. It took place from November 2018 to March 2020. Sociodemographic data were obtained via a questionnaire. Stool and blood samples were collected and analyzed for microscopy and Nested PCR detection of Cryptosporidium spp. Blood samples were analyzed for CD4+ count. A total of 363 stool samples were collected from the three sites. Individuals aged 40 - 50 years (36.52%) were most likely to participate in the study. HIV Type 1 accounted for 86.22% of the study population. The samples collected consisted of 47.65% diarrheal stool. Microscopic examination of the stool yielded a prevalence of 3.86% for Cryptosporidium spp. while the prevalence was 3.96% with molecular identification. No statistically significant difference was observed between these two prevalences (χ<sup>2</sup> = 0.26;p = 0.609). CD4+ count was the factor associated with Cryptosporidium spp. infection for both microscopy (OR = 0.887, p = 0.001) and PCR (OR = 0.896, p = 0.001). This study demonstrated that Nested PCR improves the detection of Cryptosporidium spp. in patient diagnosis.
