Establishment Of Nest PCR-DGGE Detection Method And Studies On Clinical Practice Efficacy Of Dairy Cattle Endometritis On Fungal Pathogens

Cow endometritis is a major obstetric disease which is harmful to the development of livestock production. It results huge direct and indirect losses in the worldwide each year. So far it has been studied for 100 years, but because its variable pathogens and pathological mechanisms were not clear and so many other causes, there still have no effective methods to the prevention of this disease. Therefore, the main points of this research are to find fast and effective methods of pathogen diagnosis, to explore the relative treating measures, and to give drugs more targeted. Through establishing the nest PCR-DGGE diagnosis method to cow endometritis fungal pathogen and studying the effect of clinical diagnosis, i.e., to establish accurate, rapid and effective techniques and tools for the clinical diagnosis of the etiology of cow endometritis. This research gives guidance and theory assistance for the targeted drug research and affords helps for the effective prevention and treatment of cow endometritis.This paper is divided into four parts, which the first part is that used a traditional method to isolate and to indentify the fungi in the cow's womb which the cow has suffered from endometritis, regarded this as a comparison, and provided bacteria for the establishment of method. The second part is that established a nest PCR-DGGE diagnosis method to cow endometritis fungal pathogen under the present laboratory conditions, and tested its detection limit, repeatability, and so on. The third part is that combined with the actual clinical condition to apply this method and to improve it, and compared with the traditional method and made it more suitable for practical applications. The last part is that tested the effect of diagnosis by giving drugs purposely, observed the treating condition after giving medicine, and combined with the clinical symptoms to assure the disease prognosis. The main test and result are as follow:1,Regarded pathogenic fungi which was isolated by traditional method as reference strain, and established method under the present laboratory condition. Designed universal primers by the public 18S rDNA conserved regions of the fungi, and established target genes for analysis. The result shows that nest PCR-DGGE method was with highly sensitive detection, strong specificity and good reproducibility. Compared with the traditional method, it is more quickly and more accurate. It is superior to the traditional method on clinical application, and the isolation and identification of pathogenic fungi were more comprehensive. This method is adapted to the diagnosis of cow endometritis fungi pathogen.2,Tested the diagnosis effect of drug by using this method, the result shows that it had a stronger direction to give treating drug according to nest PCR-DGGE analysis method. This method had exact effect on the treatment of fungal endometritis cows, the disease prognosis of experimental animals were very clear, it greatly shorten the treating time, and it could reduce the risk of the abuse of antibiotics.3,The researchers applied nest PCR-DGGE analysis method to make epidemiological survey on the main pathogenic fungi in cow's womb from Jilin dairy cow uterus. From the survey we found that the main pathogenic fungi were white candidiasis,candida tropicalis, which were belong to candidiasis, and aspergillus terreus. Endometritis have occurred throughout the year. Autumn and winter were the higher occurrence times of fungal endometritis.