Solunum Sisteminden İzole Edilen Mannheimia haemolytica izolatlarının tiplendirilmesi ve virulens genlerinin araştırılması

Year 2024, Volume: 13 Issue: 3, 21 - 30, 26.09.2024

Cihat Öztürk , İsmail Hakkı Ekin

https://doi.org/10.46810/tdfd.1402193

Abstract

Bu çalışmada, hasta/sağlıklı sığır ve koyunların solunum yollarından izole edilen Mannheimia haemolytica izolatlarının biyokimyasal özellikleri, önemli virülens genlerinin dağılımı tespit edilmesi amaçlandı. Nazo-farengiyal ve trake-bronşiyal svap örneklerinden Real-Time PCR ile 48 (%87.3) adet Mannheimia haemolytica izolatı kullanıldı. İncelenen izolatlarda, hastalık ve virülens genleri ile ilişkili biyokimyasal özelliklerin arginin ve sorbitol testlerindeki farklılıklara göre 4 farklı biyokimyasal profil belirlendi. Real Time-PCR yöntemiyle virülens genleri incelenen izolatlarda, %37.5’i I, %33.3’ü III ve %12.5’i II olmak üzere 3 farklı virülens gen profili tespit edildi. Virülens gen profil II’ye sahip izolatlarda virülens ile ilişkili genlerin tamamı belirlenirken, profil I’e sahip izolatlarda nmaA, profil III’e sahip izolatlarda ise nmaA ve tbpB genlerinin bulunmadığı tespit edildi. Bununla birlikte biyokimyasal profil II’nin hastalık olguları ile ilişkili ve bunun arginin negatiflik ile ilişkili olduğu belirlendi. Ayrıca virülens gen profil I özelliğine sahip izolatların sadece biyokimyasal profil I ile ilişkili olduğu ve bunun arginin negatiflikten kaynaklandığı belirlenirken, arginin pozitif izolatlar ile virülens gen profil III arasındaki ilişkinin önemli olduğu gözlendi. Sonuç olarak; arginin negatiflik ile gcp, gs60, tbpB, lktC, adh pozitif, nmaA negatif izolatların kommensal ve patojen Mannheimia haemolytica izolatlarının ayrımında kullanılabilecek epidemiyolojik kriter olabileceği ve konu ile ilgili yeni çalışmaların yapılması gerektiği düşünülmektedir.

Keywords

Mannheimia haemolytica, Tiplendirme, Virülens gen

Ethical Statement

Çalışma Van Yüzüncü Yıl Üniversitesi Hayvan Deneyleri Yerel Etik Kurul 01.03.2018 tarih ve 02 sayılı karar gereğince izinleri alınarak yapılmıştır.

Supporting Institution

Çalışma; Van Yüzüncü Yıl Üniversitesi Bilimsel Araştırma Projeleri Başkanlığı tarafından desteklenmiştir

Project Number

TDK-2018-7156

Typing of Mannheimia haemolytica isolates isolated from respiratory tract and investigation of virulence genes

Abstract

In this study, it was aimed to determine the biochemical properties, distribution of important virulence genes of Mannheimia haemolytica isolates identified from the respiratory tracts of sick and healthy cattles and sheeps. 48 (87.3%) Mannheimia haemolytica isolates from naso-pharyngeal and trachea-bronchial swaps were identified as Mannheimia haemolytica by Real Time-PCR. According to the differences in arginine and sorbitol tests, 4 different biochemical profiles were determined in the isolates examined. Three virulence gene profiles were detected in the isolates examined by Real-Time PCR. 37.5%, 33.3%, 12.5% of the isolates examined were identified as I, III and II, respectively. While all virulence-related genes were identified in the isolates with virulence gene profile II, it was determined that there were no nmaA gene in profile I isolates and nmaA and tbpB genes in profile III isolates. At the same time, it was determined that biochemical profile II was associated with disease cases and this was related to arginine negativity. In addition, it was determined that isolates with virulence gene profile I were associated only with biochemical profile I and that this was due to arginine negativity, whereas the relationship between arginine positive isolates and virulence gene profile III was found to be significant. As a result; arginine negativity and gcp, gs60, tbpB, lktC, adh positive, nmaA negative isolates may be the epidemiological criteria that can be used to differentiate commensal and pathogen Mannheimia haemolytica isolates and new studies on the subject should be done.

