The Use Of Polymerase Chain Reaction In Diagnosis Of Sheep Pox And Goat Pox Outbreaks

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Zein Elabdeen, Ahmed
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Two outbreaks of sheeppox that occurred in Gedarif State in June 2003 and one outbreak of goatpox that occurred in Khartoum State in March 2005 were investigated. Clinical signs were reported, morbidity rates were estimated and the total mortalities were recorded. Clinically, the disease was characterized by fever, depression and eruption of generalized pox lesions. Mortality rate ranged between 5.2 and 6.7 with a mean of 6.1. All age, sex and breed groups were affected. However, more than 50% of deaths were reported in young animals in comparison to adult sheep. Skin Scabs were collected from sick animals, homogenized and inoculated in cell culture and embryonated eggs. Two virus isolates were obtained when 4 skin samples were inoculated onto Lamb testes (LT) cell culture. These viruses as well as one previous isolate of sheeppox virus (SP4) and the vaccine strain 0240 induced no lesions on the chorioallantoic membrane (CAM) of embryonated chicken eggs. On the other hand, sheeppox and goatpox isolates grew well in lamb testes (LT) cell culture with no difference in virus yield or type of CPE produced by the two viruses. In Vero cells, both viruses produced more than 90% CPE within 7 days PI. In MDBK however, both viruses induced slight CPE that reached 60% in 9 days. On the other hand, both viruses induced no CPE in chick embryo fibroblast (CEF) cells. Titration of an isolate of sheeppox virus (SP1O) was performed in Vero cells and resulted in a titer of 105.2/ml. The same virus was identified as sheeppox in virus neutralization test with 1.4 neutralization index. Polymerase chain reaction (PCR) was used in this study to determine the feasibility of improving diagnosis of sheep pox and goat pox in scabs collected from the field as well as in supernatant of infected cell cultures. Three methods of DNA extraction were tried. The first method (PCR 1) employed no DNA extraction step with scab homogenate or cell culture supernatant added directly to the PCR mix. This method failed to detect pox virus DNA in all scab homogenates tested but three out of six cell culture supernatants gave positive results. In the second method (PCR 2), viral DNA was extracted by phenol-chloroform from skin scabs; three samples out of six gave positive results. In the third method (PCR 3), viral DNA was extracted using a commercial DNA extraction Kit; two out of three gave positive result. Accordingly, the best PCR method for sheeppox and goatpox diagnosis should include an initial step of DNA extraction.