Bronkopnömonili Köpeklerde Mycoplasma spp., Streptococcus spp., Bordetella bronchiseptica, Klebsiella spp. ’nin Real-time PCR ve Patolojik Yöntemler ile Teşhisi

Year 2023, Volume: 34 Issue: 2, 129 - 138, 28.12.2023

Gökhan Akçakavak Nevin Tuzcu Özgür Özdemir Mehmet Tuzcu Osman Dağar Ayşenur Tural Fatih Mehmet Öztürk Zeynep Çelik Nida Sönmezler Mehmet Halit Atik Özgür Kanat

https://doi.org/10.35864/evmd.1354193

Abstract

Köpeklerde solunum yolu hastalıklarına sebep olan çok sayıda bakteriyel etken vardır. Bakteriyel bronkopnömoni, alt solunum yollarında ve akciğer parankiminde bakterilerin neden olduğu akciğer hastalığıdır. Bu çalışmada bronkopnömonili köpeklerde Mycoplasma spp., Streptococcus spp., Bordetella bronchiseptica, Klebsiella spp.’nin Real-time PCR ile belirlenmesi ve etkenlere göre belirlenen histopatolojik bulguların karşılaştırılması amaçlanmıştır. Çalışmanın materyalini farklı sebeplerden dolayı ölen ve mikroskobik muayenede bronkopnömoni belirlenen 37 adet köpeğe ait akciğer parafin bloklar oluşturdu. Mikroskobik olarak alveol lümenlerinde ödem, alveol epitellerinde dökülme, bronş ve bronşiol epitellerinde dökülme ve bronşektazi, bronşiol lümenlerinde mononükleer hücre (MNH) ve Polimorfonükleer lökosit (PMNL) infiltrasyonu, peribronşioler MNH infiltrasyonu, bronş epitellerine lokalize olmuş bakteri kümeleri, intersitisyumda PMNL infiltrasyonu, multifokal nekroz alanları, kanama ve plöritis rastlandı. Real-time PCR incelemesinde 18 (%48.64) olguda Bordetella bronchiseptica, 9 (%24.32) olguda Mycoplasma spp., 10 (%27.02) olguda Streptococcus spp., 2 (%5.4) olguda Klebsiella spp. saptandı. Sonuç olarak bu çalışma, Real-time PCR ile köpeklerin bakteriyel bronkopnömonilerinde kültür imkânı olmayan dokularda bile etkenin belirlenebileceğini göstermiştir. Ayrıca köpeklerde polimikrobiyal alt solunum yolu enfeksiyonlarının da görülebileceğine işaret ederek teşhis için birden fazla bakteri türünün araştırılması gerektiğini ortaya koymaktadır.

Keywords

Bronkopnömoni, Köpek, Patoloji, Real-time PCR

Diagnosis of Mycoplasma spp., Streptococcus spp., Bordetella bronchiseptica, Klebsiella spp., by Real-time PCR and Pathological Methods in Dogs with Bronchopneumonia

Abstract

There are many bacterial factors that cause respiratory diseases in dogs. Bacterial bronchopneumonia is a lung disease caused by bacteria in the lower respiratory tract and lung parenchyma. In this study, it was aimed to determine Mycoplasma spp., Streptococcus spp., Bordetella bronchiseptica, Klebsiella spp., in dogs with bronchopneumonia by Real-time PCR and to compare the histopathological findings determined according to the agents. The material of the study consisted of lung paraffin blocks of 37 dogs that died due to different reasons and were found to have bronchopneumonia in microscopic examination. Microscopically, edema in the alveolar lumens, shedding of the alveolar epithelium, shedding of the bronch and bronchial epithelium and bronchiectasis, mononuclear cell infiltration (MCI) and Polymorphonuclear leukocytes (PMNL) infiltration in the bronchial lumens, peribronchiolar MCI infiltration, Bacterial clusters localized to the bronch epithelium, PMNL infiltration in the interstitium, multifocal necrosis areas, bleeding and pleuritis were observed. Real-time PCR analysis revealed Bordetella bronchiseptica in 18 (48.64%) cases, Mycoplasma spp. in 9 (24.32%) cases, Streptococcus spp. in 10 (27.02%) cases, and Klebsiella spp. in 2 (5.4%) cases. In conclusion, this study showed that the causative agent can be determined in bacterial bronchopneumonias of dogs with Real-time PCR even in tissues without culture opportunity. In addition, this study indicates that polymicrobial lower respiratory tract infections can also be seen in dogs and reveals that more than one bacterial species should be investigated for diagnosis.