Keywords

Mannheimia haemolytica, Typing, Virulence genes

Ethical Statement

The study was conducted with the permission of Van Yüzüncü Yıl University Animal Experiments Local Ethics Committee, decision numbered 02 dated 01.03.2018.

Supporting Institution

The study was supported by Van Yüzüncü Yıl University Scientific Research Projects Directorate.

Project Number

TDK-2018-7156

References

• Et ve Süt Kurumu. Et ve Süt Kurumu sektör değerlendirme raporu. Ankara. 2016.
• TÜİK Hayvancılık İstatistikleri (İnternet). 2019. [Erişim tarihi: 29.03.2020]. Erişim adresi: https://biruni.tuik.gov.tr/medas/?kn=101&locale=tr.
• Global Animal Healt Association, Animal breeding [İnternet]. 2019, [Erişim tarihi 17 Mayıs 2019]. Erişim_adresi: https://healthforanimals.org/zoonoses.html.
• Bell S. Respiratory disease in sheep: 2. Treatment and control. In Practice. 2008; 30: 278-283.
• Ermilio EM, Smith MC. Treatment of emergency conditions in sheep and goats. Vet Clin Food Anim. 2011; 27: 33-45.
• Statham J. Respiratory disease in cattle a practical approach. Livestock. 2018; 23 (5): 206-13.
• Kumar A, Tikoo SK, Malik P, Kumar AT. Respiratory diseases of small ruminants. Hindawi Publishing Corporation Veterinary Medicine International. 2014; 373642: 2.
• Şentürk B. Türkiye’de salgın hayvan hastalık sorunu ve yeni model önerileri. Harran Üniv Vet Fak Derg. 2015; 4 (1), 27-9.
• Rahal A, Ahmad AH, Prakash A, Mandil R, Kumar RT. Environmental attributes to respiratory diseases of small ruminants. Vet Med INT. 2014; 853627: 10.
• Öztürk D, Çorlu M. Pnömonili Koyun akciğerlerinden izole edilen bakteriler ve antibiyotik duyarlılıkları. Vet. Bil. Derg. 2006; 22 (1-2): 59-63
• Yüzbaşıgil AF (2010). Kuzu pnömonilerinde patolojik ve bakteriyolojik ı̇ncelemeler ile Parainfluenza 3 (PI 3) virüsünün etiyolojideki rolü. Selçuk Üniversitesi Sağlık Bilimleri Enstitüsü, Doktora Tezi.
• Rosadio R, Cirilo E, Manchego A, Rivera H. Respiratory syncytial and parainfluenza type 3 viruses coexisting with Pasteurella multocida and Mannheimia hemolytica in acute pneumonias of neonatal alpacas. Small Ruminant Res. 2011; 97: 110–6.
• Arslan HH, Özcan U. Current approach to bovine respiratory disease. Dairy and Vet Sci J. 2018; 5 (2): 1-3
• Jesse FFA, Mubin HNA, Hambali IU, Lila MAM, Chung ELT, Abba Y, et al. Review on clinical management involving respiratory diseases in ruminants. Adv Anim. Vet. Sci. 2019; 7(4): 321-5.
• Confer AW. Update on bacterial pathogenesis in BRD. Anim Health Res Rev. 2009 Dec;10(2):145-8. doi: 10.1017/S1466252309990193. PMID: 20003651.
• Tomassini L, Gonzales B, Weiser GC, Sischo W. An ecologic study comparing distribution of Pasteurella trehalosi and Mannheimia haemolytica between Sierra Nevada bighorn sheep, White Mountain Bighorn Sheep, and Domestic sheep. J Wildl Dis. 2009; 45 (4): 930-40.
• Hossain MM, Islam MS, Kamal AHM, Rahman AKMA, Cho HS. Dairy cattle mortality in an organized herd in Bangladesh. VeterinaryWorld. 2014; 7 (5): 331-6.
