| The livestock industry is one of the advantageous industries in Inner Mongolia Autonomous Region,and with the improvement of breeding level,people pay more and more attention to the prevention and control of livestock and poultry diseases.After the state invested a lot of energy and financial resources to make major infectious diseases gradually controlled,the harm of parasitic diseases gradually emerged,especially coccidiosis,which caused obvious economic losses to the farming industry.For the diagnosis of coccidiosis,a comprehensive method based on the observation of clinical symptoms combined with pathogenic examination is still in use,but due to the many species of coccidia,most of which are weakly pathogenic,the detection of pathogens does not necessarily mean that the disease is present,and this method cannot confirm the diagnosis in a timely manner when multiple intestinal protozoa with similar morphology are mixed,which affects the diagnosis and prevention of the disease.For this reason,in this experiment,an epidemiological survey of coccidiosis was conducted on sheep farms in some areas of Inner Mongolia.Based on this,morphological identification of the detected sheep Eimeria Crandallis was performed to determine the species and the dominant species of sheep Eimeria Crandallis,and then a multiplex PCR assay for the dominant sheep Eimeria Crandallis and Cryptosporidium was established in response to the findings.The main findings were as follows:(1)The results of the epidemiological survey of ovine Eimeria in five banners and counties in some parts of Inner Mongolia showed that the positive rate of Eimeria was the lowest in Ejina Banner and the highest in Tokoto County,and Eimeria infection was common in some parts of Inner Mongolia,with Eimeria Crandallis and Eimeria ovinoidails as the dominant species.At the same time,screening was done to determine the method of purifying coccidia oocysts.(2)On the basis of morphological identification,PCR methods were performed for Eimeria Crandallis and Cryptosporidium,and the optimal annealing temperature of 50℃was selected for both,and their specificity and sensitivity were verified.The minimum detection limit for Eimeria Crandallis was 0.1 ng/μL,and the minimum detection limit for Cryptosporidium was 0.01 ng/μL.The final PCR assays for Eimeria Crandallis and Cryptosporidium were established.(3)Based on the previous study,a multiplex PCR method was established for Eimeria Crandallis and Cryptosporidium,and its optimal annealing temperature of 50 ℃ was screened,and then its specificity and sensitivity were validated,specific for other Eimeria ovine,Eimeria bovis,Eimeria fowl,Nematodirus and Parascaris equorum.The minimum detection limits were 0.1 ng/μL for Eimeria crandallis and 1 ng/μL for Cryptosporidium,and the accuracy and stability of the method were verified in the clinical setting.The above study clarified the current infection status of sheep coccidia,the dominant strain and the main pathogenic strain in the investigated area,and provided basic data for the method of the disease;the established PCR identification method made up for the shortcomings of the traditional morphological identification with low detection rate and rapid species identification,and the multiplex PCR method for the dominant species can quickly and accurately determine the diagnosis and treatment of the disease,which provides technical support for the timely and accurate diagnosis of Eimeria ovine and Cryptosporidiosis.It provides technical support for the timely and accurate diagnosis of Eimeria ovine and Cryptosporidiosis,facilitates the diagnosis and treatment of sheep coccidia and other GI protozoa,and lays the foundation for the effective prevention and control of GI protozoa in sheep,which is of positive significance for the development of animal husbandry and the enrichment of farmers and herdsmen,and helps to realize the strategy of science and technology in Inner Mongolia. |
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