Simultaneous differential detection of Chlamydophila abortus, Chlamydophila pecorum and Coxiella burnetii from aborted ruminant's clinical samples using multiplex PCR

Abstract

BackgroundChlamydiosis and Q fever, two zoonosis, are important causes of ruminants' abortion around the world. They are caused respectively by strictly intracellular and Gram negative bacterium Chlamydophila abortus (Cp. abortus) and Coxiella burnetii (C. burnetii). Chlamydophila pecorum (Cp. pecorum) is commonly isolated from the digestive tract of clinically inconspicuous ruminants but the abortive and zoonotic impact of this bacterium is still unknown because Cp. pecorum is rarely suspected in abortion cases of small ruminants. We have developed a multiplex PCR (m-PCR) for rapid simultaneous differential detection of Cp. abortus, Cp. pecorum and C. burnetii in clinical samples taken from infected animals.ResultsSpecific PCR primers were designed and a sensitive and specific m-PCR was developed to detect simultaneously, in one tube reaction, three specific fragments of 821, 526 and 687-bp long for Cp. abortus, Cp. pecorum and C. burnetii respectively. This m-PCR assay was performed on 253 clinical samples taken from infected ruminant's flocks that have showed problems of abortion diseases. Thus, 67 samples were infected by either one of the three pathogens: 16 (13 vaginal swabs and 3 placentas) were positive for Cp. abortus, 2 were positive for Cp. pecorum (1 vaginal swab and 1 placenta) and 49 samples (33 vaginal swabs, 11 raw milks, 4 faeces and 1 placenta) were positive for C. burnetii. Two vaginal swabs were m-PCR positive of both Cp. abortus and C. burnetii and none of the tested samples was shown to be infected simultaneously with the three pathogens.ConclusionWe have successfully developed a rapid multiplex PCR that can detect and differentiate Cp. abortus, Cp. pecorum and C. burnetii; with a good sensitivity and specificity. The diagnosis of chlamydiosis and Q fever may be greatly simplified and performed at low cost. In addition, the improvement in diagnostic techniques will enhance our knowledge regarding the prevalence and the pathogenetic significance of Q fever and chlamydiosis.

Chlamydia pecorum

“…10 Chlamydia pecorum is commonly detected in the digestive and genital tracts of healthy ruminants; however, it has also been associated with reproductive and systemic disorders in sheep, goats, buffalo, and cattle. 3,10,11,19,21,33 Although the involvement of C. pecorum in small ruminant abortion cases has been previously recognized, 4,[28][29][30] the abortigenic and zoonotic impacts of C. pecorum are still unknown, 4 and there are no reports, to our knowledge, describing fetal lesions caused by this agent in ruminants. In the current work, we describe placental and fetal lesions caused by C. pecorum in a case of sporadic abortion in a goat.…”

“…Molecular methods such as PCR followed by sequencing are more sensitive, specific, and less time-consuming methods for the detection and identification of these bacteria. 3,4,6,14,20,29 Some of the pathological findings in the fetuses examined in our study, such as cartilaginous metaplasia of the placenta in fetus A, and brachygnathia and palatoschisis in fetus B, cannot be explained by C. pecorum infection alone. Because brachygnathia has been described in aborted ruminants with orthobunyavirus infection, 15 we tested fetus B for the detection of bunyaviruses by PCR, with negative results.…”

Prevalence of Coxiella Burnetii in Dairy Herds - Diagnostic Methods and Risk to Humans - A Review

“…has been observed (23). Molecular methods such as PCR have been developed for detection of an individual pathogen or duplex PCR for simultaneous detection of C. abortus, C. pecorum and C. burnetii (4).…”

Chlamydial Abortion in Algerian Small Ruminants

“…Members of the genus Chlamydia are gram-negative bacteria and obligate intracellular bacterial parasites responsible for a variety of diseases in humans and animals, including several zoonoses (Pannekoek et al, 2010). The reservoir is large and includes many wild and domestic mammals but domestic ruminants such as sheep, cattle and goat represent the most frequent source of human infection (Berri et al, 2009).…”

“…Chlamydia pecorum strains were isolated from abortion cases of goat and sheep (unpublished data) suggesting that this bacterium might be involved in small ruminants abortion in North African countries (Berri et al, 2009). Stillbirth and infertility, leading to important economic losses, are also associated t…”