• Caswell JL, Williams KJ. Respiratory system. Maxie M.G. (Ed.) Jubb, Kennedy and Palmer’s Pathology of Domestic. 5. baskı. Edinburgh: Elsevier; 2016.
• Angen O, Ahrens P, Kuhnert P, Christensen H, Mutters R. Proposal of Histophilus somni gen. nov., sp. nov. for the three species incertae sedis Haemophilus somnus, Haemophilus agni and Histophilus ovis. Int J Syst Evol Microbiol. 2003; 53: 1449-56.
• Christensen H, Bisgaard M. Revised definition of Actinobacillus sensu stricto isolated from animals. A review with special emphasis on diagnosis. Vet. Microbiol. 2004; 99: 13-30.
• Blackall PJ, Bojesen AM, Christensen H, Bisgaard M. Reclassification of Pasteurella trehalosi as Bibersteinia trehalosi gen. nov., comb. nov. Int J Syst Evol Microbiol. 2007; 57: 666−74.
• Yavrucu E. Mannheimia haemolytica’nın sığır epitel hücreleri üzerindeki apoptotik etkisi [Yüksek Lisans tezi]. Ankara: Ankara Üniversitesi; 2008.
• Quinn PJ (Editor), Markey BK, Leonard FC, Fitzpatrick ES, Fanning S, Hartigan PJ. Veterinary Microbiology and Microbial Disease, UK: John Wiley and Sons Ltd; 27: 300-308. 2011.
• Mutters R, Angen Q, Bisgaard M. Bergey's manual of systematics of archaea and bacteri, Online Bergey’s Manual Trust. (e-book). Published by John Wiley and Sons. in association with Bergey’s
• Manual Trust. 2015. Doi: 10.1002/9781118960608.gbm01200.
• Angen Ø, Mutters R, Caugant DA, Olsen JE, Bısgaard M. Taxonomic relationships of the [Pasteurella] haemolytica complex as evaluated by DNA-DNA hybridizations and 16S rRNA sequencing with proposal of Mannheimia haemolytica gen. nov., comb. nov., Mannheimia granulomatis comb. nov., Mannheimia glucosida sp. nov., Mannheimia ruminalis sp. nov. and Mannheimia varigena sp. nov. Int J Syst Evol Microbiol 1999; 49: 67-86.
• Omaleki L, Browning FG, Allen JL, Barber SR. Molecular epidemiology of Mannheimia haemolytica and Mannheimia glucosida associated with ovine mastitis. JVDI. 2012; 24(4): 730-4.
• Nefedchenko AV, Glotova TI, Glotov AG. Isolation and antimicrobial resistance of Mannheimia haemolytica on dairy farms in Siberia. Bulg J Vet Med. 2019; 22 (4): 428-38.
• Highlander SK. Molecular genetic analysis of virulence in Mannheimia (Pasteurella) haemolytica, Frontiers in Bioscience. 2001; 6: 1128-50.
• Lee RWH, Pool AN, Ziauddin A, Lo RYC, Shewen PE, Strommer JN. Edible vaccine development: stability of Mannheimia haemolytica A1 leukotoxin 50 during post-harvest processing and storage of field-grown transgenic white clover. Kluwer Academic Publshers Molecular Breeding. 2003; 11: 259-66.
• Zecchinon L, Fett T, Desmecht D. How Mannheimia haemolytica defeats host defence through a kiss of death mechanism. Vet Res. 2005; 36: 133-56.
• De la Mora A, Trigo F, Jaramillo L, Garfies Y, Solórzano C, Agundis C et al. The N-acetyl-D-glucosamine specific adhesin from Mannheimia haemolytica activates bovine neutrophils oxidative burst. Vet Immunol Immunopathol. 2006; 113 (1-2): 148-56.
