Kannan Ganapathy

Pathogenicity of Mycoplasma gallisepticum and Mycoplasma imitans in red-legged partridges (Alectoris rufa).

Groups of 3-day-oId red-legged partridges were infected intranasally either with the S6 strain of M. gallisepticum or with an M. imitans strain from a partridge with sinusitis. Starting 6-8 days post-infection (p.i.) birds in both groups developed signs of depression, nasal exudation, tracheal rales, sneezing, gasping, head shaking, watery eyes and eye scratching. The most outstanding feature was bilateral swelling of the infraorbital sinuses. Morbidity reached 100% in the M. gallisepticum infection and 80% in the M. imitans infection and mean clinical scores in the former were significantly greater than those of the latter group on days 11 and 14 p.i. There was also slower recovery in the M. gallisepticum infection. Necropsies at weekly intervals for 5 weeks revealed nasal and sinus exudate in both groups but tracheal exudate and cloudy airsacs were seen only in M. gallisepticum infection. M. gallisepticum was isolated from both upper and lower respiratory tract throughout the experiment while M. imitans was recovered less frequently from the upper respiratory tract and from the lungs and air sacs only at 7 days p.i. The numbers of isolations from eyes, tracheas, lungs and thoracic air sacs of the M. gallisepticum group were significantly greater than those from the M. imitans group. Seroconversion occurred in both groups using homologous antigen.

Detection of variant infectious bronchitis viruses in broiler flocks in Libya

Abstract A number of broiler flocks with respiratory disease and high mortality in five broiler farms in Libya were sampled for detection of infectious bronchitis virus (IBV). Twelve IBV strains from these farms were detected by reverse transcription polymerase chain reaction (RT-PCR) and differentiated by nucleotide sequencing of the hypervariable region of the S1 gene. A pair-wise comparison of the sequences showed two distinctive patterns. Those from farms 1, 2, 4 and 5, formed a separate cluster with 94–99% relatedness to the Egyptian IBV strains CK/Eg/BSU-2/2011, CK/Eg/BSU-3/2011 and Eg/1212B. Sequences from the farm 3 formed another cluster with 100% relatedness to Eg/CLEVB-2/IBV/012 and IS/1494/06. This appears to be the first report on the co-circulation these variant IBVs in Libya.

Effects of cyclosporin A on the immune responses and pathogenesis of a virulent strain of Mycoplasma gallisepticum in chickens.

Immune responses to the virulent S6 strain of Mycoplasma gallisepticum in immunocompetent and cyclosporin A (CsA)-treated specific pathogen free chickens were investigated, and pathogenesis of the M. gallisepticum strain was also examined. Ten-day-old specific pathogen free chickens were inoculated by eye-drop with M. gallisepticum, and a control uninfected group was inoculated with mycoplasma broth. Blood was collected weekly for 4 weeks from five birds in each group and whole blood lymphocyte transformation assayed against concanavalin A and lipopolysaccharide. Blood samples were also collected at intervals for serological assays. Live body weight, clinical signs and lesions were monitored, and recovery of M. gallisepticum was attempted from choanal cleft of live birds and also from various sites at necropsy. In parallel to the aforementioned groups, another set of two groups of chicks treated with CsA was infected with M. gallisepticum S6 or mycoplasma broth. These groups were subjected to the same experimental procedures. In the immunocompetent chickens, M. gallisepticum caused temporary T-cell suppression at 2 weeks post-infection. Comparison of the clinical signs and macroscopic lesions produced in immunocompetent and CsA-treated chickens indicated that T cells may not play an active role in disease development. The percentage of birds with mycoplasma isolation and the load of mycoplasmas suggested that T cells may have some role in resisting mycoplasma colonization or in the elimination of the infection.

Genotypes of infectious bronchitis viruses circulating in the Middle East between 2009 and 2014.

