Biomedical subjects

S Sapats

Publications and source records attributed to S Sapats.

8 recordsLinked to original sources

PubMed2006

Isolation of a variant infectious bronchitis virus in Australia that further illustrates diversity among emerging strains.

Australian infectious bronchitis viruses (IBV) have undergone a separate evolution due to geographic isolation. Consequently, changes occurring in Australian IBV illustrate, independently from other countries, types of variability that could occur in emerging IBV strains. Previously, we have identified two distinct genetic groups of IBV, designated subgroups 1 and 2. IBV strains of subgroup 1 have S1 and N proteins that share a high degree of amino acid identity, 81 to 98% in S1 and 91 to 99% in N. Subgroup 2 strains possess S1 and N proteins that share a low level of identity with subgroup 1 strains: 54 to 62% in S1 and 60 to 62% in N. This paper describes the isolation and characterisation of a third, previously undetected genetic group of IBV in Australia. The subgroup 3 strains, represented by isolate chicken/Australia/N2/04, had an S1 protein that shared a low level of identity with both subgroups 1 and 2: 61 to 63% and 56 to 59%, respectively. However, the N protein and the 3' untranslated region were similar to subgroup 1: 90 to 97% identical with the N protein of subgroup 1 strains. This N4/02 subgroup 3 of IBV is reminiscent of two other strains, D1466 and DE072, isolated in the Netherlands and in the USA, respectively. The emergence of the subgroup 3 viruses in Australia, as well as the emergence of subgroup 2 in 1988, could not be explained by any of the mechanisms that are currently considered to be involved in generation of IBV variants.

Amino Acid Sequence↗

PubMed2006

Chicken recombinant antibodies against infectious bursal disease virus are able to form antibody-virus immune complex.

Virus particles exposed to specific anti-virus antibodies result in the formation of immune complexes (Icx). Recent vaccination strategies have employed this feature, and an infectious bursal disease virus (IBDV) vaccine based on Icx has been released and is expected to replace conventional IBDV vaccines. We evaluated whether chicken recombinant antibodies (rAb) specific for IBDV, rather than conventional chicken anti-IBDV sera, could be used to generate Icx. Out of 14 rAb expressed as soluble single-chain variable fragments (scFv), nine were able to completely neutralize Bursavac, a live IBDV vaccine, when tested in ovo. When these rAb were mixed with IBDV and inoculated into either 18-day-old embryos, or 1-day-old or 2-week-old specific pathogen free chicks, a rAb.IBDV complex was formed. These Icx were similar to those produced by polyclonal chick anti-IBDV sera and IBDV. Following inoculation of the rAb.IBDV complex, the virus was rendered non-infectious for 5 to 7 days. After this time virus was released from the Icx, resulting in infection of the inoculated chicks and subsequent induction of an immune response and protection against virulent IBDV challenge. The results indicated that genetically derived antibodies can replace polyclonal sera in the formulation of Icx vaccines.

Animals↗

PubMed2005

Identification of previously unknown antigenic epitopes on the S and N proteins of avian infectious bronchitis virus.

This paper describes mapping of antigenic and host-protective epitopes of infectious bronchitis virus proteins by assessing the ability of defined peptide regions within the S1, S2 and N proteins to elicit humoral, cell-mediated and protective immune responses. Peptides corresponding to six regions in the S1 (Sp1-Sp6), one in the S2 (Sp7) and four in the N protein (Np1-Np4) were synthesized and coupled to either diphtheria toxoid (dt) or biotin (bt). Bt-peptides were used to assess if selected regions were antigenic and contained B- or T-cell epitopes and dt-peptides if regions induced an antibody response and protection against virulent challenge. All S1 and S2 peptides were antigenic, being recognised by IBV immune sera and also induced an antibody response following inoculation into chicks. Three S1-and one S2-bt peptides also induced a delayed type hypersensitivity response indicating the presence of T-cell epitopes. The S2 peptide Sp7 (amino acid position 566-584) previously identified as an immundominant region, was the most antigenic of all peptides used in this study. Two S1 (Sp4 and Sp6) and one S2 peptide (Sp7), protected kidney tissue against virulent challenge. From four N peptides located in the amino-terminal part of the N protein, only one, Np2 (amino acid position 72-86), was antigenic and also induced a delayed type hypersensitivity response. None of the N peptides induced protection against virulent challenge. The results suggest that the S1 glycoprotein carries additional antigenic regions to those previously identified and that two regions located in the S1 and one in the S2 at amino acid positions 294-316 (Sp4), 532-537 (Sp6) and 566-584 (Sp7) may have a role in protection.

Animals↗

PubMed2005

An ELISA for detection of infectious bursal disease virus and differentiation of very virulent strains based on single chain recombinant chicken antibodies.

