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Seminar 789  
Seminar:  789
Kuan, Nan-Ling
The Development of a Toxoid Vaccine againstActinobacillus pleuropneumoniae
Actinobacillus pleuropneumoniaeAPis the pathogen that causes hemorrhagic, necrotizing, fibrinous pleuropneumonia in swine. Out of 15 known serotypes of AP, serotypes 1, 5 and 2 are the predominant strains in Taiwan. With a complex pathogenicity, ApxI is one of the most important virulence factors of AP. The aim of this study is to select the appropriate seed strains, so as to develop a multivalent inactive vaccine with important toxins.
  The best toxin-producer was selected as the seed strain using a hemolytic titer test and immune dots from 5 isolates of AP serotype 1 (AP1). In establishing an optimal vaccine composition, the effective concentration of toxins and AP1 were tested. ICR mice were immunized with the crude toxins and the inactivated AP1, with the defense index (DI) evaluated two weeks later after immunization. In the test of effective concentration of AP1, it was found that the DI forthe 10X concentrated toxins with different concentration of AP1 ? 10CFU/ml, 108 CFU/ml, 10CFU/ml and without AP1 ? were 1.5, 1.4, 0.7 and 0.7, respectively. In another test of effective concentration of toxins, the DI for the 10CFU/ml of AP1 with different concentration of toxins ? 10X, 5X, 2.5X concentrated and without toxins ? were 1.2, 1.2, 1.0 and 1.0, respectively. As a result, the 10X concentrated toxins and 108CFU/ml of AP1 were ensured to be the basic components, while AP5 and AP2 will be added into the formula in the future.
Shih, Yu-Hua
The Application of a One-Step Real-Time RT-PCR Assay for the Detection of Duck Hepatitis Virus
Duck virus hepatitis (DVH) causes opisthotonus and hepatitis among ducklings, resulting in acute death and rapid transmission among ducklings within a flock. Duck hepatitis can be caused either by DHAV-1, DHAV-2, DHAV-3, DHV-II or DHV-III. Among them, DHAV-1 is the most virulent strain and is widely distributed. The susceptibility of ducks for DHAV-1 infection decreases as age increases. The disease can cause a mortality rate of up to 80% for ducklings under the age of 3-weeks, but decreases when the age reaches over 4-weeks.
Diagnosis of duck hepatitis cases usually depend on clinical symptoms, pathological lesions, and virus isolation. Methods of diagnoses include neutralization tests, ELISA, and RT-PCR, among others. However, these tests are laborious and time consuming. Therefore, this study aimed to develop a one-step real-time reverse transcription polymerase chain reaction (one-step rRT-PCR) to detect and distinguish between DHAV-1 and DHAV-2. In this study, we detected 5 cases of duck hepatitis in 2008 and 11 cases in 2009 with all cases proving to have been caused by DHAV-1. After applying the rRT-PCR assays for DHAV-1 and DHAV-2, the DHAV-1 viral nucleic acids were detected in the lungs, liver and intestine of 1-day-old ducklings with DHAV-1, at 24th hour post inoculation, and the DHAV-2 viral nucleic acids were detected in the lungs, pancreas, liver and intestine of 1-day-old ducklings with DHAV-2 at 32nd hour post inoculation. These findings suggests that the real-time RT-PCR assays we developed are  fast, highly sensitive and specific diagnostic methods; they will help and improve the identification of duck hepatitis cases once its effective application has been established in the laboratory.