Detection of Neospora caninum in aborted bovine embryos using Histopathology, ELISA, PCR and Real time PCR

Document Type : Full Research Paper

Authors

1 pathology department , Tehran university,Tehran,Iran

2 pathology department , Tehran university,Tehran, Iran

3 Iran veterinary Organization

Abstract

Background:Bovine neosporosis is caused by Neosporacaninum, A parasite that causes abortion is often the only observed sign of disease.
Objective: The aim of this study was to investigate the prevalence of Neospora caninum in some Alborz and Qazvin farms.
Methods: in this reaserch 105 samples from bovine abortions were sent to the laboratory, from January 2018 to May 2019. Samples were screened for N. caninum in the brain, heart, kidney, liver, lung, spleen, and placenta, using HistoPatology, ELISA, conventional, PCR and Real Time PCR.. Elisa was used as an initial screen for indication of infectious abortion and the second was histopathology and PCR 
Results: A mononuclear inflammatory infiltrate was predominant in the heart, kidney, liver, lung, brain, and placenta. 
N. caninum causes mononuclear inflammatory infiltrate in various organs and focal necrosis with mononuclear infiltrate in the brain, Non-suppurative lesions were also present in the lung, liver, kidney, and placenta, indicating that N. caninum infection is systemic. The rate of fetal infection with N. caninum, represented separately by each of the diagnostic techniques, was 8/6% with Elisa, 20/9% with Real Time PCR and 6/7% with conventional -PCR.
 Conclusions :The diagnosis of abortion due to N. caninum requires detection of the parasite through techniques that are specific, such as Histopathology, or PCR that are both specific and more sensitive, preferably associated with identification of histopathological lesions placenta and, importantly, fetal brain, should be encouraged to increase the success of the definitive diagnosis of neosporosis by tests of conventionalPCR and Real Time PCR.

