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Söndürülen bruselloz salgınlarından sonra sığır çiftliklerinden alınan toprak-gübre karışımları ve hayvan yemi örneklerinde Brucella etkenlerinin araştırılması

Year 2025, Volume: 96 Issue: 2, 131 - 139, 15.06.2025

Ahmet Murat Saytekin , Ayfer Güllü Yücetepe , Songül Ötkün , Sevil Erdenlig Gürbilek

https://doi.org/10.33188/vetheder.1631029

Abstract

Bruselloz gelişmiş ülkelerde kontrol altına alınmış olsa da endemik bölgelerde hayvan ve insan sağlığını olumsuz etkilemektedir. Bu bölgelerdeki sığır çiftliklerinde çıkan hastalıklar resmi olarak söndürülmüş olsa da, çiftçiler söndürme işlemlerinden sonra çiftliklerinde bulunan bazı materyallerde bakterilerin mevcut olup olmadığını merak etmektedir. Çiftlik üretim faaliyetlerinin devam edebilmesi ve yeni hayvanların zamanında tedarik edilebilmesi için bu hassas durumun açığa kavuşturulması bir ihtiyaç haline gelmiştir. Bu çalışmada, söndürme işlemlerinden sonra hastalığı etkili bir şekilde bulaştırabilecek kritik materyallerde Brucella bakterilerinin varlığının araştırılması amaçlamıştır. Bu sebeple, bruselloz salgınlarından sonra hastalığın resmi olarak sonlandırıldığı üç sığır çiftliğinde toprak-gübre karışımları ve hayvan yemleri seçildi ve incelendi. Örnekler, bruselloz için söndürme prosedürleri tamamlandıktan yaklaşık yirmişer gün sonra, toprak-gübre karışımlarından 68, hayvan yemlerinden 55 adet toplandı. Brusella bakterilerinin teşhisi için klasik kültür yöntemleri bazı modifikasyonlarla kullanıldı. Bakteri kolonileri seçici besiyerlerinde üretildikten sonra, Brusella bakterileri için cins ve tür spesifik PCR testleriyle araştırıldı. Cins-spesifik PCR ile 5 pozitiflik elde edildi. Bunlardan sadece bir tanesi tür-spesifik multipleks PCR ile doğrulanabildi. Kalan dördünün, genetik olarak yakın bazı toprak bakterilerine ait olabilecekleri düşünüldü. Sonuç olarak, bruselloz söndürme prosedürlerinin uygun olduğu ve çiftliklerin bulaş açısından güvenli sayılabileceği kanaatine varıldı. Hastalık çıkışlarından sonra hastalığın yok edilmesine yönelik resmi kurallar tam olarak uygulansa da, daha güvenilir değerlendirmeler için daha fazla çiftlikte benzer çalışmalara ihtiyaç duyulmaktadır.

