There was no evidence to implicate C. burnetii as an important contributor to abortion or perinatal lamb mortality in 30 primiparous ewe flocks located across southern Australia. The very low C. burnetii seroprevalence was consistent with the absence of detection of C. burnetii in tissues from aborted or stillborn lambs from a subset of farms. These findings are consistent with recent reviews of veterinary laboratory investigations that reported coxiellosis to be an uncommon diagnosis in Australian sheep abortion investigations [5, 6]. Coxiella burnetii control programs such as routine vaccination of breeding ewes are not warranted for sheep farms in southern Australia in the absence of further evidence that coxiellosis is contributing to lamb mortality. Nonetheless, C. burnetii should continue to be included in sheep abortion and perinatal mortality investigation protocols due to the sporadic nature of disease and important zoonotic implications.
This was an observational study with sheep managed extensively, reflecting standard sheep management in these regions of Australia. Although foetal and lamb mortality between scanning and lamb marking were high for some flocks, average lamb mortality in the primiparous flocks was consistent with ranges previously reported in Australian studies . It is unclear from the current study whether reproductive performance of maiden ewes would be impacted in flocks where seroprevalence was greater.
Serological testing targeted primiparous ewes with evidence of abortion and perinatal lamb mortality. Bias towards ewes that failed to rear lambs could be expected to overestimate prevalence in the general sheep population if C. burnetii was an important contributor to abortion and perinatal deaths. Blood samples for primiparous ewe samples were collected close to the time of lambing or abortion, and this may increase probability of detection for C. burnetii seroconversion . Very low seroprevalence in this sampled population suggests that coxiellosis was not an important contributor to abortion and perinatal lamb mortality in these flocks and was consistent with very low seroprevalence in adult ewes on these farms.
This is the most geographically widespread seroprevalence study for C. burnetii in Australian sheep. Very low C. burnetii seroprevalence was consistent with previous studies from Western Australia  and Victoria  that reported individual seroprevalence ranging from 0 - 4.1%, and flock-level seroprevalence ranging from 0 – 17.6%. Our study did not include sheep flocks from New South Wales, Queensland or Tasmania. New South Wales and Queensland have the highest rates of human Q-fever reported in Australia . The most recent studies reporting C. burnetii prevalence in sheep from New South Wales and Queensland are considerably dated and involve either single farms [15, 16] or abattoir surveys . Increased incidence of local acquisition of human infection may be associated with high prevalence in livestock . Hence, investigation of C. burnetii seroprevalence in sheep from New South Wales and Queensland is warranted.
Sampling younger ewes likely contributed to the low C. burnetii seroprevalence reported in this study. Age is recognised as an important risk factor for C. burnetii seropositivity, with older animals more likely to be seropositive [33–35]. Notwithstanding this, no apparent difference in seroprevalence was observed between primiparous ewe lambs (approximately 13 months old at lambing), yearlings (2 years old at lambing) and mature ewes (3 years or older).
There is no reference test for serological diagnosis of coxiellosis, and sensitivity and specificity for C. burnetii serological tests are not well described . The commercial indirect ELISA for C. burnetii that was used in this study has been used in other seroprevalence studies in sheep [25, 36–40] and the World Organisation for Animal Health (OIE) recommends ELISA as the preferred method for C. burnetii seroprevalence studies . In our study, the three samples categorised as seropositive using indirect ELISA were negative by CFT. It was not possible to determine if these were false positives. Complement fixation tests are reported to have lower sensitivity than ELISA, but high specificity for elevated levels of anti-C. burnetii antibodies in flocks with C. burnetii-associated abortions . Discordant results can be observed using different ELISA kits , therefore testing samples with more than one kit is an alternative option for validating animal status . Validation for commercial ELISA in Australian sheep under field conditions could better inform estimation of true prevalence. However, coxiellosis is not frequently diagnosed in Australian sheep which presents challenges for evaluating assay sensitivity and specificity under field conditions.
Seroprevalence surveys may underestimate C. burnetii shedding in livestock. Banazis, Bestall, Reid and Fenwick  detected C. burnetii in Australian sheep faecal samples in the absence of C. burnetii seroconversion. Other studies have also demonstrated poor correlation between seroconversion and antigen detection [10, 12, 36, 43]. Joulié, Rousset, Gasqui, Lepetitcolin, Leblond, Sidi-Boumedine and Jourdain  reported good correlation between high C. burnetii burden on vaginal swabs and seroconversion one-month post-abortion or post-lambing using the same commercial indirect ELISA kit as used in our study. However, it is possible that some ewes in our study were shedding C. burnetii without evidence of seroconversion, and thus represent a reservoir of C. burnetii infection for other sheep or humans. Nevertheless, the combination of testing methodology used and timing of blood sample collection (within 6 weeks of parturition) in conjunction with the absence of detection of C. burnetii using molecular techniques on tissues from aborted or stillborn lambs suggests that coxiellosis was not a major contributor to abortion and lamb mortality observed on these farms.
Despite low seroprevalence detected in this study, contact with sheep should still be considered a risk factor for Q-fever in humans and precautions should be taken to reduce the risk of zoonotic C. burnetii transmission. Sheep have been associated with cases of Q-fever in humans in Australia and overseas [10, 15, 32, 44–48]. Coxiella burnetii shedding can occur from both symptomatic and asymptomatic sheep, and in the absence of detectable seroconversion [49, 50]. Control strategies include use of appropriate personal protective clothing when handling birth material or lambing ewes, good hygiene practices, controlling dust and vaccination of people with an occupational risk including farm, abattoir and veterinary staff.