Impregnation and storage of Newcastle disease virus on to filter papers and detection of viral RNA by a single tube RT-PCR assay

Suitability of storing infected allantoic fluid (AF) and cell culture supernatants (CCS) with strain I-2 of Newcastle disease virus (NDV) on Whatman filter papers at room temperature (22-25 °C) and 37 °C was determined. RNA was extracted from filter papers or liquid aliquots and subjected to reverse transcriptase-polymerase chain reactions (RT-PCR). The results showed that filter papers soaked with NDV infected AF or CCS stored at 37°C yielded amplicons with intensity similar to that kept at room temperature for up to 150 days. The study demonstrates that NDV infected samples can be soaked onto filter papers, stored and subsequently detected by RT-PCR. This method might be safely used for storage and transportation of NDV samples to the designated laboratories for molecular studies without the need for cooling

A case study of Ngorongoro Conservation Area, Arusha, Tanzania

Tanzania Veterinary Journal Vol. 26, (1): 2009; pp 44-50A limited study was conducted to determine prevalence of brucellosis in domestic ruminants kept in a free range grazing system in Ngorongoro Conservation Area (NCA) which is a world heritage site in which pastoralists communities have been living harmoniously with wildlife for decades. Blood samples from 200 cattle, 87 goats and 13 sheep were collected by venipuncture into plain vacutainer tubes. Rose Bengal Plate Test (RBPT) and Microagglutination Test were used to detect antibodies against brucellosis in sera obtained from sampled blood. It was observed that 14.28% adult cows, 7.54% heifers, 2.38% bulls, 11.9 does, 10.7% bucks, and 10% ewes showed positive reactions to RBPT. When same samples were tested with MAT, 10.05% adult cows, 7.54% heifers, 2.38% bulls, 13.8% does, 14.3% bucks, and 3% ewes tested positive. Based on these serological tests it was concluded that brucellosis is endemic in pastoral livestock in NCA and that the reported increase in human brucellosis among pastoralists living in the NCA might be associated with domestic ruminants which are the sole source of food and income for the pastoralist in the area. Wildlife-domestic animals interaction phenomenon in NCA can as well be viewed as a significant means with which zoonoses are maintained in such ecosyste

Genomic sequence of infectious bursal disease virus from Zambia suggests evidence for genome re-assortment in nature

Infectious bursal disease virus (IBDV) is a bi-segmented RNA virus, which belongs to the genus Avibirnavirus of the family Birnaviridae. Two serotypes, 1 and 2, exist in IBDV. The serotype 1 IBDVs are the causative agents of infectious bursal disease (IBD) in chickens worldwide and lead to immunosuppression in young birds. Genome re-assortment has been speculated to occur and contribute to the emergence of new IBDV strains. However, evidence was lacking until recently when two re-assortant viruses were detected in China. In this study, we determined the complete nucleotide sequence of an IBDV, designated KZC-104, from a confirmed natural IBD outbreak in Lusaka, Zambia in 2004. The genome consisted of 3074 and 2651 nucleotides in the coding regions of segments A and B, respectively. Alignment of both nucleotide and deduced amino acid sequences, and phylogenetic analysis revealed that the genome segment A of KZC-104 was derived from a very virulent strain, whereas its segment B was derived from a classical attenuated strain. On BLAST search, the full-length segments A and B sequences showed 98% closest nucleotide homology to the very virulent strain D6948 and 99.8% closest nucleotide homology to the classical attenuated strain D78, respectively. This is a unique IBDV reassortant strain, which has emerged in nature involving segment B of a live attenuated vaccine. This observation provides direct evidence for the involvement of vaccine strains in the emergence of reassortant IBDV in the field. Taken together, these findings suggest an additional risk of using live IBDV vaccines, which may act as genetic donors for genome re-assortment. Further studies are required to investigate the epidemiology and biological characteristics of reassortant strains so that the appropriate and safe IBDV vaccines can be recommended

Deduced amino acid sequences surrounding the fusion glycoprotein cleavage site and of the carboxyl-terminus of haemagglutinin-neuraminidase protein of the avirulent thermostable vaccine strain I-2 of Newcastle disease virus

A single-tube RT-PCR technique generated a 387 bp or 300 bp cDNA amplicon covering the F-0 cleavage site or the carboxyl (C)-terminus of the HN gene, respectively, of Newcastle disease virus (NDV) strain 1-2. Sequence analysis was used to deduce the amino acid sequences of the cleavage site of F protein and the C-terminus of HN protein, which were then compared with sequences for other NDV strains. The cleavage site of NDV strain 1-2 had a sequence Motif of (112)RKQGRLIG(119), consistent with an avirulent phenotype. Nucleotide sequencing and deduction of amino acids at the C-terminus of HN revealed that strain 1-2 had a 7-amino-acid extension (VEILKDGVREARSSR). This differs from the virulent viruses that caused outbreaks of Newcastle disease in Australia in the 1930s and 1990s, which have HN extensions of 0 and 9 amino acids, respectively. Amino acid sequence analyses of the F and HN genes of strain 1-2 confirmed its avirulent nature and its Australian origin