Keywords

B. abortus Kültür PCR Sığır Toprak

References

  • Manual of Diagnostic Tests and Vaccines for Terrestrial Animals (WOAH), Chapter 3.1.4. Brucellosis (Infection with B. abortus, B. melitensis and B. suis) [Internet]. 2024 Nov 29 [cited 2024 Dec 17]. Available from: https://www.woah.org/fileadmin/Home/eng/Health_standards/tahm/3.01.04_BRUCELLOSIS.pdf
  • O’Callaghan D. Human brucellosis: recent advances and future challenges. Infect Dis Poverty. 2020;9:1-2.
  • Cutler SJ, Whatmore AM, Commander AJ. Brucellosis, new aspects of an old disease. J Appl Microbiol. 2005;98:1270-81.
  • Refai M. Incidence and control of brucellosis in the Near East region. Vet Microbiol. 2002;90:81-110.
  • Taleski V, Zerva L, Kantardjiev T, Cvetnic Z, Erski-Biljic M, Nicolovski B, et al. An overview of the epidemiology and epizootology of brucellosis in selected countries of Central and Southeast Europe. Vet Microbiol. 2002;90(1-4):147-55.
  • Blasco JM. Control and eradication strategies for Brucella melitensis infection in sheep and goats. Prilozi. 2010;31(1):145-65.
  • Morales-Estradaa AI, Hernández-Castrob R, López-Merinoa A, Singh-Bedic J, Contreras-Rodrígueza A. Isolation, identification, and antimicrobial susceptibility of Brucella spp. cultured from cows and goat manure in Mexico. Arch Med Vet. 2016;48:231-5.
  • Aydın N. Gram negatif küçük çomaklar – Brucella infeksiyonları. In: Arda M, editor. Özel Mikrobiyoloji. Ankara: Medisan Yayınevi; 1998. p. 110–25.
  • Alton GG, Jones LM, Angus RD, Verger JM. Techniques for the brucellosis laboratory. 1st ed. Paris: Institut National de la Recherche Agronomique; 1998.
  • Hagan WA, Bruner DW, Timoney JF. The genus Brucella. In: Hagan WA, Bruner DW, Timoney JF, editors. Hagan and Bruner´s microbiology and infectious diseases of domestic animals. UK: Cornell University Press; 1988. p. 135-52.
  • U.S. Environmental Protection Agency (EPA). Persistence of Categories A and B Select Agents in Environmental Matrices-EPA/600/R-14/074 [Internet]. 2014 Jun 1 [cited 2024 Oct 17]. Available from: https://www.researchgate.net/publication/289245437_Persistence_of_Categories_A_and_B_Select_Agents_in_Environmental_Matrices
  • Farrell ID. The development of a new selective medium for the isolation of Brucella abortus from contaminated sources. Res J Vet Sci. 1974;16:280-6.
  • Saytekin AM, Adıgüzel A, Al-Kilani K, Güllü Yücetepe A, Keskin O. Some virulence genes and biofilm formation capabilities of Listeria monocytogenes isolates from different sources. Ankara Univ Vet Fak Derg. 2025;72(1):35-45.
  • Queipo-Ortuno MI, Morata P, Ocon P, Manchado P, Colmenero JD. Rapid diagnosis of human brucellosis by peripheral blood PCR assay. J Clin Microbiol. 1997;35(11):2927-30.
  • Baily GG, Krahn JB, Drasar BS, Stoker NG. Detection of Brucella melitensis and Brucella abortus by DNA amplification. J Trop Med Hyg. 1992;95:271-5.
  • Mayer-Scholl A, Draeger A, Golner C, Scholz HC, Nockler K. Advancement of a multiplex PCR for the differentiation of all currently described Brucella species. J Microbiol Methods. 2010;80:112-4.
  • Lopez-Goni I, Garcia-Yoldi D, Marin CM, De Miguel MJ, Munoz PM, Blasco JM, et al. Evaluation of a multiplex PCR assay (Bruce-ladder) for molecular typing of all Brucella species and of the vaccine strains. J Clin Microbiol. 2008;46:3484-7.
  • Charters AD. Brucellosis. Aust Fam Physician. 1980;9(10):707-12.
  • Nicoletti P. The epidemiology of bovine brucellosis. Adv Vet Sci Comp Med. 1980;24:69-98.
  • Franz DR, Jahrling PB, Friedlander AM, McClain DJ, Hoover DL, Bryne WR, et al. Clinical recognition and management of patients exposed to biological warfare agents. J Am Med Assoc. 1997;278(5):399-411.
  • Jones JD, Treanor JJ, Wallen RL, White PJ. Timing of parturition events in Yellowstone bison Bison bison: implications for bison conservation and brucellosis transmission risk to cattle. Wildl Biol. 2010;16(3):333-9.
  • Leski TA, Malanoski AP, Gregory MJ, Lin B, Stenger DA. Application of a broad-range resequencing array for detection of pathogens in desert dust samples from Kuwait and Iraq. Appl Environ Microbiol. 2011;77(13):4285-92.
  • Aune K, Rhyan JC, Russell R, Roffe TJ, Corso B. Environmental persistence of Brucella abortus in the Greater Yellowstone Area. J Wildl Manage. 2012;76(2):253-61.
  • Ahmed R, Muhammad K, Rabbani M, Khan MS. Spatial distribution of soil borne Brucella species specific DNA in Punjab, Pakistan. Pak J Zool. 2017;49(5):1739-48.
  • Calfee WM, Wendling M. The effects of environmental conditions on persistence and inactivation of Brucella suis on building material surfaces. Lett Appl Microbiol. 2012;54:504-10.
  • Garrido-Haro AD, Falconí M, Moreno-Caballeros P, Elena-Rovalino M, Rosero-Mayanquer H, Yugcha-Díaz M, et al. Determination and Characterization of (Novel) Circulating Strains of Brucella sp. Within the National Bovine Brucellosis Control Program in Ecuador. Pathogens. 2025;14(2):158.
  • Mazwi KD, Kolo FB, Jaja IF, Byaruhanga C, Hassim A, Van Heerden H. Polyphasic Characterization of Brucella spp. in Livestock Slaughtered from Abattoirs in Eastern Cape, South Africa. Microorganisms. 2024;12(1):223.
  • Tekle M, Legesse M, Edao BM, Ameni G, Mamo G. Isolation and identification of Brucella melitensis using bacteriological and molecular tools from aborted goats in the Afar region of north-eastern Ethiopia. BMC Microbiol. 2019;19:108.
  • Jumas-Bilak E, Michaux-Charachon S, Bourg G, Ramuz M, Allardet-Servent A. Unconventional genomic organization in the alpha subgroup of the Proteobacteria. J Bacteriol. 1998;180(10):2749-55.
  • Whatmore AM, Foster JT. Emerging diversity and ongoing expansion of the genus Brucella. Infect Genet Evol. 2021;92:104865.
  • Yagupsky P, Morata P, Colmenero JD. Laboratory diagnosis of human brucellosis. Clin Microbiol Rev. 2019;33(1):e00073-19.
  • Oren A, Garrity GM. List of new names and new combinations previously effectively, but not validly, published. Int J Syst Evol Microbiol. 2020;70:4043-9.
  • Hördt A, López MG, Meier-Kolthoff JP, Schleuning M, Weinhold L-M, Tindall BJ, et al. Analysis of 1,000+ type-strain genomes substantially improves taxonomic classification of Alphaproteobacteria. Front Microbiol. 2020;11:468.
  • Da Costa M, Guillou JP, Garin-Bastuji B, Thie´baud M, Dubray G. Specificity of six gene sequences for detecting the genus Brucella by DNA amplification. J Appl Bacteriol. 1996;81:267-75.
  • O’Leary S, Sheahan M, Sweeney T. Brucella abortus detection by PCR assay in blood, milk, and lymph tissue of serologically positive cows. Res J Vet Sci. 2006;81(2):170-6.
  • Romero C, Gamazo C, Pardo M, Lopez-Goni I. Specific detection of Brucella DNA by PCR. J Microbiol Methods. 1995;33(3):615-7.
  • Velasco J, Romero C, Lopez-Goni I, Leiva J, Diaz R, Morion I. Evaluation of the relatedness of Brucella spp. and Ochrobactrum anthropi and description of Ochrobactrum intermedium sp. nov., a new species with a closer relationship to Brucella spp. Int J Syst Bacteriol. 1998;48:759-68.
  • Saytekin AM, Ak S. Direct diagnosis of Brucella species through multiplex PCR formed by a new method. J Microbiol Methods. 2018;154:86-94.

