The discovery of antibodies against Coxiella burnetii in cattery-confined breeding cats indicating prior or current exposure (Shapiro et al., 2015) prompted an investigation into possible sources of infection. One hypothesis was that raw meat diets containing reservoir species may provide a source of C. burnetii transmission. The aim of this pilot study was to determine whether C. burnetii DNA was present in raw meat sold exclusively for companion animal consumption. The sample population consisted of raw meat packages (n = 58) of primarily kangaroo origin, with three to four aliquots (50-120 mg) randomly selected from each package. Genomic DNA was extracted from whole tissue in each of these aliquots using a modified protocol.Three quantitative PCR assays were used for the detection of C. burnetii targeting the IS1111 gene, the heat shock operon htpAB and the C. burnetii outer membrane protein-coding gene, com1. Coxiella burnetii DNA was detected in 25/58 samples (43%) using the IS1111, htpAB and/or com1 PCR assays and confirmed by DNA sequencing.All samples amplifying a product in the com1 assay also amplified a product in the htpAB and IS1111 assays. A total of 17/58 (29%) packets were positive with all three genes, 4/58 (7%) were positive with two genes (IS1111 and htpAB) and 4/58 (7%) were positive with the IS1111 gene only. Coxiella burnetii DNA was five times more likely to be found in offal than skeletal muscle meat samples. All meat samples in which C. burnetii DNA was found were from kangaroo tissues, while samples labelled as non-kangaroo meat (n = 4) were negative. Multi-locus variable number of tandem repeat analysis (MLVA) identified three different genotypes of C. burnetii that have all been identified previously from Australian human clinical Q fever cases. Further investigations are required to determine the potential role of certain raw meats in the transmission of C. burnetii to cats and humans. K E Y W O R D Scats and dogs, Coxiella burnetii, kangaroo, pet food, Q fever, quantitative Polymerase Chain Reaction (qPCR), raw meat

The Australian fur seal (Arctocephalus pusillus doriferus) has experienced a slower than expected population recovery since the end of the commercial sealing era, with a high third trimester abortion rate. There is currently no known proximate cause. Coxiella burnetii (Cb) is a well-known cause of abortion in domestic and wildlife species and an important zoonotic pathogen. It has been recorded from a small number of northern hemisphere marine mammals and may be a potential contributory factor to decreasing populations of northern fur seals (Callorhinus ursinus) and Steller sealions (Eumetopias jubatus). It has not been recorded from marine mammals in the southern hemisphere but is well documented in ruminants and wildlife in Australia as a cause of reproductive failure. Third trimester aborted fetuses (n = 46) and full-term placentas (n = 66) from Australian fur seals, were collected on Kanowna Island and Seal Rocks in Bass Strait, south-eastern Australia. Utilizing routine hematoxylin and eosin histopathology, Cb immunohistochemistry and two different qPCR targets–htpAB and com1, Cb was identified. Routine histopathology and immunohistochemistry were insensitive for the detection of Cb. The detected Cb prevalence ranged from 10.6% for com1 up to 40.9% with htpAB. Coxiella burnetti was readily detected in full-term placentas but in aborted fetal material only in a single placenta associated with a still birth. The exact significance is currently unclear, but this highlights that Cb is present in Australian fur seals, breeding in Bass Strait. Bass Strait is in one of the world’s fastest warming oceanic regions and marine mammals breeding in the area are likely to be key indicators of marine ecosystem stressors. This first description of Cb in a marine mammal from the southern hemisphere, highlights the need to further investigate the potential risks this pathogen poses to Australian fur seals and sympatric marine mammals. Additionally, it is important to determine the zoonotic risk of this pathogen to persons working with, and in proximity of, Australian fur seal breeding colonies.

