Comparison of On-farm Culture and Conventional Culture Methods in the Diagnosis of Mastitis in Cattle

Abstract

This study aimed to compare the microbiological culture and farm-culture methods used to diagnose mastitis in cattle. In the study, blood agar, MacConkey agar, Edwards medium, and Columbia CNA (CNA) agar were inoculated in the media. The samples were incubated under appropriate conditions to perform microbiological analysis of milk samples. While bacterial isolation and identification were performed in 90.3% (75) of the milk samples examined in the study, it was determined that there was no growth in 9.7% (8) of the milk samples, and two different types of pathogens were found in some samples. In blood agar isolation, 17.9% (n: 14) of the culture positive samples were Staphylococcus aureus, 61.6% (n: 48) Streptococcus spp, E. coli in 7.7% (n: 6), Enterococcus spp. in 6.4% (n: 5), yeast in 3.8% (n: 3) and coagulase-negative staphylococci (CNS) in 2.6% (n: 2). E. coli was detected in 60% (n: 6), Enterococcus spp. in 30% (n: 3), and yeast in 10% (n: 1) of the inoculations on MacConkey agar. Streptococcus spp. was observed in 91.5% (n: 43), Enterococcus spp. in 6.4% (n: 3), and yeast and CNS in 2.1% (n: 1) of the cultures of Edwards medium. S. aureus was isolated from 20% (n: 14), Streptococcus spp. from 68.6% (n: 48), Enterococcus spp. from 4.3% (n: 3), yeast from 4.3% (n: 3) and CNS from 2.8% (n: 2) of CNA inoculations. As a result, it was concluded that the on-farm culture method for detecting some aerobic bacterial pathogens in mastitic cow milk gave similar results to the conventional culture method. This method can be used in infection prevention and control programs.

Keywords

• Antibiogram
• On-farm culture
• Cow
• Mastitis

Sığırlarda Mastitisin Teşhisinde Mikrobiyolojik Kültür ve Çiftlikte-Kültür Yöntemlerinin Karşılaştırılması

Abstract

Bu çalışmada, bakteriyel nedenli inek mastitislerinin teşhisinde çiftlikte kültür ve konvansiyonel kültür yöntemlerinin karşılaştırılması, izole edilen bakterilerin bazı anibiyotiklere in vitro duyarlıklıklarının ortaya konulması amaçlandı. Çalışmada kanlı agar, MacConkey agar, Edwards medium ve Columbia CNA agar (CNA) kullanıldı. İn vitro antibiyotik duyarlılığının belirlenmesinde neomisin/basitrasin/tetrasiklin (30 μg/10 IU/30), sefapirin (30 μg), amoksisilin/klavulanik asit (2/1) (30 μg), penisilin (10 IU), tetrasiklin (30 μg), klindamisin (2 μg), kanamisin (30 μg) ve sefalotin (30 μg) diskleri kullanıldı. Konvansiyonal izolasyon ve identifikasyon yöntemlerinin uygulandığı çalışmada, örneklerin %90,3’ünden kültür pozitif sonuç alınırken, %9,7’sinde ise herhangi bir aerobik bakteri üremedi. Kanlı agarda yapılan izolasyonda kültür pozitif örneklerinin %17,9’unda (n: 14) Staphylococcus aureus, %61,6’sında (n: 48) Streptococcus spp., %7,7’sinde (n: 6) E. coli, %6,4’ünde (n: 5) Enterococcus spp., %3,8’inde (n: 3) maya ve %2,6’sında (n: 2) koagulaz negatif stafilokok (KNS) saptandı. MacConkey agara yapılan ekimlerin %60’ında (n: 6) E. coli, %30’unda (n: 3) Enterococcus spp. ve %10’unda (n: 1) maya belirlendi. Edwards mediuma ait ekimlerin %91,5’inde (n: 43) Streptococcus spp., %6,4’ünde (n: 3) Enterococcus spp., %2,1’inden (n: 1) maya ve KNS görüldü. CNA’a yapılan ekimlerin %20’sinden (n: 14) S. aureus, %68,6’sından (n: 48) Streptococcus spp., %4,3’ünden (n: 3) Enterococcus spp., %4,3’ünden (n: 3) maya ve %2,8’inden (n: 2) ise KNS izole edildi. Sonuç olarak, mastitisli inek sütlerinde bazı aerobik bakteriyel patojenlerin saptanmasına yönelik uygulanan çiftlikte kültür yönteminin, konvansiyonel kültür yöntemine benzer sonuçlar verdiği ve bu yöntemin enfeksiyona yönelik koruma ve kontrol programlarında kullanılabileceği kanısına varıldı.

