Investigation of the Effectiveness of Chromogenic Media in the Isolation of Pasteurella multocida and Mannheimia haemolytica from Calf Nasal Swab Samples

Öz

Accurate and rapid diagnosis of disease agents is the most important step in terms of control
practices. Chromogenic media are culture media that allow the formation of colonies in colors
specific to target microorganisms. Because they are target-specific, they do not require validation
of results but provide ease of use and time savings. In this study, it was aimed to evaluate
the effectiveness of a chromogenic medium (Pasteurella BDR kit) in the detection of Pasteurella
multocida and Mannheimia haemolytica agents in the Pasteurelleceae family, which cause respiratory
disease in cattle. In this study, nasal swab samples taken from calves showing symptoms
of respiratory disease were cultured in chromogenic and standard media. Suspicious growing
colonies were confirmed by polymerase chain reaction for P. multocida and M. haemolytica. While
31 (36.9%) samples formed colonies with the chromogenic medium in the color specific to the
target bacteria, 28 (33.3%) samples were positively determined for the target bacteria using the
standard cultural method. The results of 26 samples were positive by both cultural diagnosis
methods. When the results were compared with the traditional cultural diagnosis, agreement
was found to be 92.86%. All colored colonies grown on the chromogenic medium were also tested
by polymerase chain reaction (PCR). It was determined that the chromogenic medium detected P.
multocida at a rate of 92.86% (n = 26) and M. haemolytica at a rate of 100% (n = 2) by forming colonies
with a family-specific color. As a result, it was concluded that the use of chromogenic media
is beneficial in the practical, rapid, and high-accuracy diagnosis of target agents.

Anahtar Kelimeler

• Kromojenik besiyeri
• Pasteurella
• sığır solunum hastalığı
• kültür

Buzağı Nasal Svap Örneklerinden Pasteurella multocida ve Mannheimia haemolytica İzolasyonunda Kromojenik Besiyerinin Etkinliğinin Araştırılması

Öz

Hastalık etkenlerinin doğru ve hızlı teşhisi, kontrol uygulamaları açısından en önemli basamaktır. Kromojenik besiyerleri hedef mikroorganizmalara özgü renkte kolonilerin meydana gelmesini sağlayan kültür ortamlarıdır. Bunlar hedefe özgü olduklarından sonuçların doğrulama ihtiyacı duymamakla birlikte, kullanım kolaylığı ve zaman tasarrufu sağlarlar. Bu çalışmada, sığırlarda solumun hastalığına neden olan Pasteurelleceae ailesinde bulunan Pasteurella multocida ve Mannheimia haemolytica etkenlerinin tespitinde, kromojenik besiyerinin (Pasteurella BRD Kit) etkinliğini değerlendirmek amaçlandı. Çalışmada solunum hastalığı belirtisi gösteren buzağılardan alınan nasal svap örnekleri kromojenik ve standart besiyerlerinde kültüre edildi. Üreyen şüpheli koloniler Pasteurella multocida ve Mannheimia haemolytica yönünden Polimeraz Zincir Reaksiyonu (PCR) ile doğrulandı. Kromojenik besiyeri ile 31 (%36,9) numune, hedef bakterilere özgü renkte koloni oluştururken, standard kültürel yöntem ile 28 (%33,3) numune hedef bakteriler yönünden pozitif olarak belirlendi. Her iki kültürel tanı yöntemiyle 26 numunenin sonuçları pozitif olarak tespit edildi. Sonuçlar geleneksel kültürel tanı ile karşılaştırıldığında %92,86 oranında uyumlu sonuç bulundu. Kromojenik besiyerinde üreyen renkli kolonilerin tamamı PCR ile de test edildi. Kromojenik besiyeri P. multocida’yı %92,86 (n:26), M. haemolytica’yı %100 (n:2) oranında aileye özgü renkte koloni meydana getirme suretiyle tespit ettiği belirlendi. Sonuç olarak, hedef etkenin pratik, hızlı ve yüksek doğrulukla teşhisinde kromojenik besiyerinin kullanımının yararlı olduğu kanısına varıldı.