Investigation of Brucella agents in soil-fertilizer mixtures and animal feed samples from cattle farms after extinguished brucellosis outbreaks

Year 2025, Volume: 96 Issue: 2, 131 - 139, 15.06.2025

Ahmet Murat Saytekin , Ayfer Güllü Yücetepe , Songül Ötkün , Sevil Erdenlig Gürbilek

https://doi.org/10.33188/vetheder.1631029

Abstract

Although brucellosis is under control in developed countries, it still threatens the health of animals and humans in the endemic regions. In these endemic regions although the outbreaks in some cattle farms have been officially extinguished, the farmers have wondered whether the bacteria are still present in some farm materials after extinguishment processes. The situation must be clarified so that farm production activities can continue and new animals can be procured in time. With this study, it was aimed to investigate the presence of Brucella bacteria in critical materials that may effectively transmit the disease after extinguishment procedures. For this reason, soil-fertilizer mixtures and animal feed were selected and investigated in three cattle farms where the disease was officially extinguished after the brucellosis outbreaks. The samples were collected approximately twenty days after completing extinction procedures for brucellosis. The soil-fertilizer mixtures and animal feed samples were collected in 68 and 55 pieces, respectively. The classic culture methods for the bacterial diagnosis of brucellosis were used with some modifications. After the growth of bacterial colonies on selective media, they were diagnosed by genus and species-specific PCRs. Five positive results were obtained by genus-specific PCR, but only one could be confirmed with species-specific multiplex PCR. For the remaining four, it was thought that they could belong to some soil bacteria genetically close to the Brucella genus. As a result, the brucellosis extinguishment procedures implemented could be considered adequate, and the farms were safe regarding the contamination. Although the official rules for disease extinction are fully implemented after outbreaks, similar studies are needed on more farms for more robust evaluations.

Keywords

B. abortus Cattle Culture PCR Soil

Ethical Statement

The ethical declaration has been obtained from the authors that the data, information and documents presented in this article have been obtained within the framework of academic and ethical rules, and that all information, documents, evaluations and results are presented in accordance with the rules of scientific ethics and ethics.

