Meningoencephalomyelitis caused by Neospora caninum in a juvenile fallow deer (Dama dama)

Neosporosis, caused by the protozoan parasite Neospora caninum, is a serious cause of bovine abortion, stillbirth and perinatal death. This paper reports a clinical neosporosis in a 3-week-old fallow deer (Dama dama). The fawn was full term and appeared normal at birth. Histological lesions consisted of a multifocal necrotizing and granulomatous meningoencephalomyelitis, with intralesional protozoal cysts. Positive immunohistochemical staining and characteristic PCR products confirmed the diagnosis of N. caninum infection.     

Isolation of Neospora caninum from naturally infected white-tailed deer (Odocoileus virginianus)

Attempts were made to isolate Neospora caninum from naturally infected white-tailed deer (Odocoileus virginianus). A total of 110 deer killed during the 2003 hunting season in Virginia region were used for the isolation of N. caninum. Of these, brains from 28 deer that had NAT titer of 1:200 were inoculated into interferon-gamma gene knock out (KO) mice. N. caninum was isolated from the tissues of three deer and all three isolates were mildly virulent to KO mice. Only one of the isolates could be adapted to in vitro growth. Protozoa in the tissues of KO mice reacted with N. caninum-specific polyclonal antibodies and N. caninum DNA was demonstrated in infected tissues by PCR assays; sequences of portions of the ITS-1 and gene 5 loci were identical to those in the public database. This is the first record of in vitro isolation of N. caninum from white-tailed deer and lends credence to the white-tailed deer as an intermediate host for this parasite.     

Xylazine-induced sedation in axis deer (Axis axis) and its reversal by atipamezole

Eight free-ranging axis deer (Axis axis) were captured in drive nets and injected with xylazine (3.4±0.1 mg/kg; mean ±SEM) intramuscularly using a hand-held syringe. Xylazine induced complete immobilization and sedation in three animals, heavy sedation in three, and moderate sedation in two. The mean induction time was 10.4±1.0 min. The mean rectal temperature, heart and respiratory rates of immobilized animals were 39.2±0.4°C, 75.5±6.5 beats/min and 62.1±4.2 breaths/min, respectively.All the animals were given atipamezole intravenously for reversal. The mean time from injection of xylazine to administration of atipamezole was 37.8±4.6 min. A dose ratio (w/w) for xylazine:atipamezole-HCl of 10:1 was used. The mean time from injection of atipamezole to mobility was 2.41±0.58 min.Atipamezole given intravenously effectively antagonized xylazine-induced sedation in axis deer. Only one animal showed signs of overalertness after reversal and no cases of resedation were observed.Abbreviations i.m. intramuscular(ly) - i.v. intravenous(ly) - SEM standard error of the mean     

Seroprevalence of Neospora caninum and Toxoplasma gondii in black-tailed deer (Odocoileus hemionus columbianus) and mule deer (Odocoileus hemionus hemionus)

Deer are considered important intermediate hosts for the coccidian parasites, Toxoplasma gondii and Neospora caninum. Antibodies to N. caninum and T. gondii were determined in sera of 42 mule deer (Odocoileus hemionus hemionus) and 43 black-tailed deer (Odocoileus hemionus columbianus) from Washington state, USA, using direct agglutination test with specific antigens. A titer of 1:25 was considered diagnostic for both parasites. N. caninum antibodies were found in 7 of 42 mule deer and 8 of 43 black-tailed deer. T. gondii antibodies were found in 14 black-tailed deer but not in any of the mule deer. This is probably the first report of seroprevalence of N. caninum in these hosts.     

Seroprevalence of Neospora caninum in free living and farmed red deer (Cervus elaphus) in Poland

Serum samples from 47 free living and 106 farmed red deer (Cervus elaphus) from the Mazurian Lake District in north-east Poland were investigated for the presence of antibodies to Neospora caninum. A modified Neospora iscom-ELISA was used for initial screening. All sera with optical density (OD) values exceeding 0.400 absorbance units were further investigated by Western blot analysis. Eighteen sera were positive in both tests. Six of these were from free living and 12 from farmed animals giving prevalence of 13 and 11%, respectively. This is the first report of N. caninum infection in farmed and free-living red deer living in the same region where neosporosis was confirmed in cattle and the first evidence of exposure to the parasite in red deer in Poland.     

