A Comparative Study on Detection of Bartonella henselae Infection by Culture Followed by PCR, Nested-PCR and IFA [1]

Kediler kedi tırmalama hastalığının etkeni olan zoonoz Bartonella henselae bakterisinin ana rezervuarıdır. Bu çalışmanın amacı tam kandan kültür sonrası PCR, tam kan ve oral svaptan nested-PCR ve serum örneklerinden IFA yöntemlerini içeren 3 farklı teşhis yöntemini karşılaştırmaktır. B. henselae tanısı için İstanbul, Türkiye'de yaşayan 81 ev ve sokak kedilerinden toplanan kan örneklerinden bakteriyolojik kültürü takiben yapılan konvansiyonel PCR, tam kan örnekleri ve ağız boşluğundan alınan svapların 2 farklı nested PCR'I ile karşılaştırıldı. Ayrıca aynı hayvanlarda indirect floresan antikor (IFA) tekniği ile seroprevalance belirlendi. Kültür sonucunda 26 (32%) kan örneğinde Bartonella spp saptandı. Konvansiyonel PCR testleri sonucunda bunların 20 adeti B. henselae, altı adeti B. clarridgeiae olarak identifiye edildi. Nested PCR sonucu, 81 tam kan örneğinin 29'u (36%) pozitifti. Bunların 20 adeti B. henselae, sekiz adeti B. clarridgeiae olarak identifiye edildi. Aynı zamanda, 1 örnekte hem B. henselae hem de B. clarridgeiae identifiye edildi. PCR'ı yapılan 81 oral svabın 25'i (31%) nested PCR ile pozitif bulundu. Bunların 19 adeti B. henselae, altı adeti B. clarridgeiae olarak identifiye edildi. B.henselae'ya karşı oluşmuş IgG antikorlarının varlığı incelenen serum örneklerinde seroprevalans %67 (54/81) olarak belirlendi. Sonuç olarak kan ve oral svap örneklerinden Nested-PCR kombinasyonunun kullanımı testlerin sensitivitesini arttırabilmektedir. Ayrıca kan kültürünün nested-PCR ve seroloji ile kombinasyonu kedilerde bartonellosis tanısında muhtemelen en kesin bilgiyi vermektedir

Bartonella henselae Infeksiyonunun Saptanmasında Kültür Sonrası PCR, Nested-PCR ve IFA Yöntemlerinin Karşılaştırılması

Cats are the main reservoirs of zoonotic Bartonella henselae which are the causative agents of Cat Scratch Disease (CSD). The aim of this study is to compare three diagnostic methods including culture followed by PCR from whole blood, nested-PCR from oral swab and whole blood, and IFA from serum samples. The diagnosis of B. henselae was compared with the bacteriological methods following conventional PCR and by two separate nested PCR from blood, and oral cavity swabs which were collected from 81 pet and stray cats in Istanbul, Turkey. Also the seroprevalence was determined by indirect fluorescent antibody (IFA) technique in the same animals. Bartonella spp. was determined in 26 (32%) of the blood samples by culture. Twenty of them were identified as B. henselae and 6 of them were B. clarridgeiae by following conventional PCR assay. Of 81 whole blood samples subjected to PCR, 29 (36%) were positive in the nested reaction. Of these, 20 were identified as B.henselae and 8 were B. clarridgeiae. However, one of the samples was found to be positive for both B. henselae and B. clarridgeiae DNA by the nested reaction. Of 81oral swab samples subjected to PCR, 25 (31%) were positive in the nested reaction. Of these, 19 were identified as B. henselae and 6 were B. clarridgeiae. B.henselae IgG antibody seroprevalence was detected as 67% (54/81). Using the combination of blood and oral samples by Nested-PCR simultaneously may increase the sensitivity of the test. Also, the combination of the blood culture with nested- PCR and serology is likely to give the most definitive information in the diagnosis of bartonellosis in cats