We are reporting on the infectious bronchitis virus (IBV) genotypes circulating within seven Middle East countries and the alterations in genotype distributions between 2009 and 2014. Tissue samples on FTA cards were received over the six-year period. Viral RNA was extracted using phenol chloroform and subjected to nested RT-PCR targeting a 393 bp region of the S1 gene before being followed by sequencing. From the 461 submitted samples, 363 were IBV positive by RT-PCR (77.01%). Of these, 355 (97.80%) gave sequences that can be genotyped. They belonged to six genotypes; 793B (43.66%), IS/1494/06 (18.31%), Massachusetts (Mass) (12.96%), IS/885/00 (11.27%), Q1 (11.27%) and D274 (2.25%). The prominence of 793B is not surprising, given that 793B vaccine strains are widely used in the Middle East. Sequence analysis demonstrated that the majority of 793B (67.13%) and Mass (81.13%) strains were closely related to vaccine strains based on 99-100% homology with the partial-S1 gene. Vaccinal strains belonging to the D274 genotype were present but only at a low level. Variable proportions of 793B, Mass, D274, IS/1494/06, IS/885/00 and Q1 field strains were identified in different countries. After 2012, the 793B field strain showed distinct clustering compared to strains from earlier years. Translated amino acid alterations were minimal but still may have played an important role in the persistence of this virus despite the use of live 793B vaccines. Huge challenges for an efficient protection against virulent IBVs and chicken production are posed by co-circulating793B, Mass and D274 viruses with less than 99% homology to the respective vaccine strains, along with the recently emerged variant IBVs, despite active IBV vaccination strategies in the Middle East, continuous surveillance of IBV genotypes is essential in formulating optimal control strategies, including the choice and development of new vaccine strains and formulation of vaccination programmes.

Survey of campylobacter, salmonella and mycoplasmas in house crows (Corvus splendens) in Malaysia

House crows (Corvus splendens) in Selangor, Malaysia were examined for the presence of Campylobacter species, Salmonella species, Mycoplasma gallisepticum and Mycoplasma synoviae by serology, culture and pcr. For the detection of Campylobacter and Salmonella species swabs were taken either from the intestine or cloaca. For the detection of mycoplasmas, swabs were taken either from the choanal cleft or trachea for culture and pcr and serum samples were tested by the rapid serum agglutination (rsa) and monoclonal antibody-blocking elisa (mbelisa) for antibodies to M gallisepticum and M synoviae. For campylobacter, 25·3 per cent of the crows were positive by culture, and the species identified were Campylobacter jejuni and Campylobacter coli. No Salmonella species were isolated. Four of 24 swabs were positive for M gallisepticum dna but none gave positive results for M synoviae dna. No M gallisepticum or M synoviae antibodies were detected by rsa but 60 per cent of the sera gave positive reactions for M gallisepticum and 13 per cent gave positive reactions for M synoviae by mbelisa.

Pathogenicity of in vivo-passaged Mycoplasma imitans in turkey poults in single infection and in dual infection with rhinotracheitis virus

Mycoplasma imitans (Mim) is a close relative of Mycoplasma gallisepticum (Mg), but has been isolated from ducks, geese and partridge. In order to investigate its potential pathogenicity for turkeys a UK isolate of Mim from a partridge with sinusitis was first passaged through turkey poults and then assessed for pathogenicity in turkey embryo tracheal organ cultures (TOCs) and in one-day-old turkey poults with or without turkey rhinotracheitis virus (TRTV). Mim appeared to gain virulence on passage through poults and this was confirmed by an increased ciliostatic effect in TOCs. In single infection in poults the organism caused mild upper respiratory disease but in dual infection with TRTV there was a significant increase in clinical signs and lesions. The mycoplasma was only isolated from upper respiratory tract in single infection but was recovered also from lung and airsacs in the presence of the virus. There was also a higher humoral immune response in the dual infection than the single Mim infection and there were some cross-reactions with commercial Mg stained antigen in the rapid serum agglutination test.

Molecular detection of infectious bronchitis and avian metapneumoviruses in Oman backyard poultry

Abstract Infectious bronchitis virus (IBV) and avian metapneumovirus (aMPV) are economically important viral pathogens infecting chickens globally. Identification of endemic IBV and aMPV strains promotes better control of both diseases and prevents production losses. Orophrayngeal swab samples were taken from 2317 birds within 243 different backyard flocks in Oman. Swabs from each flock were examined by RT-PCR using part-S1 and G gene primers for IBV and aMPV respectively. Thirty-nine chicken flocks were positive for IBV. Thirty two of these were genotyped and they were closely related to 793/B, M41, D274, IS/1494/06 and IS/885/00. 793/B-like IBV was also found in one turkey and one duck flock. Five flocks were positive for aMPV subtype B. Though no disease was witnessed at the time of sampling, identified viruses including variant IBV strains, may still pose a threat for both backyard and commercial poultry in Oman.