• Lo RYC, Sathiamoorthy S, Shewen PE. Analysis of in vivo expressed genes in Mannheimia haemolytica A1. FEMS Microbiol Lett. 2006; 265: 18-25.
• Lee RWH, Cornelisse M, Ziauddin A, Slack PJ, Hodgins DC, Strommer JN, et al. Expression of a modified Mannheimia haemolytica gs60 outer membrane lipoprotein in transgenic alfalfa for the development of an edible vaccine against Bovine Pneumonic Pasteurellosis. J Biotechnol. 2008; 135: 224-31.
• Mohamed RA, Abdelsalam EB. A review on Pneumonic Pasteurellosis (Respiratory Mannheimiosis) with emphasis on pathogenesis, virulence mechanisms and predisposing factors. Bulg J Vet Med. 2008; 11 (3): 139-60.
• Rice JA, Carrasco-Medina L, Hodgins DC, Shewen PE. Mannheimia haemolytica and bovine respiratory disease. Anim Health Res Rev. 2008; 8 (2): 117-28.
• Hounsome JDA. Comparative outer membrane proteomic analyses of bovine and ovine isolates of Mannheimia haemolytica and Mannheimia glucosida [PhD thesis]. UK; University of Glasgow. 2012.
• Puchalski A, Chmiel RU, Dec M, Wernicki A. An electrophoretic characterization of ıron-transporting proteins in Mannheimia haemolytica A1. Pol J Vet. 2013; 3: 527-32.
• Legesse A, Abayneh T, Mamo G, Gelaye E, Tesfaw L, Yami M, et al. Molecular characterization of Mannheimia haemolytica Isolates associated with Pneumonic cases of sheep in selected Areas of Central Ethiopia. BMC Microbiol. 2018; 18: 205.
• Singh F, Sonawane GG, Meena RK. Molecular detection of virulent Mannheimia haemolytica and Pasteurella multocida in lung tissues of pneumonic sheep from semiarid tropics, Rajasthan, India. Turk J Vet Anim Sci. 2018; 42: 556-1.
• Özbey G, Muz A. Pnömonili Koyun ve Keçilerin Akciğerlerinden Aerobik Bakteri izolasyonları ve İzole Pasteurella multocida ve Mannheimia haemolytica’nın Polimeraz Zincir Reaksiyonu ile Saptanması. Turk J Vet Anim Sci. 2004; 28: 209-16.
• Gioia J, Qin X, Jiang H, Clinkenbeard K, L0 R, Liu Y, et al. The genome sequence of Mannheimia haemolytica A1: Insights into virulence, natural competence, and Pasteurellaceae phylogeny. J. Bacteriol. 2006; 7257-66.
• Alexander TW, Cook SR, Yanke LJ, Booker CW, Morley PS, Read RR, et al. A multiplex polymerase chain reaction assay for the identification of Mannheimia haemolytica, Mannheimia glucosida and Mannheimia ruminalis. Vet Microbiol. 2008; 130 (1-2): 166-75.
• Kumar J, Shivendra D, Kumar R. Rapid detection of Mannheimia haemolytica in lung tissues of sheep and from bacterial culture. Veterinary World. 2015; 8 (9): 1073-7.
• Shanthalingam S, Goldy A, Bavananthasivam J, Subramaniam R, Batra SA, Kugadas A, et al. PCR assay detects Mannheımıa haemolytıca in culture negatıve pneumonıc lung tıssues of Bıghorn Sheep (Ovıs Canadensıs) from outbreaks in the Western Usa, 2009–2010. J Wildl Dis. 2014; 50 (1): 1-10.
• Ahmed WA, Mohammed RJ, Khalaf IA. Molecular and phenotypical characterization of Mannheimia haemolytica ısolated from goats in Baghdad Province. AIM. 2017; 7: 304-14.