Keywords

• Sığır
• bakteri
• mastitis
• izolasyon

References

1. 1. Atasever S, Erdem H. (2008): Süt sığırlarında mastitis ile sütün elektriksel iletkenliği arasındaki ilişkiler. Anadolu Journal of Agricultural Sciences, 23 (2), 131-136.
2. 2. Barrett DJ, Healy AM, Leonard FC, Doherty ML. (2005): Prevalence of pathogens causing subclinical mastitis in 15 dairy herds in the R.I. Irish Veterinary Journal, 58, 333-337.
3. 3. Cheng WN, & Han SG. (2020): Bovine mastitis: risk factors, therapeutic strategies, and alternative treatments - A review. Asian-Australasian journal of animal sciences, 33(11), 1699–1713.
4. 4. CLSI. (2017): Performance Standards for Antimicrobial Susceptibility Testing. 27th ed. CLSI supplement M100. Wayne, PA: Clinical and Laboratory Standards Institute.
5. 5. Çelik Ö, Sur E. & Çetin H. (2021): Aydın İli Söke İlçesinde Sütçü İneklerde Subklinik Mastitis Prevalansının ve Mastitise Neden Olan Aerobik Bakterilerin Belirlenmesi . Harran Üniversitesi Veteriner Fakültesi Dergisi, 10 (2), 100-106.
6. 6. Çokal Y, Konuş R. (2012): Subklinik mastitisli ineklerin sütlerinden aerobik bakterilerin izolasyonu. Balıkesir Sağlık Bilimleri Dergisi, 1(2), 65-69.
7. 7. Dufour S, Labrie J, Jacques M. (2019): The Mastitis Pathogens Culture Collection. Microbiol Resour Announc. 8:e00133-19.
8. 8. Elizabeth A, McInnis EA, Kalanetra KM, Mills DA, Maga EA (2015): Analysis of raw goat milk microbiota: impact of stage of lactation and lysozyme on microbial diversity. Food Microbiol. 46, 121-131.