PDF

___

1. Kordick DL, Breitschwerdt EB: Intraerythrocytic presence of Bartonella henselae. J Clin Microbiol, 33, 1655-1656, 1995.
2. Năsoiu L, Mitrea I, Ioniță M: Advanced diagnostic methods as tools to investigate the exposure to bartonella infections in cats. Proc Rom Acad (Series B, Suppl. 1): 145-150, 2015.
3. Sander A, Buhler C, Pelz K, von Cramm E, Bredt W: Detection and identification of two Bartonella henselae variants in domestic cats in Germany. J Clin Microbiol, 35, 584-587, 1997.
4. Zanutto MdS, Mamizuka EM, Raiz-Junior R, Diogo CL, Okay TS, Hagiwara MK: Experimental infection and horizontal transmission of Bartonella henselae in domestic cats. Rev Inst Med Trop São Paulo, 43, 257- 261, 2001. DOI: 10.1590/S0036-46652001000500004
5. Koehler JE, Quinn FD, Berger TG, LeBoit PE, Tappero JW: Isolation of Rochalimaea species from cutaneous and osseous lesions of bacillary angiomatosis. N Engl J Med, 327, 1625-1631, 1992. DOI: 10.1056/ NEJM199212033272303
6. Jensen WA, Fall MZ, Rooney J, Kordick DL, Breitschwerdt EB: Rapid identification and differentiation of Bartonella species using a single-step PCR assay. J Clin Microbiol, 38, 1717-1722, 2000.
7. Rampersad JN, Watkins JD, Samlal MS, Deonanan R, Ramsubeik S, Ammons DR: A nested-PCR with an internal amplification control for the detection and differentiation of Bartonella henselae and B. clarridgeiae: an examination of cats in Trinidad. BMC Infect Dis, 5, 63, 2005. DOI: 10.1186/1471-2334-5-63
8. Jekel JF, Katz DL, Elmore JG, Wild D: Epidemiology, biostatistics and preventive medicine. Elsevier Health Sciences, Philadelphia, 2007.
9. Fabbi M, De Giuli L, Tranquillo M, Bragoni R, Casiraghi M, Genchi C: Prevalence of Bartonella henselae in Italian stray cats: evaluation of serology to assess the risk of transmission of Bartonella to humans. J Clin Microbiol, 42, 264-268, 2004. DOI: 10.1128/JCM.42.1.264-268.2004
10. Pennisi MG, Marsilio F, Hartmann K, Lloret A, Addie D, Belak S, Boucraut-Baralon C, Egberink H, Frymus T, Gruffydd-Jones T, Hosie MJ, Lutz H, Möstl K, Radford AD, Thiry E, Truyen U, Horzinek MC: Bartonella species infection in cats: ABCD guidelines on prevention and management. J Feline Med Surg, 15, 563-569, 2013. DOI: 10.1177/ 1098612X13489214
11. Engvall EO, Brandstrom B, Fermer C, Blomqvist G, Englund L: Prevalence of Bartonella henselae in young, healthy cats in Sweden. Vet Rec, 152, 366-369, 2003. DOI: 10.1136/vr.152.12.366
12. Bai Y, Rizzo MF, Alvarez D, Moran D, Peruski LF, Kosoy M: Coexistence of Bartonella henselae and B-clarridgeiae in populations of cats and their fleas in Guatemala. J Vector Ecol, 40, 327-332, 2015. DOI: 10.1111/ jvec.12171
13. Pennisi MG, La Camera E, Giacobbe L, Orlandella BM, Lentini V, Zummo S, Fera MT: Molecular detection of Bartonella henselae and Bartonella clarridgeiae in clinical samples of pet cats from Southern Italy. Res Vet Sci, 88, 379-384, 2010. DOI: 10.1016/j.rvsc.2009.11.005
14. Kim YS, Seo KW, Lee JH, Choi EW, Lee HW, Hwang CY, Shin NS, Youn HJ, Youn HY: Prevalence of Bartonella henselae and Bartonella clarridgeiae in cats and dogs in Korea. J Vet Sci, 10, 85-87, 2009. DOI: 10.4142/jvs.2009.10.1.85
15. Quimby JM, Elston T, Hawley J, Brewer M, Miller A, Lappin MR: Evaluation of the association of Bartonella species, feline herpesvirus 1, feline calicivirus, feline leukemia virus and feline immunodeficiency virus with chronic feline gingivostomatitis. J Feline Med Surg, 10, 66-72, 2008. DOI: 10.1016/j.jfms.2007.05.007
16. Namekata DY, Kasten RW, Boman DA, Straub MH, SipersteinCook L, Couvelaire K, Chomel BB: Oral shedding of Bartonella in cats: Correlation with bacteremia and seropositivity. Vet Microbiol, 146, 371- 375, 2010. DOI: 10.1016/j.vetmic.2010.05.034
17. Ficociello J, Bradbury C, Morris A, Lappin MR: Detection of Bartonella henselae IgM in serum of experimentally infected and naturally exposed cats. J Vet Intern Med, 25, 1264-1269, 2011. DOI: 10.1111/j.1939- 1676.2011.00820.x
18. Park HJ, Lee SE, Hong SH, Lee WJ, Seo KW, Song KH: Seroprevalence of Toxoplasma gondii and Bartonella henselase infection in stray cats of the Daejeon City, Korea. Korean J Vet Res, 54, 87-89, 2014. DOI: 10.14405/ kjvr.2014.54.2.87
19. Lappin MR, Breitschwerdt E, Brewer M, Hawley J, Hegarty B, Radecki S: Prevalence of Bartonella species antibodies and Bartonella species DNA in the blood of cats with and without fever. J Feline Med Surg, 11, 141-148, 2009. DOI: 10.1016/j.jfms.2008.06.005