Coxiella burnetii is suspected as a novel pathogen contributing to decreased pup production in Australian fur seals (Arctocephalus pusillus doriferus). It has recently been described from a single breeding colony in Bass Strait, has previously been associated with two decreasing populations of northern hemisphere pinnipeds and is a known reproductive pathogen. Data around its disease ecology in marine mammals are sparse. Aims. To determine whether environmental DNA (eDNA) can be used to survey for C. burnetii in Australian fur seal breeding colonies. To determine whether C. burnetii in Australian fur seals is the same genotype as terrestrial Australian C. burnetii. Methods. Soil samples were collected from Kanowna Island and Seal Rocks. Placental samples were collected from Kanowna Island. Soil was evaluated for eDNA using a quantitative polymerase chain reaction (qPCR) for com1 gene. Placental samples were evaluated with com1, htpAB and IS1111 markers. Multiple-locus variable number of tandem repeats analysis for three microsatellite loci (ms-24, ms-28 and ms-33) was used to determine relatedness to Australian C. burnetii genotypes. Key results. eDNA results varied between preand post-pupping at Seal Rocks. When targeting the com1 gene, the post-pupping prevalence at Kanowna Island and Seal Rocks was 59.6% and 90%, respectively. eDNA PCR inhibition of samples was low at 1.9%. There was very poor, sporadic to absent IS1111 amplification in placental samples. The com1 and htpAB qPCRs had an overall prevalence across placental samples of 39.2% and 56.7% respectively. In 90.1% of placental samples (n = 11), the ms-28 locus amplified. Neither ms-24 nor ms-33 amplified. Conclusions. eDNA is an effective tool to survey Australian fur seal breeding colonies in the post-pupping period for C. burnetii. The prevalence appears to be much higher in the Seal Rocks colony than in the Kanowna Island colony. It appears that this is not a terrestrial Australian genotype but rather closely related to genotypes detected in marine mammals in the northern hemisphere. Implications. This research significantly expands our ability to survey for C. burnetii in Australian fur seals and other marine mammals. It highlights knowledge gaps in our understanding of the disease ecology and phylogeny of C. burnetii in marine mammals.

Tick-borne infectious diseases caused by obligate intracellular bacteria of the genus Rickettsia are a growing global problem to human and animal health. Surveillance of these pathogens at the wildlife interface is critical to informing public health strategies to limit their impact. In Australia, reptile-associated ticks such as Bothriocroton hydrosauri are the reservoirs for Rickettsia honei, the causative agent of Flinders Island spotted fever. In an effort to gain further insight into the potential for reptile-associated ticks to act as reservoirs for rickettsial infection, Rickettsia-specific PCR screening was performed on 64 Ambylomma albolimbatum ticks taken from shingleback skinks (Tiliqua rugosa) located in southern Western Australia. PCR screening revealed 92% positivity for rickettsial DNA. PCR amplification and sequencing of phylogenetically informative rickettsial genes (ompA, ompB, gltA, sca4, and 17kda) suggested that the single rickettsial genotype detected represented a novel rickettsial species, genetically distinct from but closely related to Rickettsia gravesii and within the rickettsia spotted fever group (SFG). On the basis of this study and previous investigations, it would appear that Rickettsia spp. are endemic to reptile-associated tick species in Australia, with geographically distinct populations of the same tick species harboring genetically distinct SFG Rickettsia species. Further molecular epidemiology studies are required to understand the relationship between these diverse Rickettsiae and their tick hosts and the risk that they may pose to human and animal health.

Tick bites in Australia are linked to the transmission of a variety of infectious diseases in humans, livestock and wildlife. Despite this recognition, little is currently known about the variety of potential pathogens that are carried and transmitted by Australian ticks. In this study, we attempted to expand knowledge of Australian tick-borne bacterial pathogens by analyzing various tick species from the state of Queensland for potential human pathogens belonging to the Rickettsia, Coxiella and Borrelia genera. A total of 203 ticks, comprising of four genera and nine different tick species, were screened by specific qPCR assays. An overall Rickettsia qPCR positivity of 6.4% (13/203) was detected with rickettsial DNA found in four tick species (Ixodes holocyclus, I. tasmani, Amblyommatriguttatum, and Haemaphysalis longicornis). Amplification and analysis of several rickettsial genes from rickettsial qPCR positive samples identified sequences closely related to but genetically distinct from several previously described cultured and uncultured rickettsial species in the Rickettsia spotted fever subgroup. No ticks were positive for either Coxiella or Borrelia DNA. This work suggests that a further diversity of rickettsiae remain to be described in Australian ticks with the full importance of these bacteria to human and animal health yet to be elucidated.