Anahtar Kelimeler

• Kromojenik besiyeri
• Pasteurella
• sığır solunum hastalığı
• kültür

Kaynakça

1. 1. Peel DS. The effect of market forces on bovine respiratory disease. Vet Clin North Am Food Anim Pract. 2020;36(2):497-508. [CrossRef]
2. 2. Kudirkiene E, Aagaard AK, Schmidt LMB, Pansri P, Krogh KM, Olsen JE. Occurrence of major and minor pathogens in calves diagnosed with bovine respiratory disease. Vet Microbiol. 2021;259:109135. [CrossRef]
3. 3. Gorden PJ, Plummer P. Control, management, and prevention of bovine respiratory disease in dairy calves and cows. Vet Clin North Am Food Anim Pract. 2010;26(2):243-259. [CrossRef]
4. 4. Pratelli A, Cirone F, Capozza P, et al. Bovine respiratory disease in beef calves supported long transport stress: an epidemiological study and strategies for control and prevention. Res Vet Sci. 2021;135:450- 455. [CrossRef]
5. 5. Chernitskiy AE, Safonov VA. Early detection of bovine respiratory disease in calves by induced cough. In IOP Conference Series. IOP Conf Ser.: Earth Environ Sci. 2021;677(4). [CrossRef]
6. 6. Dorso L, Rouault M, Barbotin C, Chartier C, Assié S. Infectious bovine respiratory diseases in adult cattle: an extensive necropsic and etiological study. Animals (Basel). 2021;11(8):2280. [CrossRef]
7. 7. Zhang M, Hill JE, Alexander TW, Huang Y. The nasal viromes of cattle on arrival at Western Canadian feedlots and their relationship to development of bovine respiratory disease. Transbound Emerg Dis. 2021;68(4):2209-2218. [CrossRef]
8. 8. Bell CJ, Blackburn P, Elliott M, et al. Investigation of polymerase chain reaction assays to improve detection of bacterial involvement in bovine respiratory disease. J Vet Diagn Invest. 2014;26(5):631-634. [CrossRef]

