Rabbit viral haemorrhagic disease (RVHD) is a highly contagious, highly fatal, peracute and acute viral disease of both wild and domestic rabbits caused by rabbit haemorrhagic disease virus (RHDV). Testing for haemagglutination activity (HA) in processed liver samples is one of the cornerstones for rapid diagnosis of RHDV outbreaks in national rabbitries. However, RHDV isolates exhibiting no HA activity are increasingly being reported. The extent of deviation from classical HA activity patterns for RHDV strains in Egypt has not been investigated. This study compared the HA activity patterns of samples collected from 61 RHDV outbreaks that occurred between 1999 and 2005 to determine whether dependence on HA test (HAT) for diagnosis of RHDV outbreaks needs to be reviewed. All samples were confi rmed RHDV positive using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and western blotting. Using slide HAT, only 36.1% of samples were positive (22 samples). Plate HAT conducted at 4ºC detected an additional 16 positive samples bringing the total HA-positive samples to 38 (62.3%). Plate HAT conducted at 22ºC failed to detect additional positive samples. The majority of samples detected after plate HA testing (62.5%) had HA titres comparable to those obtained from slide-HA-positive samples, indicating that the difference in HA activity is dependent on the nature of the HA antigen rather than its presence. Direct detection of HA activity failed in 37.7% of samples despite the presence of classical signs, pathology, and being reverse transcription polymerase chain reaction (RT-PCR) positive for three different VP60 regions. Experimental infection of seronegative rabbits with 9 HA negative RHDV samples showed that 5 isolates were in-fact HA positive, while only 4 isolates remained HA negative. The increased detection of viruses lacking HA activity and the low HAT sensitivity mandates the use of molecular techniques for rapid confi rmation of RHDV diagnosis in the Egyptian environment.
Vaccination is the major control measure for rabbit haemorrhagic disease virus (RHDV). The co-circulation of different RHDV genotypes in Egypt has led to the need to determine the most effective vaccine strain and the cross-protection between these genotypes. Rabbits seronegative for RHDV were vaccinated with the commercial GI.1a (RHDVa) vaccine strain Giza2006 and the GI.1d (G5) vaccine strain Giza97. The rabbits were challenged three weeks post vaccination with GI.1a (RHDVa) strains Giza2010 and Kal2012 and GI.1d (G5) RHDV Giza97 and RHDV2014 to determine the degree of cross-protection and evaluate immunity and cross-reactivity by haemagglutination inhibition (HI) and indirect enzyme-linked immunosorbent assay (iELISA). Both vaccines were fully protective three weeks post vaccination, with 95% protection for the GI.1a vaccine and 94.7% for the GI.1d vaccine, with no direct relationship between mortality rates and the genotype of the challenge strain. The antibody titres obtained using the HI test were one log higher for the GI.1a compared with the GI.1d vaccine, but post-challenge titres showed increased responses, expressed as 1−3 log 2 higher titres, for the GI.1d vaccine. Sequence and phylogenetic analysis of the Egyptian strain RHDV2014 revealed its relatedness to the GI.1d genotype and showed no evidence of the presence of GI.2 in Egypt until 2014. In conclusion, both GI.1d (G5) and GI.1a (RHDVa)-based vaccines are protective against both RHDV genotypes present in Egypt but continuous monitoring of circulating strains is essential because the arrival of GI.2 in Egypt will require new vaccination strategies.
The poultry industry depends heavily on immunization, particularly with live attenuated vaccines. These vaccines are commonly not adjuvanted and can be either injected or delivered in birds’ mucosa. In the current study we evaluated the protective efficacy of adjuvanted and non-adjuvanted live Newcastle disease virus (LaSota strain) vaccines. Three non-adjuvanted live NDV vaccines were used to vaccinate three groups of chickens. The same immunizations were administered to three additional groups employing adjuvant technology where a mucosal adjuvant (Montanide TM IMS 1313 nanoparticles) was used. Under experimental conditions, humoral and cellular immune responses were assessed, and challenge test was done for evaluation of the vaccine efficacy in the vaccinated chickens. RT qPCR was used for determination of viral shedding in oropharyngeal swabs of challenged chickens. Mucosal nanoparticles adjuvanted live NDV vaccines significantly improved the antibody titer and the cell mediated immune response in comparison with the non-adjuvanted ones. In the challenged chicken groups with highly virulent NDV sub-genotype VIId at the third week post vaccination, the Montanide adjuvanted vaccines were fully protective and prevention of virus shedding was also noticed while the protection rate of non-adjuvanted vaccines ranged from 90% to 100% and the virus shedding was reduced. The study indicated that, efficacy of live vaccines could be improved by using Montanide™ IMS 1313 nanoparticles adjuvant in a model of mucosal delivery of live NDV vaccine in chickens.
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