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Molecular Epidemiology of Cryptosporium in Scale Chicken Farms in Henan Province

Author: SunYanPing
Tutor: NingChangShen；ZhangLongXian
School: Henan Agricultural University
Course: Preventive Veterinary Medicine
Keywords: Chicken Cryptosporidium Epidemiological investigation Parasite location Nested PCR Phylogenetic
CLC: S858.31
Type: Master's thesis
Year: 2008
Downloads: 28
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Abstract

Cryptosporidium (Cryptosporidium) is an important zoonotic parasitic protozoa, its wide host range, more than 240 kinds of animals, including humans, can be infected; cryptosporidiosis it caused serious harm to humans and animals. In this paper, saturated sugar flotation technique and modified acid-fast staining of large-scale farms in Henan Province cryptosporidiosis epidemiological investigation and through the the infected chicks trial and nested PCR based on the 18S rRNA gene amplification, obtained pathogenicity and molecular biological characteristics of some of the isolates, in order to provide a theoretical basis for the research and control of the molecular epidemiology of cryptosporidiosis. 1, in order to understand the scale farms in Henan Province cryptosporidiosis infection, I in August 2006 to July 2007 with a saturated sugar flotation method and modified acid-fast staining Henan scale farms investigation. Investigated a total of 10 cities in eight large-scale farms and 37 small and medium-scale farmers, check samples 2015 copies. The results show that the layers, breeders Cryptosporidium infection rate was 10.57% (163/1542), broiler infection rate was 3.17% (15/473), 0 ~ 31,31 ~ 60d chicks and 121 ~ 180d early egg Chicken relatively high infection rate, 13.70%, 24.60% and 12.87%, respectively. Grower (60 ~ 120d) and laying hens (180d later) infection rates are relatively low at 2.96%, 4.78%. Bayesian Cryptosporidium parvum (C. baileyi) according to the preliminary identification of the morphological data. The data analysis showed that the chicken Cryptosporidium insect highest infection rate in the March-May spring, the lowest rates of infection in 12 to February winter. 3d chicks with separation at the beginning of laying hens Cryptosporidium oocysts inoculated observational study the oocyst law and clinical symptoms, and tissue sections and electron microscopy parasite location. The results show that: the latent period of 4d, the oocyst open period 18d, the peak of infection to 14d. The major cause of respiratory symptoms. Tissue sections and electron microscopy can be found in other parts of the larynx, trachea, bursa, the genital cavity parasitic worms. 3, according to published Cryptosporidium 18S rRNA gene sequence of 32 Cryptosporidium isolates nested PCR product sequences are about 837 bp in length; using the restriction enzyme SspI and VspI of PCR products to digest digestion to determine the species and genotypes. SspI digestion results: after 28 Cryptosporidium isolates digested 573 bp, 267 bp two pieces, four isolates 450bp, 267bp, 111bp fragment; VspI digested results: 28 Cryptosporidium isolates digested generate 621bp, 115 bp, 104 bp fragment, four isolates produce 457bp, 171bp, 115bp fragment. There are two kinds of of C. baileyi C. meleagridis fragment length initially identified Cryptosporidium infected chickens. 4, in order to determine the species or genotype isolates from phylogenetic relationship using nested PCR amplification of the 18S rRNA gene of 32 Cryptosporidium isolates, and the amplified fragments were sequenced, sequencing sequence Blast in the NCBI, EMBL and DDBJ three nucleic acid sequence database search for homologous sequences, then to utilize the the ClustalX1.81, Phylip3.65 and DNAStar4.0 biology software on sequence comparison, to build molecular evolution tree and homology analysis. Analysis to determine the evolutionary relationships based on the 18S rRNA gene locus germline 28 isolates of C. baileyi, and isolates of the same evolutionary branches, for the same kind; four isolates with GenBank EF641025, EF179381 in the same evolutionary branches, is the same. It can determine the infected chicken Cryptosporidium C. baileyi and C. meleagridis. Comprehensive analysis based on the results of the epidemiological investigation, and domestic and foreign literature found Cryptosporidium in Henan Province farms widespread; species identification analysis showed that chickens infected with Cryptosporidium C. baileyi and C. meleagridis, which C. baileyi as the dominant species. Cryptosporidium isolated from laying hens infected chicks prove the C. baileyi mainly parasitic in chicken bursal disease and respiratory tract. Evolutionary relationship analysis based on 18S rRNA gene nested PCR amplification and restriction enzyme digestion kinds germline confirmed, the 28 isolates C. baileyi, 4 isolates C.meleagridis. 11 isolates part of the nucleotide mutations. The test for mastering the the Henan Province farms Cryptosporidium molecular popular dynamic reference, to provide a theoretical basis for the control of zoonotic cryptosporidiosis outbreak, has some public health significance.

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CLC: > Agricultural Sciences > Livestock, animal medicine,hunting,silkworm,bee > Animal Medicine ( Veterinary Medicine) > Livestock, poultry, wildlife diseases > Poultry > Chicken
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