• Dağ S, Gürbüz A, Özen H, Büyük F, Çelebi Ö, Karaman M, et al. Immunohistochemical and molecular detection of Mannheimia spp. and Pasteurella spp. in sheep with pneumonia in Kars Province – Turkey. Kafkas Univ Vet Fak Derg. 2018; 24 (2): 281-8.
• Rawat N, Gilhare VR, Kushwaha KK, Hattimare DD, Khan FF, Shende RK et al. Isolation and molecular characterization of Mannheimia haemolytica and Pasteurella multocida associated with pneumonia of goats in Chhattisgarh. Vet World. 2019; 12 (2): 331-6.
• Koneman EW (Editor), Allen SD, Dowell VR, Janda WM, Sommers HM, Winn WC. Color Atlas and Text Book of Diagnostic Microbiology. Third Edition. Lippincott Comp., Philadelphia, USA. 1988.
• Carter GR (1984). Pasteurella, Yersinia and Francisella, chapter 11, in “Diagnostic Procedures in Veterinary Bacteriology and Mycologyˮ Fourth Edition, 111-121, Charles C Thomas, U.S.A.
• Önat K, Kahya S, Çarlı T. Frequency and antibiotic susceptibility of Pasteurella multocida and Mannheimia haemolytica isolates from nasal cavities of Cattle. Turk J Vet Anim Sci. 2010; 34 (1): 91-4.
• Angen Ø, Ahrens P, Bisgaard M. Phenotypic and genotypic characterization of Mannheimia (Pasteurella) haemolytica-like strains isolated from diseased animals in Denmark. Vet Mic. 2002; 84: 103-14.
• Laishevtcev AI, ОСОБЕННОСТИ БИОХИМИЧЕСКОЙ ИДЕНТИФИКАЦИИ И ДИФФЕРЕНЦИАЦИИ БАКТЕРИЙ ВИДА Mannheimıa haemolytica (Features of biochemical identification and differentiation of Mannheimıa haemolytica). RJOAS. 2016; 6 (54): 70-7.
• Hawari AD, Hassawi DS, Sweiss M. Isolation and identification of Mannheimia haemolytica and Pasteurella multocida in sheep and goats using biochemical tests and Random Amplified Polymorphic DNA (RAPD) Analysis. J Biol Sci. 2008; 8(7): 1251-4.
• Klima CL, Alexander TW, Hendrick S, McAllister AT. Characterization of Mannheimia haemolytica isolated from feedlot cattle that were healthy or treated for bovine respiratory disease. Can J Vet Res.2014; 78 (1): 38-45.
• Gülaydın Ö, Gürtürk K. İdentification of Pasteurella multocida strains isolated from respiratory tract of healthy and diseased cattle and determination of capsular types by PCR in Van Region. Van Vet J. 2018; 29 (3): 143-6.
• Ülker H, Kucuk D, Cantekin Z, Solmaz H (2012). Hatay yöresinde kesimhanede kesilen sığır akciğerlerinden Pasteurella multocida ve Mannheimia haemolytica izolasyonu ve antibiyotiklere duyarlılığı. AVKAE Derg, 2, 2, 10-14.
• Gürbüz A. Sığır ve koyunlara ait pnömonili akciğerlerden Pasteurella haemolytica’nın izolasyonu, identifikasyonu, biyotiplendirilmesi ve antibiyotiklere olan duyarlılıklarının belirlenmesi. [Doktora Tezi]. Kars: Kafkas Üniversitesi. 2003.
• Noyes NR, Benedict KM, Gow SP, Booker CW, Hannon SJ, McAllister TA, Morley PS (2015). Mannheimia haemolytica in feedlot cattle: prevalence of recovery and associations with antimicrobial use, resistance, and health outcomes. Journal of Veterinary Internal Medicine 29, 705–713.