9. 9. Gülaydın O, Gürtürk K. Identification of Pasteurella multocida strains isolated from respiratory tract of healthy and diseased cattle and determination of capsular types by PCR. Van Veterin J. 2018;29(3):143-146.
10. 10. McGuirk SM. Disease management of dairy calves and heifers. Vet Clin North Am Food Anim Pract. 2008;24(1):139-153. [CrossRef]
11. 11. Çelik M, Erdenliğ-Gürbilek S. Mannheimia haemolytica suşlarının farklı besiyerlerinde üreme ve lökotoksin oluşturma özelliklerinin incelenmesi. J Adv Vetbiosci Tech. 2020;5(2):33-42.
12. 12. Perry JD, Freydière AM. The application of chromogenic media in clinical microbiology. J Appl Microbiol. 2007;103(6):2046-2055. [CrossRef]
13. 13. Reissbrodt R. New chromogenic plating media for detection and enumeration of pathogenic Listeria spp.—an overview. Int J Food Microbiol. 2004;95(1):1-9. [CrossRef]
14. 14. Alizadeh N, Rezaee MA, Kafil HS, et al. Detection of carbapenemresistant Enterobacteriaceae by chromogenic screening media. J Microbiol Methods. 2018;153:40-44. [CrossRef]
15. 15. Rank EL. Chromogenic agar media in the clinical, food, and environmental testing arenas, part I. Clin Microbiol Newsl. 2012;34(6):43-47. [CrossRef]
16. 16. Perry JD. A decade of development of chromogenic culture media for clinical microbiology in an era of molecular diagnostics. Clin Microbiol Rev. 2017;30(2):449-479. [CrossRef]
17. 17. Garcia BLN, Fidelis CE, Freu G, Granja BM, Dos Santos MV. Evaluation of chromogenic culture media for rapid identification of Gram-positive bacteria causing mastitis. Front Vet Sci. 2021;8:662201. [CrossRef]
18. 18. Granja BM, Fidelis CE, Garcia BLN, Dos Santos MV. Evaluation of chromogenic culture media for rapid identification of microorganisms isolated from cows with clinical and subclinical mastitis. J Dairy Sci. 2021;104(8):9115-9129. [CrossRef]
19. 19. Payne M, Roscoe D. Evaluation of two chromogenic media for the isolation and identification of urinary tract pathogens. Eur J Clin Microbiol Infect Dis. 2015;34(2):303-308. [CrossRef]
20. 20. Pletinckx LJ, De Bleecker Y, Dewulf J, Rasschaert G, Goddeeris BM, De Man I. Evaluation of salt concentrations, chromogenic media and anatomical sampling sites for detection of methi cilli n-res istan t Staphylococcus aureus in pigs. Vet Microbiol. 2012;154(3-4):363- 368. [CrossRef]
21. 21. Quinn PJ, Markey BK, Leonard FC, Hartigan P, Fanning S, Fitzpatrick EI. Veterinary Microbiology and Microbial Disease. Chichester, UK: John Wiley & Sons; 2011.
22. 22. Ewers C, Lübke-Becker A, Bethe A, Kießling S, Filter M, Wieler LH. Virulence genotype of Pasteurella multocida strains isolated from different hosts with various disease status. Vet Microbiol. 2006;114(3- 4):304-317. [CrossRef]
23. 23. Deressa A, Asfaw Y, Lubke B, Kyule MW, Tefera G, Zessin KH. Molecular detection of Pasteurella multocida and Mannheimia haemolytica in sheep respiratory infections in Ethiopia. J Appl Res Vet Med. 2010;8(2):101.
24. 24. Thomas AC, Bailey M, Lee MRF, et al. Insights into Pasteurellaceae carriage dynamics in the nasal passages of healthy beef calves. Sci Rep. 2019;9(1):11943. [CrossRef]
25. 25. Bosch AA, Biesbroek G, Trzcinski K, Sanders EA, Bogaert D. Viral and bacterial interactions in the upper respiratory tract. PLoS Pathog. 2013;9(1):e1003057. [CrossRef]
26. 26. Pansri P, Katholm J, Krogh KM, et al. Evaluation of novel multiplex qPCR assays for diagnosis of pathogens associated with the bovine respiratory disease complex. Vet J. 2020;256:105425. [CrossRef]
27. 27. Cirone F, Padalino B, Tullio D, et al. Prevalence of pathogens related to bovine respiratory disease before and after transportation in beef steers: preliminary results. Animals (Basel). 2019;9(12):1093. [CrossRef]
28. 28. Cole SD, Swiderski M, Dietrich J, McGonigle KM. Comparison of a chromogenic urine culture plate system (UTid+) and conventional urine culture for canine and feline specimens. Vet Sci. 2022;9(3):138. [CrossRef]
29. 29. Al-Anbagi NA. Isolaton and ıdentıfıcatıon some bacterıal causes of lung abscessessheep by chromogenıc medıa. Basrah J Vet Res. 2016;15(2):360-370.
30. 30. Beehan DP, McKinnon AO. How to Diagnose Common Equine Reproductive Tract Bacterial Pathogens Using Chromogenic Agar. Proc Am Assoc Eq Pract. 2009;55:320-325.
31. 31. DeRosa DC, Mechor GD, Staats JJ, Chengappa MM, Shryock TR. Comparison of Pasteurella spp. simultaneously isolated from nasal and transtracheal swabs from cattle with clinical signs of bovine respiratory disease. J Clin Microbiol. 2000;38(1):327-332. [CrossRef]
32. 32. Abed AH, El-Seedy FR, Hassan HM, et al. Serotyping, genotyping and virulence genes characterization of Pasteurella multocida and Mannheimia haemolytica isolates recovered from pneumonic cattle calves in North Upper Egypt. Vet Sci. 2020;7(4):174. [CrossRef]
33. 33. Cengiz S, Adıgüzel MC, Dinç G. Detection of Pasteurella multocida, Mannhemia haemolytica, Histophilus somni and Mycoplasma bovis in cattle lung. Rev Mex Cienc Pecuarias. 2021;12(3):710-720. [CrossRef]
34. 34. Timsit E, Hallewell J, Booker C, Tison N, Amat S, Alexander TW. Prevalence and antimicrobial susceptibility of Mannheimia haemolytica, Pasteurella multocida, and Histophilus somni isolated from the lower respiratory tract of healthy feedlot cattle and those diagnosed with bovine respiratory disease. Vet Microbiol. 2017;208:118-125. [CrossRef]
35. 35. Jamali H, Rezagholipour M, Fallah S, et al. Prevalence, characterization and antibiotic resistance of Pasteurella multocida isolated from bovine respiratory infection. Vet J. 2014;202(2):381-383. [CrossRef]
36. 36. Tel OY, Keskin O. Koyun akciğerlerinden Pasteurella multocida ve Mannheimia haemolytica izolasyonu ve antibiyotiklere duyarlılığı. Yüzüncü Yıl Univ Vet Fak Derg. 2010;21(1):31-34.
37. 37. Christensen H, Kuhnert P, Norskov-Lauritsen N, Planet PJ, Bisgaard M. The family Pasteurellaceae. In: Rosenberg E, DeLong EF, Lory S, Stackebrandt E, Thompson F, eds. The Prokaryotes. 4th ed. Berlin, Germany: Springer; 2014:535-564. Vet Sci