Gray wolf (Canis lupus) is a natural definitive host for Neospora caninum

The gray wolf (Canis lupus) was found to be a new natural definitive host for Neospora caninum. Neospora-like oocysts were found microscopically in the feces of three of 73 wolves from Minnesota examined at necropsy. N. caninum-specific DNA was amplified from the oocysts of all three wolves. Oocysts from one wolf were infective for the gamma interferon gene knock out (KO) mice. Viable N. caninum (designated NcWolfUS1) was isolated in cell cultures seeded with tissue homogenate from the infected mouse. Typical thick walled tissue cysts were found in outbred mice inoculated with the parasite from the KO mouse. Tissue stages in mice stained positively with N. caninum-specific polyclonal antibodies. Our observation suggests that wolves may be an important link in the sylvatic cycle of N. caninum.     

Neospora caninum infection as a cause of reproductive failure in a sheep flock

Neospora caninum has been detected only sporadically in cases of ovine abortion, and it has therefore traditionally been considered as an unimportant parasite in small ruminants. This study was carried out with the aim of identifying the pathogen causing serious reproductive problems on a commercial sheep farm. Sera from all rams and ewes tested negative for antibodies against Border disease virus, Schmallenberg virus and Coxiella burnetii, and infections by these agents were therefore ruled out. Nevertheless, seropositivity to N. caninum and/or Toxoplasma gondii was detected, although the seroprevalence was higher in the case of N. caninum. The percentage of lambings and the number of lambs per dam were significantly lower in ewes that were seropositive to N. caninum while no effect on these parameters was detected in ewes that were seropositive to T. gondii. There was also no evidence of infection by T. gondii in the foetal/lamb tissues analyzed by PCR and/or immunohistopathological techniques. On the contrary, the DNA of N. caninum was detected in 13 out of 14 foetuses/lambs descendant from dams seropositive to this parasite. Characteristic lesions caused by N. caninum and/or its antigen were also detected. Genotyping of the N. caninum DNA revealed only two closely related microsatellite multilocus genotypes. The results clearly demonstrate that infection by N. caninum was the cause of the low reproductive performance of this sheep flock.     

Causes of perinatal mortality in farmed fallow deer (Dama dama)

The causes of perinatal mortality in farmed fallow deer in the Camden region of New South Wales were determined between 1982 and 1987. The results of 144 necropsies on fawns showed that ante-parturient and parturient deaths occurred in 33 (22.9%) of cases, with post-parturient deaths accounting for 99 (69.8%), and undiagnosed causes the remaining 12 (8.3%). The most common cause of post-parturient death (43.1%) was exposure/starvation of fawns that had birthweights lower than the average for surviving fawns on these same farms (3.01 kg and 4.12 kg, respectively). These results provide good evidence of the correlation between low birthweight and perinatal mortality in fallow deer on Australian deer farms.     

Neospora caninum antibodies detected in Midwestern white-tailed deer (Odocoileus virginianus) by Western blot and ELISA

White-tailed deer (Odocoileus virginianus) serve to maintain the Neospora caninum life cycle in the wild. Sera from white-tailed deer from south central Wisconsin and southeastern Missouri, USA were tested for antibodies to N. caninum by Western blot analyses and two indirect ELISAs. Seroreactivity against N. caninum surface antigens was observed in 30 of 147 (20%) of WI deer and 11 of 23 (48%) of MO deer using Western blot analysis. Compared to Western blot, the two indirect ELISAs were found to be uninformative due to degradation of the field-collected samples. The results indicate the existence of N. caninum antibodies in MO and WI deer, and that Western blot is superior to ELISA for serologic testing when using degraded blood samples collected from deer carcasses.     

Normal haematological values of axis deer (Axis axis), Père David's deer (Elaphurus davidianus) and barasingha (Cervus duvauceli)

Full blood counts and fibrinogen estimations were carried out on a number of clinically normal adult axis deer (Axis axis), Père David's deer (Elaphus davidianus) and barasingha (Cervus duvauceli). Apart from the finding of higher total red cell counts and a lower mean red cell volume in axis deer than in Père David's deer and barasingha, there was little interspecies variation. Some individual Père David's deer showed eosinophilia, thought to be associated with subclinical intestinal nematode infestation. The findings should provide a useful reference base for monitoring health in these species.     

