Abstract: The ecology of virulent strains of Rhodococcus equi on horse farms is likely to influence the prevalence and severity of R. equi pneumonia in foals. This study examined the association between the ecology of virulent R. equi and the epidemiology of R. equi pneumonia by collecting air and soil samples over two breeding seasons (28 farm-year combinations) on Thoroughbred breeding farms with different reported prevalences of R. equi pneumonia. Colony blotting and DNA hybridization were used to detect and measure concentrations of virulent R. equi. The prevalence of R. equi pneumonia was associated with the airborne burden of virulent R. equi (both the concentration and the proportion of R. equi bacteria that were virulent) but was not associated with the burden of virulent R. equi in the soil. Univariable screening and multivariable model building were used to evaluate the effect of environmental and management factors on virulent R. equi burdens. Lower soil moisture concentrations and lower pasture heights were significantly associated with elevated airborne concentrations of virulent R. equi, as were the holding pens and lanes, which typically were sandy, dry, and devoid of pasture cover. Few variables appeared to influence concentrations of virulent R. equi in soil. Acidic soil conditions may have contributed to an elevated proportion of virulent strains within the R. equi population. Environmental management strategies that aim to reduce the level of exposure of susceptible foals to airborne virulent R. equi are most likely to reduce the impact of R. equi pneumonia on endemically affected farms.

Abstract: Equine thelaziosis caused by the eyeworm Thelazia lacrymalis is a parasitic disease transmitted by muscid flies. Although equine thelaziosis is known to have worldwide distribution, information on the epidemiology and presence of the intermediate hosts of T. lacrymalis is lacking. In the present work, a PCR-RFLP based assay on the first and/or second internal transcribed spacer (ITS1 and ITS2) of ribosomal DNA was developed for the detection of T. lacrymalis DNA in its putative vector(s). The sensitivity of the technique was also assessed. The restriction patterns obtained readily differentiated T. lacrymalis from four species of Musca (Diptera, Muscidae) (i.e. Musca autumnalis, Musca domestica, Musca larvipara and Musca osiris), which are potential vectors of equine eyeworms. The molecular assay presented herein is a useful tool to identify the intermediate host(s) of T. lacrymalis in natural conditions and to study its/their ecology and epidemiology.

Abstract: Habronema microstoma and Habronema muscae (Spirurida: Habronematidae) are parasitic nematodes which infect the stomach and/or skin of equids. The accurate diagnosis of gastric habronemosis is central to studying its epidemiology, but data on its distribution and prevalence are lacking, mainly due to the limitations of clinical and coprological diagnosis in live horses. To overcome this constraint, a two-step, semi-nested PCR-based assay was validated (utilizing genetic markers in the nuclear ribosomal DNA) for the specific amplification of H. microstoma or H. muscae DNA from the faeces from horses (n = 46) whose gastrointestinal parasite status had been determined at autopsy and whose faeces were examined previously using a conventional parasitological approach. Of these horses examined at autopsy, some harboured adults of either H. microstoma (n= 19) or H. muscae (n =4), and others (n = 7) harboured both species. Most of them were also infected with other parasites, including strongylid nematodes (subfamilies Cyathostominae and Strongylinae), bots and/or cestodes; there was no evidence of metazoan parasites in 2 horses. Larvated spirurid eggs were detected in the faeces of 1 of the 30 horses (3.3 %) shown to be infected with Habronema at autopsy. For this set of 46 samples, the PCR assay achieved a diagnostic specificity of 100 % and a sensitivity of approximately 97 % (being able to specifically detect as little as approximately 0.02 fg of Habronema DNA). The specificity of the assay was also tested using a panel of control DNA samples representing horse, the gastric spirurid Draschia megastoma and 26 other species of parasites from the alimentary tract of the horse. H. microstoma, H. muscae and D. megastoma could be readily differentiated from one another based on the sizes of their specific amplicons in the PCR. The results of this study showed that the performance of the PCR for the diagnosis of gastric habronemosis was similar to that of autopsy but substantially better than the traditional coprological examination procedure used. The ability to specifically diagnose gastric habronemosis in equids should have important implications for investigating the epidemiology and ecology of H. microstoma and H. muscae.

Abstract: Gastric or cutaneous habronemosis caused by Habronema microstoma Creplin, 1849 and Habronema muscae Carter, 1865 is a parasitic disease of equids transmitted by muscid flies. There is a paucity of information on the epidemiology of this disease, which is mainly due to limitations with diagnosis in the live animal and with the identification of the parasites in the intermediate hosts. To overcome such limitations, a molecular approach, based on the use of genetic markers in the second internal transcribed spacer (ITS-2) of ribosomal DNA, was established for the two species of Habronema. Characterisation of the ITS-2 revealed sequence lengths and G+C contents of 296 bp and 29.5% for H. microstoma, and of 334 bp and 35.9% for H. muscae, respectively. Exploiting the sequence difference (approximately 40%) between the two species of nematode, primers were designed and tested by the polymerase chain reaction (PCR) for their specificity using a panel of control DNA samples from common equid endoparasites, and from host tissues, faeces or muscid flies. Effective amplification from each of the two species of Habronema was achieved from as little as 10 pg of genomic DNA. Hence, this molecular approach allows the specific identification and differentiation of the DNA from H. microstoma and H. muscae, and could thus provide a molecular tool for the specific detection of Habronema DNA (irrespective of developmental stage) from faeces, skin and muscid fly samples. The establishment of this tool has important implications for the specific diagnosis of clinical cases of gastric and cutaneous habronemosis in equids, and for studying the ecology and epidemiology of the two species of Habronema.

Abstract: Jamestown Canyon (JC) and Saint Louis encephalitis (SLE) viruses are mosquito-transmitted viruses that have long been present in California. The objective of this study was to determine the seroprevalence of these two viruses in horses prior to the introduction of West Nile (WN) virus. Approximately 15% of serum samples collected in 1998 from 425 horses on 44 equine operations horses throughout California had serum antibodies to JC virus, whereas antibodies were not detected to SLE virus. The results indicate that horses in California were commonly infected prior to 1998 with mosquito-transmitted Bunyaviruses that are identical or closely related to JC virus, but not with SLE virus. The different seroprevalence of SLE and JC viruses in horses likely reflects the unique ecology of each virus, and it is predicted that WN virus will have a wider distribution in California than closely related SLE virus.