Keywords

Bronchopneumonia, Dog, Pathology, Real-time PCR

References

• Akcakavak G, Tuzcu M, Tuzcu N, Celik Z, Tural A, Dagar O. (2023) Investigation with Real- Time PCR and Histopathology on the presence of H. felis, H. heilmannii and H. pylori in dogs, Rev Cient. 33, 1-7.
• Aher T, Roy A, Kumar P. (2012) Molecular detection of virulence genes associated with pathogenicity of Klebsiella spp. isolated from the respiratory tract of apparently healthy as well as sick goats. Isr J Vet Med. 67, 249-52.
• Armstrong D, Morton V, Friedman M, Steger L, Tully J. (1972) Canine pneumonia associated with mycoplasma infection. Am J Vet Res., 33, 1471-1478.
• Buonavoglia C, Martella V. (2007) Canine respiratory viruses. Vet res. 38, 355-73.
• Carvalho I, Alonso CA, Silva V, Pimenta P, Cunha R, Martins C, Igrejas G, Torres C, Poeta P. (2020) Extended-spectrum beta-lactamase-producing Klebsiella pneumoniae isolated from healthy and sick dogs in Portugal. Microb Drug Resist. 26, 709-715.
• Chalker VJ, Brooks HW, Brownlie J. (2003) The association of Streptococcus equi subsp. zooepidemicus with canine infectious respiratory disease. Vet Microbiol. 95, 149-156.
• Chambers J, Matsumoto I, Shibahara T, Haritani M, Nakayama H, Uchida K. (2019) An outbreak of fatal Bordetella bronchiseptica bronchopneumonia in puppies. J Comp Pathol. 167, 41-45.
• Chandler JC, Lappin MR. (2002) Mycoplasmal respiratory infections in small animals: 17 cases (1988–1999). JAAHA 38, 111-119.
• Dear JD. (2020) Bacterial pneumonia in dogs and cats: An update. Veterinary Clinics: J Small Anim Pract. 50, 447-465.
• Ellis JA, Haines DM, West KH, Burr JH, Dayton A, Townsend HG, Kanara EW, Konoby C, Crichlow A, Martin K. (2001) Effect of vaccination on experimental infection with Bordetella bronchiseptica in dogs. J Am Vet Med Assoc. 218, 367-375.
• Facklam R. (2002) What happened to the streptococci: overview of taxonomic and nomenclature changes. Clin Microbiol Rev., 15, 613-630.
• Ford RB. (2006) Canine infectious tracheobronchitis. Infectious diseases of the dog and cat.
• Foster S, Martin P, Braddock J, Malik R. (2004) A retrospective analysis of feline bronchoalveolar lavage cytology and microbiology (1995–2000). J Feline Med Surg, 6, 189-198.
• Greisen K, Loeffelholz M, Purohit A, Leong D. (1994) PCR primers and probes for the 16S rRNA gene of most species of pathogenic bacteria, including bacteria found in cerebrospinal fluid. J Clin. Microbiol. 32,335-351.
• Hulse LS, Hickey D, Mitchell JM, Beagley KW, Ellis W, Johnston SD. (2018) Development and application of two multiplex real-time PCR assays for detection and speciation of bacterial pathogens in the koala. J Vet Diagn Invest. 30, 523-529.
• Jameson P, King L, Lappin M, Jones R. (1995) Comparison of clinical signs, diagnostic findings, organisms isolated, and clinical outcome in dogs with bacterial pneumonia: 93 cases (1986-1991). J Am Vet Med Assoc, 206, 206-209.
• Johnson L, Queen E, Vernau W, Sykes J, Byrne B. (2013) Microbiologic and cytologic assessment of bronchoalveolar lavage fluid from dogs with lower respiratory tract infection: 105 cases (2001–2011). J Vet Intern Med., 27, 259-267.
