ISOLATION AND IDENTIFICATION ORNITHOBACTERIUM RHINOTRACHEALE FROM CHICKEN IN VIETNAM Nguyen Thi Bich Lien1, Vo Thi Tra An1, Tran Thi Ngoc Han1, Ho Hoang Dung1 and Niwat Chansiripornchai2 Nong Lam University, Vietnam Chulalongkorn University, Thailand

Content ORT infection ORT - Isolation ORT - Identification Further study

Introduction

Ornithobacterium infection Ornithobacterium rhinotracheale (ORT) Symptoms: sneezing, moist eyes, swollen heads, difficult breathing, mortality (2-10%) Lesions: airsacculitis, tracheitis, pneumonia Risk factors: virus (ND, AI, IBD, IB), ventilation (NH3, CO2) Transmission: horizontal (aerosols, direct contact, drinkers), vertical (egg shell)

Ornithobacterium rhinotracheale Gram-negative, rod shaped bacterium, pleomorphic Slow growing, easily overgrown by less fastidious growing bacteria (gentamicin supp.) For optimal growth 5–10% sheep blood agar For 48 hours at 37°C Microaerophilic conditions (5-10% CO2) Small circular grey to grey-white colonies

Antibiotic susceptibility Pathogenic characteristics 1981 Turkey 1987 duck 1991 chicken 1993 Characterized Isolation Antibiotic susceptibility Pathogenic characteristics Vietnam: no publication on isolation ORT

The study

swab (30), trachea/ lungs (75) Chicken: difficult breathing , sneezing , facial edema Samples swab (30), trachea/ lungs (75) DNA extraction Columbia + 5% sheep blood + gentamicin 5 µg/ml PCR amplification pinpoint small-colony, grey, greydish white, non-heamolysis 16S rRNA ORT Gram stain (-), mobility (-), oxidase (+), catalase (-), MacConkey (-) Nucleotide sequencing

Results Generally, positive 4.7% No swab samples found carrying ORT 5 isolates cultured from lung/ tracheal samples (6.6%) 7 samples were positive by PCR assay (6.7%) (9.3%) Asadpour et al (2008), 2.3% Turan và Ak (2002), 11,46%

RESULT: isolation Samples from No (swab No.) Pos. Rate (%) Breeding hens 5 (3) 1 20 Layer 8 (5) Broiler 21 (2) 2 9,5 Native chicken 40 (17) Local breed 18 (3) 5,5 Black-Bone Silky Fowl  13 (0) 7,7 Total 105 (30) 5 4,7 Seifi, 2002 (2,6%); Turkyilmaz, 2005 (1,2%), Ghanbarpour and Salehi, 2009 (3,5%)

RESULT: PCR Positive control (ORT, N. Chansiripornchai, 2007) Negative control (E.coli) - 6: positive samples 7. negative sample . PCR product was nucleotide-sequenced and compared with reference gene in GenBank (KC454288). . The homology of 100% found (several ORT sequences)

RESULT: PCR (direct from samples) Samples from No Pos. Rate (%) Breeding hens 5 (3) 1 20 Layer 8 (5) Broiler 21 (2) 2 9,5 Native chicken 40 (17) Local breed 18 (3) 5,5 Black-Bone Silky Fowl  13 (0) 7,7 Total 105 (30) 7 4,7 Asadpour et al (2008): ELISA, 62,8%; PCR, 3,1%; Ozbey et al (2004): ELISA 10,2%; PCR, 2,4%

Conclusion First study on isolation and identification of ORT in Vietnam. Further study on antimicrobial susceptibility, pathogenic characteristics

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