References

  • Manual of Diagnostic Tests and Vaccines for Terrestrial Animals (WOAH), Chapter 3.1.4. Brucellosis (Infection with B. abortus, B. melitensis and B. suis) [Internet]. 2024 Nov 29 [cited 2024 Dec 17]. Available from: https://www.woah.org/fileadmin/Home/eng/Health_standards/tahm/3.01.04_BRUCELLOSIS.pdf
  • O’Callaghan D. Human brucellosis: recent advances and future challenges. Infect Dis Poverty. 2020;9:1-2.
  • Cutler SJ, Whatmore AM, Commander AJ. Brucellosis, new aspects of an old disease. J Appl Microbiol. 2005;98:1270-81.
  • Refai M. Incidence and control of brucellosis in the Near East region. Vet Microbiol. 2002;90:81-110.
  • Taleski V, Zerva L, Kantardjiev T, Cvetnic Z, Erski-Biljic M, Nicolovski B, et al. An overview of the epidemiology and epizootology of brucellosis in selected countries of Central and Southeast Europe. Vet Microbiol. 2002;90(1-4):147-55.
  • Blasco JM. Control and eradication strategies for Brucella melitensis infection in sheep and goats. Prilozi. 2010;31(1):145-65.
  • Morales-Estradaa AI, Hernández-Castrob R, López-Merinoa A, Singh-Bedic J, Contreras-Rodrígueza A. Isolation, identification, and antimicrobial susceptibility of Brucella spp. cultured from cows and goat manure in Mexico. Arch Med Vet. 2016;48:231-5.
  • Aydın N. Gram negatif küçük çomaklar – Brucella infeksiyonları. In: Arda M, editor. Özel Mikrobiyoloji. Ankara: Medisan Yayınevi; 1998. p. 110–25.
  • Alton GG, Jones LM, Angus RD, Verger JM. Techniques for the brucellosis laboratory. 1st ed. Paris: Institut National de la Recherche Agronomique; 1998.
  • Hagan WA, Bruner DW, Timoney JF. The genus Brucella. In: Hagan WA, Bruner DW, Timoney JF, editors. Hagan and Bruner´s microbiology and infectious diseases of domestic animals. UK: Cornell University Press; 1988. p. 135-52.
  • U.S. Environmental Protection Agency (EPA). Persistence of Categories A and B Select Agents in Environmental Matrices-EPA/600/R-14/074 [Internet]. 2014 Jun 1 [cited 2024 Oct 17]. Available from: https://www.researchgate.net/publication/289245437_Persistence_of_Categories_A_and_B_Select_Agents_in_Environmental_Matrices
  • Farrell ID. The development of a new selective medium for the isolation of Brucella abortus from contaminated sources. Res J Vet Sci. 1974;16:280-6.
  • Saytekin AM, Adıgüzel A, Al-Kilani K, Güllü Yücetepe A, Keskin O. Some virulence genes and biofilm formation capabilities of Listeria monocytogenes isolates from different sources. Ankara Univ Vet Fak Derg. 2025;72(1):35-45.
  • Queipo-Ortuno MI, Morata P, Ocon P, Manchado P, Colmenero JD. Rapid diagnosis of human brucellosis by peripheral blood PCR assay. J Clin Microbiol. 1997;35(11):2927-30.
  • Baily GG, Krahn JB, Drasar BS, Stoker NG. Detection of Brucella melitensis and Brucella abortus by DNA amplification. J Trop Med Hyg. 1992;95:271-5.
  • Mayer-Scholl A, Draeger A, Golner C, Scholz HC, Nockler K. Advancement of a multiplex PCR for the differentiation of all currently described Brucella species. J Microbiol Methods. 2010;80:112-4.
  • Lopez-Goni I, Garcia-Yoldi D, Marin CM, De Miguel MJ, Munoz PM, Blasco JM, et al. Evaluation of a multiplex PCR assay (Bruce-ladder) for molecular typing of all Brucella species and of the vaccine strains. J Clin Microbiol. 2008;46:3484-7.
  • Charters AD. Brucellosis. Aust Fam Physician. 1980;9(10):707-12.
  • Nicoletti P. The epidemiology of bovine brucellosis. Adv Vet Sci Comp Med. 1980;24:69-98.
  • Franz DR, Jahrling PB, Friedlander AM, McClain DJ, Hoover DL, Bryne WR, et al. Clinical recognition and management of patients exposed to biological warfare agents. J Am Med Assoc. 1997;278(5):399-411.
  • Jones JD, Treanor JJ, Wallen RL, White PJ. Timing of parturition events in Yellowstone bison Bison bison: implications for bison conservation and brucellosis transmission risk to cattle. Wildl Biol. 2010;16(3):333-9.