Keywords

References

1- Alvarez-García, G., Culebras, A.G., Expósito, D.G., Navarro-Lozano,V., Pastor-Fernández, I., Ortega-Mora, L.M. (2013). Serological diagnosis of bovine neosporosis: a comparative study of commercially available ELISA tests, Vet Parasitol, 198(1-2), 85-95. https://doi.org/10.1016/j.vetpar.2013.07.033 PMID: 23953144.
2- Anderson, M.L., Blanchard, P.C., Barr, B.C., Dubey, J.P., Hoffman, R.L., Conrad, P.A. (1991). Neospora-like protozoan infection as a major cause of abortion in California dairy cattle, J Am Vet Med Assoc,198(2):241-4. https://doi.org/.
3- Barratt, J., Al Qassab, S.,  Reichel, M.P., Ellis J.T. (2008). The development and evaluation of a nested PCR assay for detection of Neospora caninum and Hammondia heydorni in feral mouse tissues, https://doi.org/10.1016/j.mcp.2008.03.001 PMID: 18420378.
4-Baszler, T.V.,  Gay, L.J.,  Long, M.T  Mathison, B.A. (1999). Detection by PCR of Neospora caninum in fetal tissues from spontaneous bovine abortions, J Clin Microbiol (12):4059-64. https://doi.org/10.1128/JCM.37.12.4059-4064.1999 PMID: 10565932.
5- Bjerkås, I.,  Mohn, S. F.,  Presthus, J. (1984). Unidentified cyst-forming sporozoon causing encephalomyelitis and myositis in dogs, Z Parasitenkd.,70(2):271-4. https://doi.org/10.1007/bf00942230  PMID: 6426185.
6-BOENISCH, T. (2005). Effect of heat-induced antigen retrieval following inconsistent formalin fixation, Appl Immunohistochem Mol Morphol(3):283-6 https://doi.org/10.1097/01.0000146524.74402.a4  PMID: 16082257.
7-Collantes-Fernández , E.,  Zaballos, A.,  Alvarez-García, G., Luis M Ortega-Mora, L. M. Quantitative detection of Neospora caninum in bovine aborted fetuses and experimentally infected mice by real-time PCR. (2002). J Clin Microbiol.,40(4):1194-8. https://doi.org/10.1128/JCM.40.4.1194-1198.2002 PMID: 11923330.
8-  Corbellini , L. G., Driemeier, D.,  Cruz, C. F. E.,  Gondim, L. F. P., Wald, V. (2002). Neosporosis as a cause of abortion in dairy cattle in Rio Grande do Sul, southern Brazil, Vet Parasitol.,103(3):195-202. https://doi.org/10.1016/s0304-4017(01)00600-8  PMID: 11750112.
9-De Craeye , S., Speybroeck, N., Ajzenberg, D.,  Dardé, M. L., Collinet, F., Tavernier, P., Van Gucht, S., Dorny, P., Dierick, K. (2011). Toxoplasma gondii and Neospora caninum in wildlife: Common parasites in Belgian foxes and Cervidae, Vet Parasitol.,178(1-2):64-9 https://doi.org/10.1016/j.vetpar.2010.12.016PMID: 21236577.
10-Dubey , J. P. (2003). Review of Neospora caninum and neosporosis in animals, Korean J Parasitol.,41(1):1-16. https://doi.org/10.3347/kjp.2003.41.1.1 PMID: 12666725.
11-DUBE, J. P., Carpenter, J. L.,  Speer, C. A.,  Topper, M. J., Uggla, A. (1988(. Newly recognized fatal protozoan disease of dogs. J Am Vet Med Assoc.,192(9):1269-85. PMID: 3391851.
12- Dubey, J. P.,  Lindsay, D. S. )1996(. A review of Neospora caninum and neosporosis, Vet Parasitol.,67(1-2):1-59 .https://doi.org/10.1016/s0304-4017(96)01035-7 PMID: 9011014.
13-  Dubey, J. P.,  Lindsay, D.S.,  Speer, C. A. (1998). Structures of Toxoplasma gondii tachyzoites, bradyzoites and sporozoites and biology and development of tissue cysts. Clin Microbiol Rev.,11(2):267-99.
https://doi.org/ 10.1128/CMR.11.2.267  PMID: 9564564
14- Dubey , J. P., Schares, G.,  Ortega-Mora L. M. (2007(. Epidemiology and control of neosporosis and Neospora caninum, Clin Microbiol Rev.,20(2):323-67. https://doi.org/10.1128/CMR.00031-06 PMID: 17428888.
15- Eperon , S., Brönnimann, K.,  Hemphill, A.,  Gottstein, B. (1999).  Susceptibility of B-cell deficient C57BL/6 (microMT) mice to Neospora caninum infection. Parasite Immunol (5):225-36. https://doi.org/10.1046/j.1365-3024.1999.00223.x PMID: 10320620.
16-  Gharekhani ,J., Yakhchali,M.(2010).  Neospora caninum infection in dairy farms with history of abortion in West of Iran. Vet Anim Sci; 8:100071. https://doi.org/10.1016/j.vas.2019.100071 PMID: 32734088.