• Castillo JLR, Valencia GL, Monge-Navara FJ, Medina-Basulto GE, Hori-Oshima S, Cueto-Gonzalez SA, ve ark. Detection and economic impact related to bovine respiratory disease, shrink, and traveling distance in feedlot cattle in Northwest Mexico. Turk J Vet Anim Sci. 2017; 41: 294-301.
• Demissie T, Dawo F, Sisay T. Biochemical and antigenic characterization of Mannheimia, pasteurella and Mycoplasma species from naturally infected pneumonic sheep and goats, Bishoftu, Ethiopia. AJBAS. 2014; 6 (6): 198-204.
• Kırkan Ş, Kaya O. Serotyping of Mannheimia haemolytica Strains Isolated from Pneumonic Lungs of Sheep in the Aydın Region of Turkey. Turk J Vet Anim Sci. 2005; 29: 491-4.
• Tel OY, Keskin O. Koyun akciğerlerinden Pasteurella multocida ve Mannheimia haemolytica izolasyonu ve antibiyotiklere duyarlılığı. YYÜ Vet Fak Derg. 2010; 21 (1): 31.
• Bisgaard M. Incidence of Pasteurella haemolytica in the respiratory tract of Apparently healthy chickens and chickens with infectıous bronchitis. Characterisation of 213 Strains. Avian Pathology. 1977; 6: 285-292.
• Daphal SH, Mhasa PP, Pawade MM, Shelke PP, Sangle JD, Kolhe RP. Emergence of virulent Pasteurella multocida and Mannheimia hemolytica in sheep and goats of Western Maharashtra, India. Int J Curr Microbiol App Sci. 2018; 7 (9): 1990-1998.
• Angen Q, Aalbjek B, Falsen E, Olsen JE, Bisgaard M. Relationships among strains classified with the ruminant Pasteurella haemolytica-complex using quantitative evaluation of phenotypic Data. Zbl. Bakt. 1997; 285: 459-79.
• Taunde PA, Argenta FF, Bianchi RM, De Cecco BS, Vielmo A, Lopes BC, et al. Mannheimia haemolytica pleuropneumonia in goats associated with shipping stress. Ciencia Rural. 2019; 49(1): 1-6.
• İlhan Z, Keleş İ. Biotyping and serotyping of Mannheimia (Pasteurella) haemolytica isolated from lung samples of slaughtered sheep in the Van Region. Turk J Vet Anim Sci. 2007; 31 (2): 137-41
• Şahin İ. Samsun ili ve çevresinde koyun pnömonilerinden izole edilen Mannheimia haemolytica türlerinin fenotipik karakterizasyonu ve antibiyotik duyarlılıklarının belirlenmesi. Samsun, Ondokuz Mayıs Üniversitesi. 2017.
• Fisher MA, Weiser GC, Hunter DL, Ward ACS. Use of a Polymerase Chain Reaction method to detect the leucotoxine gene lktA in biogrup and biovariant isolates of Pasteurella haemolytica and Pasteurella trehalosi. Am. J. Vet. Res. 1999; 11: 1402-6.
• Kırkan Ş. Aydın yöresinde koyunların solunum sisteminde infeksiyon nedeni Mannheimia (Pasteurella) haemolytica’nın biyotip ve Serotip tayini, elektroforez ve PCR ile tanısı. [Doktora Tezi]. Aydın: Adnan Menderes Üniversitesi. 2003.
• El Dokmak MM, Khalil SA, Ebied SKHM. Genetic diversity of Mannheimia haemolytica strains. AJVS. 2015; 47: 166-74.
• Garcia-Alvarez A, Garayzabal JF, Chaves F, Pinto C, Cid D. Ovine Mannheimia haemolytica Isolates from Lungs with and without Pneumonic Lesions Belong to Similar Genotypes. Vet Microbiol. 2018; 219: 80-6.