  • Leski TA, Malanoski AP, Gregory MJ, Lin B, Stenger DA. Application of a broad-range resequencing array for detection of pathogens in desert dust samples from Kuwait and Iraq. Appl Environ Microbiol. 2011;77(13):4285-92.
  • Aune K, Rhyan JC, Russell R, Roffe TJ, Corso B. Environmental persistence of Brucella abortus in the Greater Yellowstone Area. J Wildl Manage. 2012;76(2):253-61.
  • Ahmed R, Muhammad K, Rabbani M, Khan MS. Spatial distribution of soil borne Brucella species specific DNA in Punjab, Pakistan. Pak J Zool. 2017;49(5):1739-48.
  • Calfee WM, Wendling M. The effects of environmental conditions on persistence and inactivation of Brucella suis on building material surfaces. Lett Appl Microbiol. 2012;54:504-10.
  • Garrido-Haro AD, Falconí M, Moreno-Caballeros P, Elena-Rovalino M, Rosero-Mayanquer H, Yugcha-Díaz M, et al. Determination and Characterization of (Novel) Circulating Strains of Brucella sp. Within the National Bovine Brucellosis Control Program in Ecuador. Pathogens. 2025;14(2):158.
  • Mazwi KD, Kolo FB, Jaja IF, Byaruhanga C, Hassim A, Van Heerden H. Polyphasic Characterization of Brucella spp. in Livestock Slaughtered from Abattoirs in Eastern Cape, South Africa. Microorganisms. 2024;12(1):223.
  • Tekle M, Legesse M, Edao BM, Ameni G, Mamo G. Isolation and identification of Brucella melitensis using bacteriological and molecular tools from aborted goats in the Afar region of north-eastern Ethiopia. BMC Microbiol. 2019;19:108.
  • Jumas-Bilak E, Michaux-Charachon S, Bourg G, Ramuz M, Allardet-Servent A. Unconventional genomic organization in the alpha subgroup of the Proteobacteria. J Bacteriol. 1998;180(10):2749-55.
  • Whatmore AM, Foster JT. Emerging diversity and ongoing expansion of the genus Brucella. Infect Genet Evol. 2021;92:104865.
  • Yagupsky P, Morata P, Colmenero JD. Laboratory diagnosis of human brucellosis. Clin Microbiol Rev. 2019;33(1):e00073-19.
  • Oren A, Garrity GM. List of new names and new combinations previously effectively, but not validly, published. Int J Syst Evol Microbiol. 2020;70:4043-9.
  • Hördt A, López MG, Meier-Kolthoff JP, Schleuning M, Weinhold L-M, Tindall BJ, et al. Analysis of 1,000+ type-strain genomes substantially improves taxonomic classification of Alphaproteobacteria. Front Microbiol. 2020;11:468.
  • Da Costa M, Guillou JP, Garin-Bastuji B, Thie´baud M, Dubray G. Specificity of six gene sequences for detecting the genus Brucella by DNA amplification. J Appl Bacteriol. 1996;81:267-75.
  • O’Leary S, Sheahan M, Sweeney T. Brucella abortus detection by PCR assay in blood, milk, and lymph tissue of serologically positive cows. Res J Vet Sci. 2006;81(2):170-6.
  • Romero C, Gamazo C, Pardo M, Lopez-Goni I. Specific detection of Brucella DNA by PCR. J Microbiol Methods. 1995;33(3):615-7.
  • Velasco J, Romero C, Lopez-Goni I, Leiva J, Diaz R, Morion I. Evaluation of the relatedness of Brucella spp. and Ochrobactrum anthropi and description of Ochrobactrum intermedium sp. nov., a new species with a closer relationship to Brucella spp. Int J Syst Bacteriol. 1998;48:759-68.
  • Saytekin AM, Ak S. Direct diagnosis of Brucella species through multiplex PCR formed by a new method. J Microbiol Methods. 2018;154:86-94.
There are 38 citations in total.

Details

Primary Language English
Subjects Veterinary Microbiology
Journal Section RESEARCH ARTICLE
Authors

Ahmet Murat Saytekin 0000-0001-7486-8054

Ayfer Güllü Yücetepe 0000-0002-9842-3305

Songül Ötkün 0000-0003-2736-953X

Sevil Erdenlig Gürbilek 0000-0002-0377-2650

Early Pub Date June 13, 2025
Publication Date June 15, 2025
Submission Date February 1, 2025
Acceptance Date May 7, 2025
Published in Issue Year 2025 Volume: 96 Issue: 2

Cite

Vancouver Saytekin AM, Güllü Yücetepe A, Ötkün S, Erdenlig Gürbilek S. Investigation of Brucella agents in soil-fertilizer mixtures and animal feed samples from cattle farms after extinguished brucellosis outbreaks. Vet Hekim Der Derg. 2025;96(2):131-9.
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