Molecular typing of Coxiella burnetii from sheep in Egypt
Introduction
Coxiella burnetii is an obligate intracellular bacterium, which causes Q fever disease, and is widely distributed throughout the world. The main reservoir for the pathogen is domestic animals such as cattle, sheep and goats which have a main role in the disease cycle [1].
The diseased animals suffer from several reproductive disorders such as abortion, still birth, and delivery of weak offspring. Q fever is mainly transmitted by inhalation; it could also occur through the consumption of milk or dairy products [2].
According to published data, Q fever seroprevalence, using ELISA, among sheep and goats in Egypt was 22.7% and 12.5%, respectively [3]. While the prevalence of antibodies against C. burnetii in humans who had contacted with animals was 19% [4].
Genetic characterization of C. burnetii is required for epidemiological investigation in Q fever outbreaks and for surveillance purposes. A number of different molecular typing methods have been described to analyze the genetic variability of C. burnetii. Recently, PCR-based methods were developed including multi-locus variable number of tandem repeats analysis (MLVA) and multispacer sequence typing (MST) [5].
While MLVA is based on variable copy number in tandemly repeated DNA on multiple loci in C. burnetii genome, MST is based on DNA sequence variation in 10 short intergenic regions. Both methods can be performed directly on extracted DNA from clinical samples or on C. burnetii isolates [[6], [7], [8]].
As, there is evidence of the presence of Q fever in domestic animals in Egypt [4]. Therefore, the identification of the source of infection and molecular characterization of the circulating C. burnetii strains in Egypt is very important to enhance the wealth of molecular epidemiology data of Q fever in the country.
The aim of this study was to characterize strains of C. burnetii circulating in domestic animals in Egypt, using MLVA and MST genotyping. In addition, the profiles obtained were subsequently compared with other reported genotypes.
Section snippets
Samples
The study comprised one sheep herd which suffered from abortion and located at Alexandria governorate, Egypt. A total of 21 vaginal swabs were collected from aborted cases during the first week after abortion. All samples were transferred to Anses Laboratory, Animal Q fever Unit for molecular identification and characterization.
DNA extraction and PCR assay
Total DNA was extracted from vaginal swabs using the QIAamp Tissue and Blood Kit (Qiagen GmbH, Hilden, Germany) according to the manufacturer’s instructions. To detect
Results
All 21 vaginal swabs were positive with Coxiella specific real-time PCR but only six samples with Ct-value below 34 were analyzed for MLVA genotyping as in Table 1 and only three samples were analyzed using MST genotyping.
The six C. burnetii samples revealed different MLVA pattern in comparison with NM as reference strain based on the 17 MLVA loci used as in Table 1.
Interestingly, all six samples revealed no alleles with ms23 and ms24 markers in comparison to NM. In addition, Eg4, Eg14 and Eg18
Discussion
Molecular characterization of C. burnetii is a powerful tool to explore genetic diversity of strains and determine the relationships between isolates causing disease. Typing data can provide information about the source of infection and the risk of transmission of the infections between animals and human [14,15].
In the case of Q fever, MLVA and MST methods have been used for the typing of C. burnetii strains, both methods have high discriminatory power and can be applied directly on clinical
Conclusion
The present work supports epidemiological information about genetic diversity for C. burnetii occurring in Egypt. Examination of six C. burnetii samples originating from sheep revealed three MST types (completely novel profiles) and four MLVA types. The detected genotypes revealed close relation to C. burnetii isolated from humans. Additional analyses using larger samples from different livestock species and locations are needed to confirm these findings and to construct a reference database of
Declaration of Competing Interest
The authors declare that they have no conflicts of interest associated with this publication.
Acknowledgements
The authors wish to thank the French institute of Egypt and the Academy for Scientific Research and Technology (ASRT) for providing support to complete this work under the “Prevalence, molecular diagnosis of Q fever in Egypt and strain genotyping among ruminants” Imhotep Project.
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