17-  Gondim, L. F.,  Sartor, I. F.,  Monteiro, J.r.,  Haritani, M.(1999). Neospora caninum infection in an aborted bovine foetus in Brazil. N Z Vet J.,47(1):35. https://doi.org/10.1080/00480169.1999.36106 PMID: 16032066.
18-  Gondim , L. F.P.,  Laski, P.,  Gao, L.,  McAllister, M. M.(2004). Variation of the internal transcribed spacer 1 sequence within individual strains and among different strains of Neospora caninum. J Parasitol;90(1):119-22.  https://doi.org/10.1645/ge-134r PMID: 15040677.
19- Gondim , L. F.P., McAllister, M. M., Pitt, W. C.,  Zemlicka, D. E.(2004).  Coyotes (Canis latrans) are definitive hosts of Neospora caninum. Int J Parasitol;34(2):159-61. https://doi.org/10.1016/j.ijpara.2004.01.001 PMID: 15037103.
20- Gottstein , B., Hentrich, B., Wyss, R.,  Thür, B., Busato, A.,  Stärk, K. D.,  Müller, N.(1998).   Molecular and immunodiagnostic investigations on bovine neosporosis in Switzerland. Int J Parasitol, 28(4):679-91. https://doi.org/10.1016/s0020-7519(98)00006-x  PMID: 9602392.
21-  Holmdahl , O. J., Mattsson, J.G.(1996).  Rapid and sensitive identification of Neospora caninum by in vitro amplification of the internal transcribed spacer 1. Parasitology,112 ( Pt 2):177-82. https://doi.org/10.1017/s0031182000084742 PMID: 8851857.
22-  Hughes , J. M.,  Williams, R. H.,  Morley, E. K.,  Cook, D .A .N.,  Terry, R .S.,  Murphy, R .G.,  Smith, J. E., Hide, G.(2006). The prevalence of Neospora caninum and co-infection with Toxoplasma gondii by PCR analysis in naturally occurring mammal populations. Parasitology. 132(Pt 1):29-36. https://doi.org/10.1017/s0031182005008784 PMID: 16393351.
23-KEY, M.., BOENISCH, T. Antigen Retrieval. In: KEY, M. (ed.). Immunohistochemical staining methods. 4 ed. Carpinteria: Dako Corporation, 2006. p. 41-45.   
24-Kirkbride, C. A. Laboratory diagnosis of livestock abortion. 3 ed. Iowa State: University Press, 1990. 260 p.       
25-  Lindsay , D. S., Dubey J. P. (1989). Immunohistochemical diagnosis of Neospora caninum in tissue sections. Am J Vet Res;50(11):1981-3. PMID: 2694869.
26-  McAllister , M. M.,  Dubey, J. P.,  Lindsay, D. S., W R Jolley, W. R.,  Wills, R. A.,  McGuire A. M. (1998). Dogs are definitive hosts of Neospora caninum, Int J Parasitol;28(9):1473-8. PMID: 9770635.
27- Medina , L., Cruz-Vázquez, C ., Quezada, T., Morales, E., García-Vázquez, Z. (2006). Survey of Neospora caninum infection by nested PCR in aborted fetuses from dairy farms in Aguascalientes, Mexico, Vet Parasitol;136(3-4):187-91. https://doi.org/10.1016/j.vetpar.2005.11.003 PMID: 16332413.
28-  McNamee , P. T.,  Trees, A. J., Guy, F., Moffett, D., Kilpatrick. D. (1996). The diagnosis and prevalence of neosporosis in Northern Ireland cattle, Vet Rec;138(17):419-20  https://doi.org/10.1136/vr.138.17.419 PMID: 8733183.
29-Moayer, F., Ataie, O., Mosa Khani, F., Bahonar, A. (2011). Pathologic findings of aborted fetuses in dairy farms around Tehran,  JVCR  V:2, N:3(7):155-166.
30-  Monteiro , R. M.,  Richtzenhain, L. J.,  Pena, H. F. J.,  Souza, S. L. P.,  Funada, M. R.,  Gennari, S. M.,  Dubey, J. P., Sreekumar, C.,  Keid, L. B.,  Soares, R. M. )2007(. Molecular phylogenetic analysis in Hammondia-like organisms based on partial Hsp70 coding sequences, Parasitology;134(Pt 9):1195-203 https://doi.org/10.1017/s0031182007002612 PMID: 17462122.
31-  Moore , D. P., Regidor-Cerrillo, J., Morrell, E.,  Poso, M. A.,  Cano, D. B.,  Leunda, M. R., Linschinky, L.,  Odeón, A. C., Odriozola, E.,  Ortega-Mora, L. M.,  Campero, C. M. )2008(. The role of Neospora caninum and Toxoplasma gondii in spontaneous bovine abortion in Argentina, Vet Parasitol;156(3-4):163-7 https://doi.org/10.1016/j.vetpar.2008.06.020 PMID: 18691819.
32- Müller , N., Zimmermann, V., Hentrich, B., Gottstein B. )1996(. Diagnosis of Neospora caninum and Toxoplasma gondii Infection by PCR and DNA Hybridization Immunoassay, J Clin Microbiol;34(11):2850-2 https://doi.org/10.1128/jcm.34.11.2850-2852.1996 PMID: 8897199.
