Epidemiological Survey On Sheep Mycoplasma Pneumoniae And Partial Biological Characteristics Of Isolates From Shawan County, Xinjiang

Mycoplasma pneumoniae of sheep, also known as “contagious ovine pleuropneumonia”, is a kind of chronic respiratory disease caused by mycoplasma ovipneumoniae(MO), which is characterized by the symptoms of fever, dyspnea, cough, runny nose, progressive emaciation and interstitial proliferative pneumonia. It is mainly prevalent in countries and regions with large scales of sheep husbandry. MO was firstly separated in 1982 in Sichuan, China. Then it also spread in Inner Mongolia, Xinxiang, Qinghai, Tibet and some other areas, which caused large economic loss in animal husbandry and posed a serious threat to the rapid development of China’s sheep husbandry. Therefore, it is of important significance to carry out epidemiological survey on sheep mycoplasma pneumonia, and analyze the biological characteristics of the local epidemic strains for scientific prevention and control of the disease. In order to understand the infection and prevalence status of sheep mycoplasma pneumonia in Shawan county of Xinxiang, epidemiological investigation was conducted in this study through isolation, identification and genetic variation analysis of the MO epidemic strains, and the study of partial biological characteristics and drug sensitivity of these isolates. The main research contents and results were as follows:1. Epidemiological survey on sheep mycoplasma pneumonia in Shawan county of XinxiangTwenty-seven flocks of sheep from eighteen farms in seven towns of Shawan were investigated by the investigation methods of serology, etiology, field epidemiology and pathological anatomy. The results of serological survey showed that infection rates were between 7.14% and 20.00%, with average rate of 11.92%. PCR detection for nasal swabs of sheep with suspected clinical symptoms showed that the positive rates were between 58.33% and 77.78%, with average rate of 66.27%. The morbidity mainly concentrated in spring and winter. The infection rates varied significantly among different breeds of sheep. The infection rate of Hu sheep reached 85.71%, which is higher than that of small-tailed Han sheep(57.14%) and Altay sheep(48.15%). The 0-3-month-old sheep were susceptible populations with the infection rate as high as 68.57%. The infected sheep mainly displayed respiratory symptoms such as cough, runny nose, progressive emaciation and hyperplasia of pneumonia. Pathologic changes in the lungs of the dead sheep were in lesions of pneumonia and partly became meat-changed. It was confirmed that MO infection was commonly in sheep to some degree in the seven towns of Shawan, among which the situation in Bor Tong Gu Township is the worst.2. Isolation, identification and genetic variation analysis of Shawan MO epidemic strainsThree strains of colony were isolated from suspected infection epidemic materials by using improved KM2 medium, which display typical characteristics of MO colony. Then the isolates were identified by molecular biological techniques. 16 S r RNA and antigen P80 gene sequence was amplified from the isolates by PCR and sequenced. The genetic evolution was analyzed by sequence alignment. There were 99.35% to 99.68% identities in the nucleotide of 16 S r RNA gene sequences among the three strains and MO SC01 strains. The results of genetic evolution indicated that the isolated strains were MO. The nucleotide identities of P80 gene sequence between the separated MO strains and French 14811 MO strains is 97.11%, and the identities of deduced amino acid sequences was 98.28%. There existed 60 base differences in the P80 gene, of which 24 were synonymous mutations, 36 were mutations, and 12 deduced amino acid mutations, which confirmed that Shawan MO epidemic strains varied genetically from that of other areas.3. Partial biological characteristics and drug sensitivity of the MO epidemic strainsUsing biochemical reactions, biological membrane preparation and drug sensitivity analysis were conducted for the three MO epidemic strains. The results showed that the strains could not only ferment glucose, but also pigment. After dyed for 60 h, the strains can all produce thin biofilms, but with not obvious mesh structure. According to the results of the sensitivity against 16 kinds of drugs, these isolates were all highly sensitive to ciprofloxacin, ofloxacin and azithromycin, the MIC are 0.013-0.026, 0.0195-0.026 and 0.016-0.026μg/m L, respectively; there were all medium sensitive to florfenicol, enrofloxacin, moxifloxacin, acheomycin, lincomycin and clarithromycin, the MIC were 0.104-0.208, 0.065-0.130, 0.104-0.156, 0.065-0.104, 0.365-0.417 and 0.417-0.521μg/m L, respectively; there were all low-alcohol sensitive to terramycin, doxycycline, erythromycin, minocycline and gentamicin, the MIC were 1.042-1.667, 0.833-1.458, 1.042-1.667, 1.25-2.083 and 2.083-3.3331μg/m L, respectively; not sensitive to kanamysin; Only SW-3 strain was slightly sensitive to tylosin while the other two strains were highly sensitive. Correlation analysis showed that there were variations of 20 points of 23 S r RNA genes of the strains slightly sensitive to tylosin, compared with those slightly sensitive strains, including 559 T-C, 753 G-A, 1084 T-C, 1358 A-G, 1381 A-G, 1645 T-C, 1652 C-T, 1653 T-C, 1689 C-T, 1695 A-C, 1696 T-A, 1697 A-G, 1761 T-C, 1817 T-C, 1914 T-C, 2123 G-A, 2628 G-A, 2629 T-C, 2789 G-T, 2830 C-T, from which it was speculated that these variations might be related to the sensitivity of the strains against tylosin.In all, epidemiological survey on mycoplasma ovipneumoniae of sheep in Shawan county was carried out, and the genetic evolution of 16 S r RNA and antigen P80 gene sequences of the Shawan epidemic strains were analyzed, then partial biological characteristics were determined, which provided epidemiological data for the effective prevention and control of the disease in Shawan county.