| Cryptosporidium is an important zoonotic pathogen, which is also one of the two pathogens specified in the drinking water quality standards in China and many other countries. However, thus far, most studies of distribution, transmission and source tracking of Cryptosporidium in water and humans are carried out in developed countries, and little is known in China.To identify the role of waterborne transmission of cryptosporidiosis in Shanghai, China, in this study,50 source water samples from Huangpu River,30 tap water samples and 28 swimming pool water samples were collected and concentrated. Cryptosporidium species present in the samples were detected and identified by PCR-RFLP (restriction fragment length polymorphism) analysis. Of the source water samples,28.0%(14/50) were positive for Cryptospordium. Four species, including Cryptosporidium andersoni from cattle, C. suis from pig, and C. baileyi and C. meleagridis from poultry, were identified. Among the four species, only C. meleagridis is considered as a human pathogenic parasite, however, C. meleagridis was only detected in one sample.30 tap water samples and 28 swimming pool water samples were all Cryptosporidium negative in this study.To facilitate the characterization of the endemic transmission of cryptosporidiosis in human populations in Shanghai,90,87,109, and 100 raw domestic wastewater samples were collocted from Shanghai and three control cities, including Nanjing, Qingdao, and Wuhan, respectively. The positive rates of Cryptosporidium in wastewater samples from the four cities were 70.0%,37.9%,68.8%, and 47.0%, respectively. Anthroponotic species C. hominis was found extremely popular in Cryptospordium positive samples (93.7%) in Shanghai, significantly higher than the rates in three control cities (24.2%,1.3%, and 4.4%, respectively). In addition, subtyping results shows that there was a high genetic complexity of C. hominis in Shanghai. Subtype families including la, Ib, Id, Ie, and If were all found, with one subtype IbA21G2 the most prevalent (46.8%). The high C. hominis heterogeneity is likely an indicator of endemic cryptosporidiosis transmission in Shanghai, while the dominant subtype IbA21G2 is likely an indicator of cryptosporidiosis outbreaks among humans.Children are vulnerable to Cryptosporidium infection. To characterize the distribution and transmission of Cryptosporidium species in children in Shanghai, in this study,6,284 fecal samples were collected from three pediatric hospitals. Among them,102 (1.6%) were infected with Cryptosporidium. Four Cryptosporidium species were identified, including C. hominis (92 cases), C. meleagridis (6 cases), C. canis (2 cases), and C. felis (2 cases).6 subtypes, including IaA14R4 (36 cases), IdA19 (37 cases), IbA19G2 (1 case), IdA14 (1 case), IaA18R4 (1 case), and IgA14 (1 case), were identified in 77 C. hominis positive samples. Since the two dominant C. hominis subtypes were absent from local wastewater samples, we therefore tracked the sources of positive samples during later period of the study. And an extended cryptosporidiosis outbreak was identified in Ward A, Hospital I during our sampling period. The outbreak was dominated by two C. hominis subtypes with different virulence and population structure; one outbreak subtype IaA14R4 was significantly related with diarrhea while the other subtype IdA19 was not. In addition, by population genetics analysis, we found IaA14R4 had an epidemic population structure, and IdA19 had a clonal population structure.In this study, we tracked the infection source of cryptosporidiosis in children. Animal (birds, dogs, and cats) contact was likely the major transmission route for sporadic infections. The outbreak subtype IaA14R4 was linked to cryptosporidiosis in an orphanage in Shanghai, this subtype was still prevalent in children in this orphanage during later period of this study. In contrast, the other outbreak subtype IdA19 might have originated from Hospital I. Caregivers might have played an important role in the transmission of the cryptosporidiosis in these children.HIV+patients are also vulnerable to cryptosporidiosis. In this study, the infection rate of Cryptosporidium in HIV+patients from Henan, China was 1.5%(10/683). Four species were detected, including zoonotic species C. meleagridis (5 cases), C. parvum (2 cases), C. suis (1 case), and anthroponotic species C. hominis (2 cases). Analysis of risk factors also shows that zoonotic transmission was important for Cryptosporidium infections. Besides, data in this study show a connection of Cryptosporidium infection and using well water as water source, indicating the possibility of well water contamination by Cryptosporidium oocysts.Genotyping is important for source tracking of Cryptosporidium contamination/infection. We thus in this study established two LightCycler real-time PCR genotyping techniques based on fluorescence resonance energy transfer (FRET) probes. The two techniques can differentiate various Cryptosporidium species/genotypes by melting curve analysis. The SSU rRNA based technique can differentiate 5 common species in humans and most common species/genotypes in source water, whereas the HSP90 gene based technique can differentiate C. hominis, C. parvum, and C. meleagridis perfectly. The new established genotyping techniques in this study have shown a bright future of widely use.In conclusion, results in the study will help to understand the public health significance of Cryptosporidium species in China, and will help to make more efficient control strategy of cryptosporidiosis. In addition, this study will play an important role in promoting the development of Cryptosporidium genotyping and pathogen tracking technologies. |
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