Comparison of Light Microscopy and Polymerase Chain Reaction for the Detection of Haemoparasites in Cattle in Nigeria.

Abstract

Haemoparasitic diseases are among the important factors that threaten cattle health and productivity especially in the sub-Saharan region. In Nigeria, their detection using sensitive molecular techniques is scanty. This study was designed to investigate and to reevaluate the repertoire of haemoparasites of cattle in Ibadan, Nigeria with a comparative evaluation of light microscopy (LM) and polymerase chain reaction (PCR) methods. Blood samples from 100 cattle slaughtered at Ibadan abattoirs were examined using LM and PCR techniques for haemoparasite detection. The PCR reactions using three primer sets targeting the 16S rRNA genes for Hemoplasma spp. and Anaplasma/Ehrlichia spp. and 18S rRNA genes of Babesia/Theleiria spp. were done. A few randomly selected amplicons from each set were sequenced and analysed. A total infection rate of 34% by LM including Hemoplasma spp. (17%), Anaplasma spp. (16%), microfilaria (5%) and Trypanosoma spp. (12%) was recorded. While, 86% positivity was recorded with PCR amplification as follows: Hemoplasma spp. (64%), Babesia/Theleiria spp. (46%) and Anaplasma/Ehrlichia spp. (5%). Comparison of LM and PCR findings showed that no LM Anaplasma spp.-positive samples and 7 out of the 17 LM hemoplasma-positive cattle were confirmed by PCR. In addition, LM led to misdiagnosis of 46 Babesia/Theleiria spp.-positive samples. Amplicon sequencing and phylogenetic analysis of Babesia/Theileria spp.-positive samples revealed Theileria velifera and Theileria annulata. In the Anaplasma/Ehrlichia spp.-positive samples, only Anaplasma marginale was characterized. Mycoplasma wenyonii, "Candidatus Mycoplasma haemobos" and Pseudomonas fluorescens like were characterized among the hemoplasma-infected cattle. The first report of "Candidatus Mycoplasma haemobos" and Pseudomonas fluorescens like in Nigerian cattle is herewith documented. The alarming LM misdiagnosis of haemoparasites during this study confirms its limitations as it fails to identify many parasites and emphasizes the need for inclusion of molecular techniques to improve their detection. The study also shows for the first time the high prevalence of haemotropic mycoplasma in Nigerian cattle via molecular diagnostic methods, thus indicating a strong need for the investigation of their zoonotic implications.