Nong lam University, Ho Chi Minh City

Faculty of Animal Science and Veterinary Medicine

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, VietnamChulalongkorn University, Thailand

Faculty of Animal Science and Veterinary Medicine – NLU - HCMC

Content

• ORT infection• ORT - Isolation• ORT - Identification• Further study

INTRODUCTION

Faculty of Animal Science and Veterinary Medicine – NLU - HCMC

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)

Faculty of Animal Science and Veterinary Medicine – NLU - HCMC

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

Faculty of Animal Science and Veterinary Medicine – NLU - HCMC

1981Turkey

1987duck

1991chicken

1993Characterized

• Isolation• Antibiotic susceptibility• Pathogenic characteristics• Vietnam: no publication on isolation ORT

THE STUDY

Faculty of Animal Science and Veterinary Medicine – NLU - HCMC

Samplesswab (30), trachea/ lungs (75)

Chicken: difficult breathing , sneezing , facial edema

DNA extraction Columbia + 5% sheep blood+ gentamicin 5 µg/ml

Gram stain (-), mobility (-), oxidase (+), catalase (-), MacConkey (-)

PCR amplification pinpoint small-colony, grey, greydish white, non-heamolysis16S rRNA ORT

Nucleotide sequencing

Faculty of Animal Science and Veterinary Medicine – NLU - HCMC

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%)

30

75

Samples

SwabLung/ trachea

Asadpour et al (2008), 2.3%Turan và Ak (2002), 11,46%

Faculty of Animal Science and Veterinary Medicine – NLU - HCMC

RESULT: isolationSamples from No (swab No.) Pos. Rate (%)

Breeding hens 5 (3) 1 20Layer 8 (5) 0 0

Broiler 21 (2) 2 9,5Native chicken 40 (17) 0 0

Local breed 18 (3) 1 5,5Black-Bone Silky Fowl 13 (0) 1 7,7

Total 105 (30) 5 4,7

Seifi, 2002 (2,6%); Turkyilmaz, 2005 (1,2%), Ghanbarpour and Salehi, 2009 (3,5%)

Faculty of Animal Science and Veterinary Medicine – NLU - HCMC

RESULT: PCR

. PCR product was nucleotide-sequenced and compared with

reference gene in GenBank (KC454288).

. The homology of 100% found (several ORT sequences)

1. Positive control (ORT, N. Chansiripornchai, 2007)

2. Negative control (E.coli)3. - 6: positive samples7. negative sample

Faculty of Animal Science and Veterinary Medicine – NLU - HCMC

RESULT: PCR (direct from samples)Samples from No Pos. Rate (%)

Breeding hens 5 (3) 1 20Layer 8 (5) 0 0

Broiler 21 (2) 2 9,5Native chicken 40 (17) 1 0

Local breed 18 (3) 2 5,5Black-Bone Silky Fowl 13 (0) 1 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%

Faculty of Animal Science and Veterinary Medicine – NLU - HCMC

Conclusion

• First study on isolation and identification of ORT in Vietnam.

• Further study on antimicrobial susceptibility, pathogenic characteristics

Faculty of Animal Science and Veterinary Medicine – NLU - HCMC

Thank you for your attention!