• Jouini A, Klibi N, Dziri R, Alonso CA, Boudabous A, Slama B, Torres C. (2015) Detection of extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae in vegetables, soil and water of the farm environment in Tunisia. Int J Food Microbiol. 203, 86-92.
• Kim M, Jee H, Shin S, Lee B, Pakhrin B, Yoo H, Yoon J, Kim D. (2007) Outbreak and control of haemorrhagic pneumonia due to Streptococcus equi subspecies zooepidemicus in dogs. Vet Rec. 161, 528-530.
• Kirchner B, Port CD, Magoc T, Sidor M, Ruben Z. (1990) Spontaneous bronchopneumonia in laboratory dogs infected with untyped Mycoplasma spp. Lab Anim Sci. 40, 625-628.
• Lamm C, Ferguson A, Lehenbauer T, Love B. (2010) Streptococcal infection in dogs: a retrospective study of 393 cases. Vet Pathol., 47, 387-395.
• Luna L. (1968) Routine staining procedures. Manual of histologic staining methods of the Armed Forces Institute of Pathology. Blakiston Division McGraw-Hill, New York, 258 p.
• Maden M, Birdane FM, Atkan F, Hadimli HH, Şen İ, Aslan V. (2000) Clinical, cytologic, bacteriologic and radiographic analysis of respiratory diseases in dogs. Eurasian J Vet Sci. 16, 43-50.
• Marques C, Belas A, Aboim C, Cavaco-Silva P, Trigueiro G, Gama LT, Pomba C. (2019) Evidence of sharing of Klebsiella pneumoniae strains between healthy companion animals and cohabiting humans. J Clin Microbiol. 57, e01537-18.
• McAuliffe L, Ellis RJ, Ayling RD, Nicholas RA. (2003) Differentiation of Mycoplasma species by 16S ribosomal DNA PCR and denaturing gradient gel electrophoresis fingerprinting. J Clin Microbiol., 41, 4844-7.
• Navon-Venezia S, Kondratyeva K, Carattoli A. (2017) Klebsiella pneumoniae: a major worldwide source and shuttle for antibiotic resistance. FEMS microbiol Rev. 41, 252-275.
• O’neill E, Day M, Hall E, Holden D, Murphy K, Barr F, Pearson G. (2006) Bacterial cholangitis/cholangiohepatitis with or without concurrent cholecystitis in four dogs. J Small Anim Pract 47, 325-35.
• Pesavento P, Hurley K, Bannasch M, Artiushin S, Timoney J. (2008) A clonal outbreak of acute fatal hemorrhagic pneumonia in intensively housed (shelter) dogs caused by Streptococcus equi subsp. zooepidemicus. Vet Pathol. 45, 51-3.
• Proulx A, Hume DZ, Drobatz KJ, Reineke EL. (2014) In vitro bacterial isolate susceptibility to empirically selected antimicrobials in 111 dogs with bacterial pneumonia. J Vet Emerg Crit Care, 24, 194-200.
• Quinn P, Carter M, Markey B, Carter G. (1999) The streptococci and related cocci. Clin Vet Microbiol.. Mosby, Edinburgh, Scotland, 127-136.
• Radaelli ST, Platt SR. (2002) Bacterial meningoencephalomyelitis in dogs: a retrospective study of 23 cases (1990‐1999). J Vet intern med., 16, 159-163.
• Radhakrishnan A, Drobatz KJ, Culp WT, King LG. (2007) Community-acquired infectious pneumonia in puppies: 65 cases (1993–2002). J Am Vet Med Assoc. 230, 1493-7.