Two chicken single-chain variable antibody fragments (scFv) designated scFv154 and scFv88, previously shown to react with either all or very virulent (vv) infectious bursal disease virus (IBDV) strains, respectively, were evaluated for use in an enzyme-linked immunosorbent assay (ELISA) for differentiation of vvIBDV. Specificity and sensitivity of the vvIBDV ELISA was assessed when scFv154 and scFv88 were expressed as soluble antibodies (sAb), phage antibodies (pAb) or hyper-phage antibodies (hpAb). The highest test sensitivity and specificity was obtained using hpAb154 to detect all IBDV and pAb88 to differentiate vvIBDV strains. Such an ELISA was eight to 16 times more sensitive for IBDV antigen detection than the mouse monoclonal antibody ELISA. Using field samples, the scFv ELISA was able to differentiate between flocks infected with vvIBDV and those infected with classical or variant IBDV. In one instance IBDV was detected in a flock found to be negative by the monoclonal antibody ELISA. The results showed that scFv can be utilized as highly specific and sensitive ELISA reagents for the detection and discrimination of avian pathogens.

Animals↗

PubMed2004

Virus strains from a flock exhibiting unusually high mortality due to infectious bursal disease.

OBJECTIVE: To characterise infectious bursal disease viruses (IBDVs) isolated from commercial broiler flocks exhibiting unusually high mortality due to infectious bursal disease (IBD). DESIGN: An IBD outbreak occurred in mid 1999 on two broilers farms (A and B) in northern New South Wales amongst chickens 28 to 38 days of age, with a sharp rise in mortality of 2.5%. Initial histopathological diagnosis indicated acute IBD. Since acute IBD caused by classical pathogenic and very virulent (vv) IBDVs is exotic to Australia, samples from both farms A and B were obtained and used for virus characterisation. METHOD: Tissue homogenates were made from six bursae collected from farm B. One histological sample from farm A was also used. Nucleotide sequencing of the hypervariable region (HVR) within the VP2 gene of IBDVs was determined and the deduced amino acid sequences compared with previously characterised Australian and overseas IBDVs. The phylogenetic relationship between IBDVs from farm B and IBDVs from Australia and overseas was then determined. Pathogenicity of one isolate, N2/99 from farm B, was compared with 3 other local IBDVs, as well as with three pathogenic overseas strains in 3-week-old specific pathogen-free (SPF) chickens. RESULTS: Initial histopathological characterisation of a sample of bursa from a bird on farm A showed widespread acute lymphoid necrosis, follicular haemorrhage and stromal oedema, indicative of acute IBD. Subsequent analysis using reverse transcriptase polymerase chain reaction (RT-PCR), followed by nucleotide sequencing of the same bursal sample, as well as 6 samples from nearby farm B, showed that the IBDVs involved were similar in sequence to Australian vaccine strains and not to classical pathogenic or vvIBDVs. One isolate, N2/99 from farm B, was only marginally more pathogenic than other local IBDVs. It induced mild clinical signs in 30% of chicks and no mortality. In comparison, vvIBDV CS89 and classical pathogenic 52/70 strains induced severe clinical signs in 100% and 80% of chickens, respectively with mortalities of 27% and 12%, respectively. CONCLUSIONS: The results illustrated the value of nucleotide sequencing as a method for discrimination of local and exotic types of IBDV.

Amino Acid Sequence↗

PubMed2003

Characterization of infectious bursal disease virus isolates from Indonesia indicates the existence of very virulent strains with unique genetic changes.

Sequencing of the hypervariable region of viral protein VP2 of infectious bursal disease virus (IBDV) isolates obtained from non-vaccinated chickens in Indonesia showed that the majority (16/17) were closely related to published very virulent (vv)IBDV strains. Four isolates contained identical amino acid sequences to Asian and European vvIBDVs, sharing vv-specific amino acid residues 222(Ala), 256(Ile), and 294(Ile). Eight isolates differed by one amino acid at position 222(Ala-->Ser); however, this change did not alter the pathogenicity or antigenicity of these strains. Two isolates, with amino acid substitutions at positions 272(Ile-->Thr) and 279(Asp-->Asn), did not cause clinical disease or mortality, and were therefore considered to be naturally occurring, attenuated mutants of vvIBDV. The results illustrate variability that might occur among vvIBDV strains.

Amino Acid Sequence↗

PubMed2002

Confirmation of the existence of two distinct genetic groups of infectious bursal disease virus in Australia.