33- Müller , N., Vonlaufen, N., Gianinazzi, C.,  Leib, S. L., Hemphill A. )2002(. Application of Real-Time Fluorescent PCR for Quantitative Assessment of Neospora caninum Infections in Organotypic Slice Cultures of Rat Central Nervous System Tissue, J Clin Microbiol;40(1):252-5 https://doi.org/10.1128/jcm.40.1.252-255.2002 PMID: 11773124.
34- Otter , A., M Jeffrey, M.,  Griffiths, I. B.,  Dubey J. P. (1995). A survey of the incidence of Neospora caninum infection in aborted and stillborn bovine fetuses in England and Wales, Vet Rec;136(24):602-6. https://doi.org/10.1136/vr.136.24.602 PMID: 7571263.
35-  Paula , V. S. O.,  Rodrigues, A. A. R.,  Richtzenhain, L. J., Cortez, A., Soares, R. M.  Gennari, S. M. (2004). Evaluation of a PCR based on primers to Nc5 gene for the detection of Neospora caninum in brain tissues of bovine aborted fetuses, Vet Res Commun;28(7):581-5 https://doi.org/10.1023/b:verc.0000042877.07684.89 PMID: 15563105.
36- Payne , S., Ellis, J. (1996). Detection of Neospora caninum DNA by the polymerase chain reaction. .) Int. J. Parasitol, Int J Parasitol;26(4):347-51  https://doi.org/10.1016/0020-7519(96)00030-6 PMID: 8773521.
37- Pereira-Bueno, J., Quintanilla-Gozalo, A., Pérez-Pérez, V., Espi-Felgueroso, A., Alvarez-García, G., Collantes-Fernández, E.,  Ortega-Mora L. M. )2003). Evaluation by different diagnostic techniques of bovine abortion associated with Neospora caninum in Spain. Vet Parasitol;111(2-3):143-52. https://doi.org/10.1016/s0304-4017(02)00361-8 PMID: 12531290.
38- Pescador , C. A.,  Corbellini, L. G.,  Oliveira, E. C.,  Raymundo, D. L., Driemeier, D. (2007). Histopathological and immunohistochemical aspects of Neospora caninum diagnosis in bovine aborted fetuses, Vet Parasitol;150(1-2):159-63 https://doi.org/10.1016/j.vetpar.2007.08.028  PMID: 17904290.
39- Ramos-Vara , J.,  Kiupel, M.,  Baszler, T.,  Bliven, Laura  Brodersen, B.,  Chelack, B., Czub, S.,  Del Piero, F.,  Dial, S.,  Ehrhart, E. J., Graham, T.,  Manning, L., Paulsen, D.,  Valli, V. E.,  West, K. (2008). Suggested guidelines for immunohistochemical techniques in veterinary diagnostic laboratories, J Vet Diagn Invest ,20(4):393-413. https://doi.org/10.1177/104063870802000401 PMID: 18599844.
40-  Sager , H., Fischer, I., Furrer, K., Strasser, M., Waldvogel, A., Boerlin, P., Audigé, L., Gottstein B.  (2001).  A Swiss case-control study to assess Neospora caninum-associated bovine abortions by PCR, histopathology and serology, Vet Parasitol;102(1-2):1-15  https://doi.org/10.1016/s0304-4017(01)00524-6  PMID: 11705647.
41- Schares , G., Peters, M., Wurm, R., Bärwald, A.,  Conraths F. J. (1998). The efficiency of vertical transmission of Neospora caninum in dairy cattle analysed by serological techniques, Vet Parasitol;80(2):87-98 https://doi.org/10.1016/s0304-4017(98)00195-2  PMID: 9870361.
42-Söndgen , P., Peters, M., Bärwald, A., Wurm, R., Holling, F.,  Conraths, F. J., Schares G. (2001). Bovine neosporosis: immunoblot improves foetal serology, Vet Parasitol;102(4):279-90  https://doi.org/10.1016/s0304-4017(01)00543-x PMID: 11731071.
43-Wouda , W.,  Moen, A. R.,  Visser, I. J., van Knapen F. (1997). Bovine fetal neosporosis: a comparison of epizootic and sporadic abortion cases and different age classes with regard to lesion severity and immunohistochemical identification of organisms in brain, heart, and liver, J Vet Diagn Invest;9(2):180-5 https://doi.org/10.1177/104063879700900212 PMID: 9211238.
44- Yamage , M., Flechtner, O., Gottstein B. (1996). Neospora caninum: specific oligonucleotide primers for the detection of brain "cyst" DNA of experimentally infected nude mice by the polymerase chain reaction (PCR), PMID: 8604096.
 
Veterinary Research & Biological Products
Volume 35, Issue 3 - Serial Number 3
September 2022
Pages 62-70

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  • Receive Date: 25 September 2021
  • Accept Date: 07 October 2021

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