• Roberts DE, McClain HM, Hansen DS, Currin P, Howerth EW. (2000) An outbreak of Klebsiella pneumoniae infection in dogs with severe enteritis and septicemia. J Vet Diagn Invest, 12, 168-73.
• Rosendal S. (1978) Canine mycoplasmas: pathogenicity of mycoplasmas associated with distemper pneumonia. J Infect Dis. 138, 203-210.
• Rosendal S. (1982) Canine mycoplasmas: their ecologic niche and role in disease. J Am Vet Med Assoc. 180, 1212-1214.
• Sayin Z, Sakmanoglu A, Erganis O, Ucan US, Hadimli HH, Aras Z, Sanioglu G, Coskun AA. (2016) Genotypic Characterization of Bordetella bronchiseptica Strains Isolated from Stray and Pet Dogs. Pak Vet J 36, 482-486.
• Schulz B, Kurz S, Weber K, Balzer H-J, Hartmann K. (2014) Detection of respiratory viruses and Bordetella bronchiseptica in dogs with acute respiratory tract infections. Vet J. 201, 365-369.
• Seguin MA, Vaden SL, Altier C, Stone E, Levine JF. (2003) Persistent urinary tract infections and reinfections in 100 dogs (1989–1999). J Vet Intern Med., 17, 622-631.
• Songer JG, Post KW. (2004) Veterinary microbiology-E-book: bacterial and fungal agents of animal disease, Elsevier Health Sciences. 434 p.
• Sykes JE, Kittleson MD, Pesavento PA, Byrne BA, MacDonald KA, Chomel BB. (2006) Evaluation of the relationship between causative organisms and clinical characteristics of infective endocarditis in dogs: 71 cases (1992–2005). J Am Vet Med Assoc. 228, 1723-34.
• Taha-Abdelaziz K, Bassel LL, Harness ML, Clark ME, Register KB, Caswell JL. (2016) Cilia-associated bacteria in fatal Bordetella bronchiseptica pneumonia of dogs and cats. J Vet Diagn Invest. 28, 369-376.
• Tuzcu M, Özmen M, Karakoç SR, Tuzcu N, Yoldaş A. (2012) Diagnosis of mycoplasmosis in chicks by pathological and Real Time-PCR methods. Eurasian J Vet Sci. 28, 82-86.
• Tuzcu M, Tuzcu N, Akcakavak G, Celik Z. (2022) Diagnosis of Sarcina ventriculi-derived haemorrhagic abomasitis in lambs by histopathology and real-time PCR. Acta Vet Brno, 91, 227-233.
• Tuzcu N, Tuzcu M, Akçakavak G, Başbuğ O, Kurul M. (2021) Köpeklerde canine distemper virüs enfeksiyonunun klinik, patolojik ve real-time RT-PCR ile teşhisi. JAVST 6, 57-64
• Tuzcu N, Celik Z, Akcakavak G, Bulut A, Salik R, Oner M, Kaya V, Tuzcu M. (2023) Determination of Helicobacter heilmannii in cats by real time polymerase chain reaction and histopathology. MAE Vet Fak Derg. 8, 69-73.
• Ulstad CR, Solheim M, Berg S, Lindbæk M, Dahle UR, Wester AL. (2016) Carriage of ESBL/AmpC-producing or ciprofloxacin non-susceptible Escherichia coli and Klebsiella spp. in healthy people in Norway. Antimicrob Resist Infect Control, 5, 1-11.
• Vaissaire J, Remond M, Dufrêne M, Laroche M, Le Doujet C, Peigné F, Lebreton F. (1991) Stérilités, avortements et mortinatalité chez la chienne d’élevage dus à Streptococcus spp. du groupe G de Lancefield. Bull Acad Vet Fr. 144, 345-350.
• Yoldaş A, Tuzcu M, Özmen E, Yığın A, Özmen M, Seval K, Tuzcu N. (2009) Sığırlarda Kuduz Hastalığının Teşhisi ve Kuduz Virüsünün Anatomik Lokalizasyonunun Real Time PCR ile BelirlenmesiÜzerine Karşılaştırmalı Çalışmalar. Dicle Üniversitesi Vet Fak Derg, 2, 42-50.