OBJECTIVE: To characterise infectious bursal disease viruses (IBDVs) prevalent at major commercial sites throughout Australia and to compare the nucleic acid sequences of local strains of IBDV with those of characterised overseas strains. DESIGN: Samples of bursae were collected from 20 broiler farms that belonged to different poultry companies in New South Wales (NSW), Queensland (Qld), Victoria (Vic), Westem (WA) and South Australia (SA). METHOD: Bursae were collected from broilers between 24 and 35 days of age. Bursal tissue was homogenised and tested for the presence of IBDV antigen using four monoclonal antibodies (Mabs) which detect antigenic variation in IBDV strains. The nucleotide sequences of the hypervariable region (HVR) within the VP2 gene of IBDVs was determined and the deduced amino acid sequences compared with three vaccine strains and six previously characterised Australian IBDV strains. The deduced amino acid sequences were also compared with the published amino acid sequences of overseas strains. The phylogenetic relationships between Australian strains and overseas strains were then determined. RESULTS: IBDV was detected in birds from 14 out of 20 farms sampled. Typing with four Mabs showed that all viruses from Vic (6) and SA (10) were antigenic variants, whereas all viruses from NSW (29), Qld (4) and WA (5) were classical-like strains. Nucleotide sequencing of one sample from each of the 14 farms on which IBDV was detected confirmed results obtained with Mabs. The amino acid sequences of all Australian viruses differed from the amino acid sequences of foreign IBDV strains. Phylogenetic analysis showed that Australian IBDV viruses belonged to two distinct genetic groups. Very virulent (vv) IBDV strains belonged to a third genetic group, and overseas classical and variant strains belonged to a fourth genetic group. CONCLUSIONS: The results confirmed previous findings that there are two groups of IBDV strains circulating in commercial broilers in Australia. The majority are classical-like strains that are antigenically and genetically similar to vaccine strains 002/73 and V877. These classical strains were prevalent in broilers in three states, NSW, Qld and WA. The second group of strains are antigenic variants that were only found in broilers in two states, Vic and SA. All Australian IBDVs characterised to date are genetically distinct and can be differentiated from all other overseas strains. This enables identification of incursion of any exotic strain into Australian poultry, be it classical, US variant or wIBDV strains.

Amino Acid Sequence↗

PubMed2000

Avian infectious bronchitis virus.

Infectious bronchitis virus (IBV) is prevalent in all countries with an intensive poultry industry, with the incidence of infection approaching 100% in most locations. Vaccination is only partially successful due to the continual emergence of antigenic variants. At many sites, multiple antigenic types are simultaneously present, requiring the application of multiple vaccines. Although many countries share some common antigenic types, IBV strains within a geographic region are unique and distinct, examples are Europe, the United States of America and Australia. Measures to restrict the introduction of exotic IBV strains should therefore be considered. Infectious bronchitis has a significant economic impact; in broilers, production losses are due to poor weight gains, condemnation at processing and mortality, whilst in laying birds, losses are due to suboptimal egg production and downgrading of eggs. Chickens and commercially reared pheasants are the only natural hosts for IBV. Other species are not considered as reservoirs of IBV. The majority of IBV strains cause tracheal lesions and respiratory disease with low mortality due to secondary bacterial infections, primarily in broilers. Nephropathogenic strains, in addition to tracheal lesions, also induce prominent kidney lesions with mortality of up to 25% in broilers. Strains of both pathotypes infect adult birds and affect egg production and egg quality to a variable degree. Infected chicks are the major source of virus in the environment. Contaminated equipment and material are a potential source for indirect transmission over large distances. Virus is present in considerable titres in tracheal mucus and in faeces in the acute and recovery phases of disease, respectively. Virus spreads horizontally by aerosol (inhalation) or ingestion of faeces or contaminated feed or water. The virus is highly infectious. Clinical signs will develop in contact chicks within 36 h and in nearby sheds within one to two days. Infection is resolved within fourteen days with a rise in antibody titres. In a small number of chicks, latent infection is established with subsequent erratic shedding of virus for a prolonged period of time via both faeces and aerosol. Movement of live birds should be considered as a potential source for the introduction of IBV. Isolation and identification of IBV is needed for positive diagnosis. The preferred method of isolation is to passage a sample in embryonating specified-pathogen-free chicken eggs. Identification is either by monoclonal antibody based enzyme-linked immunosorbent assay (ELISA) or polymerase chain reaction. Virus neutralisation test in tracheal organ culture is the best method for antigenic typing. Continual use of live vaccines complicates diagnosis since no simple diagnostic tool can differentiate a field from a vaccine strain. Nucleotide sequencing of the S1 glycoprotein is the only method to discriminate between all IBV strains. Serology is also complicated by continual use of live vaccines. For surveillance purposes, ELISA is the method of choice, regardless of the antigenic type of IBV involved. The assay is used to monitor the response to vaccination, but field challenge can only be detected if flock antibody status is monitored continually. The antigenic type of a challenge strain involved cannot be ascertained by ELISA.

Animals↗