9. 9. Erskine RJ, Walker RD, Bolin CA, Bartlett PC, White DG. (2002): Trends in antibacterial susceptibility of mastitis pathogens during a seven-year period. Journal of DairyScience. 85, 1111-1118.
10. 10. Ferreira JP, Anderson KL, Correa MT, Lyman R, Ruffin F, Reller LB, Fowler VG. (2011): Transmission of MRSA between companion animals and infected human patients presenting to outpatient medical care facilities. PLoS ONE, 6,:1-6.
11. 11. Ferreira JC, Gomes MS, Bonsaglia EC, Canisso IF, Garrett EF, Stewart JL, ... & Lima FS. (2018): Comparative analysis of four commercial on-farm culture methods to identify bacteria associated with clinical mastitis in dairy cattle. PLoS One, 13(3), e0194211.
12. 12. Fox LK. (2009): Prevelance, incidence and risk factors of heifer mastitis. Veteriner Microbiology, 134, 82-88.
13. 13. Hiitiö H, Riva R, Autio T, Pohjanvirta T, Holopainen J, Pyörälä SPS. (2015): Performance of a real-time PCR assay in routine bovine mastitis diagnostics compared with in-depth conventional culture. Journal of Dairy Research, 82, 200–8.
14. 14. Jones GM, Bailey TL. (2009): Understanding the basics of mastitis. Virginia Cooperative Extension.1-5. Erişim: https://vtechworks.lib.vt.edu/bitstream/handle/10919/48392/404-233_pdf.pdf
15. 15. Kaya O, Kırkan Ş, Gülal M, Ünal B. (1999): Aydın yöresinde ineklerde klinik mastitise neden olan mikroorganizmaların saptanması ve bunların antibiyotiklere duyarlılıklarının incelenmesi. Pendik Veteriner Mikrobiyoloji Dergisi, 30, 25-29.
16. 16. Sharun K, Dhama K, Tiwari R, Gugjoo MB, Iqbal Yatoo M, Patel SK ... & Chaicumpa W. (2021): Advances in therapeutic and managemental approaches of bovine mastitis: a comprehensive review. Veterinary Quarterly, 41(1), 107-136.
17. 17. Lago A, Godden SM, Bey R, Ruegg PL, Leslie K. (2011): The selective treatment of clinical mastitis based on on-farm culture results: I. Effects on antibiotic use, milk with holding time, and short-term clinical and bacteriological outcomes. Journal of Dairy Science, 94, 4441–56,
18. 18. Lago A, and Godden SM. (2018). Use of rapid culture systems to guide clinical mastitis treatment decisions. Vet Clin North Am Food Anim Pract, 34, 389–412.
19. 19. Macun HC, Yağcı, IP, Ünal N, Kalender H, Sakarya F, Yıldırım M. (2011): Kırıkkale’de belirlenen subklinik mastitisli ineklerde etken izolasyonu ve antibiyotik direnç durumu. Erciyes Üniversitesi Veteriner Fakültesi Dergisi, 8(2), 91-95.
20. 20. McCarron JL, Keefe GP, McKenna SL, Dohoo IR, Poole DE. (2009): Evaluation of the University of Minnesota Tri-plate and 3M Petrifilm for the isolation of Staphylococcus aureus and Streptococcus species from clinically mastitic milk samples. Journal of Dairy Science, 92, 5326–33.
21. 21. McDougall SJ, Niethammer and Graham EM. (2018): Antimicrobial usage and risk of retreatment for mild to moderate clinical mastitis cases on dairy farms following on-farm bacterial culture and selective therapy. N. Z. Vet. J. 66, 98–107.
22. 22. Neeser NL, Hueston WD, Godden SM, Bey RF. (2006): Evaluation of the use of an on farm system for bacteriologic culture of milk from cows with low-grade mastitis. Journal of the American Veterinary Medical Association, 228, 254–60.
23. 23. NMC. (2022): National Mastitis Council. Erişim Adresi: https://www.nmconline.org/nmc-protocols-guidelines-and-procedures/ (Erişim:1.05.2022).
24. 24. Özdemir M. (2005): Mastitisli inek sütlerinden Staphylococcus türlerinin izolasyonu ve identifikasyonu. Pendik Veteriner Kontrol ve Araştırma Enstitüsü, İstanbul 2005.
25. 25. Pinzón-Sánchez C, Cabrera VE, Ruegg PL. (2011): Decision tree analysis of treatment strategies for mild and moderate cases of clinical mastitis occurring in early lactation. Journal of Dairy Science, 94, 1873–92.
26. 26. Stuhr T, Aulrich K. (2010): Intramammary infections in dairy goats: recent knowledge and indicators for detection of subclinical mastitis. Landbauforsch, 4, 267-280.
27. 27. Tel OY, Keskin O, Zonturlu AK, & Arserim Kaya NB. (2009): Şanlıurfa yöresinde subklinik mastitislerin görülme oranı, aerobik bakteri izolasyonu ve duyarlı antibiyotiklerin belirlenmesi. F Ü Sağ Bil Vet Derg, 23(2), 101-106.
28. 28. Tenhagen BA, Koster G, Wallmann J, Heuwieser W. (2006): Prevalence of mastitis pathogens and their resistance against antimicrobial agents in dairy cows in Brandenburg, Germany, Journal of Dairy Science, 89, 2542-2551.
29. 29. Vasquez AK, Nydam DV, Capel MB, Eicker S, Virkler PD. (2017): Clinical outcome comparison of immediate blanket treatment versus a delayed pathogen-based treatment protocol for clinical mastitis in a New York dairy herd. J. Dairy Sci. 100, 2992–3003.
30. 30. Verbeke J, Piepers S, Supré K, De Vliegher S. (2014): Pathogen-specific incidence rate of clinical mastitis in Flemish dairy herds, severity, and association with herd hygiene. J. Dairy Sci. 97, 6926–6934.