Evaluation of Flinders Technology Associates cards for storage and molecular detection of avian metapneumoviruses.

The feasibility of using Flinders Technology Associates (FTA) cards for the molecular detection of avian metapneumovirus (aMPV) by reverse transcriptase-polymerase chain reaction (RT-PCR) was investigated. Findings showed that no virus isolation was possible from aMPV-inoculated FTA cards, confirming viral inactivation upon contact with the cards. The detection limits of aMPV from the FTA card and tracheal organ culture medium were 101.5 median ciliostatic doses/ml and 100.75 median ciliostatic doses/ml respectively. It was possible to perform molecular characterization of both subtypes A and B aMPV using inoculated FTA cards stored for up to 60 days at 4 to 6°C. Tissues of the turbinate, trachea and lung of aMPV-infected chicks sampled either by direct impression smears or by inoculation of the tissue homogenate supernatants onto the FTA cards were positive by RT-PCR. However, the latter yielded more detections. FTA cards are suitable for collecting and transporting aMPV-positive samples, providing a reliable and hazard-free source of RNA for molecular characterization.

QX-like infectious bronchitis virus isolated from proventriculitis in commercial broilers in England

This short communication reports on the isolation of QX-like infectious bronchitis virus (IBV) from the proventriculus of broiler chicken in England, and its pathogenesis in specific-pathogen-free (SPF) chicks. This appears to be the first report of the isolation of this virus from the proventriculus of affected chicks in the UK. The virus was first recovered from this tissue in chickens in China in 1996 (YuDong and others 1998), and later, more cases were reported, including in flocks vaccinated against IBV (Yu and others 2001). We received samples of kidney and proventriculus from a flock of 56-days-old commercial broiler chickens, where the submitting field veterinarian reported respiratory signs, increased levels of mortality, a poor feed conversion ratio and suboptimal live body weight gain. At necropsy, proventriculitis and swollen kidneys were reported. Similar tissues were pooled and processed for IBV detection by reverse-transcriptase PCRs (RT-PCR), as previously described (Worthington and others 2008), and virus isolation (VI) were attempted. RT-PCR tests on the supernatant of the pooled tissues of kidneys were negative for IBV, while the proventriculus samples were positive. The proventriculus was negative for avian metapneumovirus (aMPV), Newcastle disease virus (NDV) and …

Heterologous live infectious bronchitis virus vaccination in day-old commercial broiler chicks: clinical signs, ciliary health, immune responses and protection against variant infectious bronchitis viruses

ABSTRACT Groups of one-day-old broiler chicks were vaccinated via the oculo-nasal route with different live infectious bronchitis virus (IBV) vaccines: Massachusetts (Mass), 793B, D274 or Arkansas (Ark). Clinical signs and gross lesions were evaluated. Five chicks from each group were humanely killed at intervals and their tracheas collected for ciliary activity assessment and for the detection of CD4+, CD8+ and IgA-bearing B cells by immunohistochemistry (IHC). Blood samples were collected at intervals for the detection of anti-IBV antibodies. At 21 days post-vaccination (dpv), protection conferred by different vaccination regimes against virulent M41, QX and 793B was assessed. All vaccination programmes were able to induce high levels of CD4+, CD8+ and IgA-bearing B cells in the trachea. Significantly higher levels of CD4+ and CD8+ expression were observed in the Mass2 + 793B2-vaccinated group compared to the other groups (subscripts indicate different manufacturers). Protection studies showed that the group of chicks vaccinated with Mass2 + 793B2 produced 92% ciliary protection against QX challenge; compared to 53%, 68% and 73% ciliary protection against the same challenge virus by Mass1 + D274, Mass1 + 793B1 and Mass3 + Ark, respectively. All vaccination programmes produced more than 85% ciliary protection against M41 and 793B challenges. It appears that the variable levels of protection provided by different heterologous live IBV vaccinations are dependent on the levels of local tracheal immunity induced by the respective vaccine combination. The Mass2 + 793B2 group showed the worst clinical signs, higher mortality and severe lesions following vaccination, but had the highest tracheal immune responses and demonstrated the best protection against all three challenge viruses.