Influence of modifiable risk factors on the incidence of stillbirth/perinatal mortality in dairy cattle

Bovine perinatal mortality is defined as the death of a full-term calf before, during or up to 48 h after calving. Recent studies indicate that the prevalence of bovine perinatal mortality is increasing, particularly in Holstein primiparae. Factors leading to a greater incidence of dystocia are the most important modifiable variables influencing the risk of perinatal mortality. Modifiable predictors are largely (age at first calving, breeding method, sire, calving management, feto-maternal health status and gestational nutrition) or moderately (calf breed, sex, gestation length) under the control of the dairy farm manager. Unlike non-modifiable risk factors, such as primiparity and fetal plurality, these factors can be manipulated to reduce the incidence of perinatal mortality.     

Prevalence of antibodies against Toxoplasma gondii and Neospora caninum in moose (Alces alces) and roe deer (Capreolus capreolus) in Sweden

Toxoplasma gondii and Neospora caninum are two coccidian parasites with a worldwide distribution. T. gondii is one of the more common parasitic zoonoses in the world and in young children and immunocompromised persons, infection can lead to severe disease and death. N. caninum is an important cause of abortions in cattle. Wildlife have been identified as reservoirs and transmitters for both parasites. The purpose of this study was to investigate the seroprevalences of T. gondii, and N. caninum in moose (Alces alces), and roe deer (Capreolus capreolus) in Sweden. Blood samples were collected from 417 moose during 2000-2005 and from 199 roe deer during 1990-2007. The samples were investigated for presence of antibodies by a T. gondii direct agglutination test and a N. caninum iscom ELISA. Because the iscom ELISA has not been validated for moose or roe deer, sera that gave a positive result were further investigated by immunoblot analysis to verify presence of antibodies. Antibodies to T. gondii were detected in 85 (20%) and 68 (34%) moose and roe deer sera, respectively. In moose the seroprevalence was higher in south and central Sweden than in the north, whereas there was no difference between the regions for roe deer. Adult moose and roe deer had higher odds of being seropositive than young animals but there were no difference in seroprevalence between males and females. One roe deer was positive by immunoblotting and was regarded as N. caninum positive, whereas all moose sera were negative. The results show that T. gondii infection is widely spread in the Swedish moose and roe deer populations. Precautions should therefore be taken when handling internal organs and carcasses of harvested cervids. Proper handling and cooking of game meat also is important to prevent toxoplasmosis in humans.     

Toxoplasmosis as a cause of ovine perinatal mortality

Aims. To determine temporal and spatial patterns of bovine tuberculosis (Tb) in a population of brushtail possums (Trichosurus vulpeculu) free from commercial and recreational hunting and living contiguously with livestock, and to relate these patterns to measures of possum health and density.

Methods. Possums were trapped on 11 lines located on a forest-rough grazing margin in the Ahaura Valley in Westland in 1979-80 and each August from 1992 to 1996, and were examined post mortem for gross lesions indicative of Tb. Infection levels in possums were compared with Th test data from cattle grazing in the same area.

Results. Tuberculosis was identified from the trapped sample of possums in 1980. Trapping of further possums on the same trap lines confirmed the presence of the disease each year from 1992 to 1996, and suggested a clustering of infection in time and space. The prevalence of Th in possums declined with increasing possum population size and well being, and decreasing age. The prevalence of Tb in possums was highest in 1980 and 1992, and appeared to roughly coincide with similar upsurges in Tb in nearby cattle.

Conclusions. Our results suggest that the exceptional levels of Tb recorded in possums in 1992 had a direct effect on possum numbers, and gave rise to very low trap catches in 1993-94. They also suggest thatTb can persist for several years in possum populations existing well below the 5% trap catch targeted by regipnal councils forTb possum control.     

Serosurvey of antibodies against Toxoplasma gondii and Neospora caninum in white-tailed deer from Northern Mexico

The objective of this study was to determine the seroprevalence of Toxoplasma gondii and Neospora caninum in white-tailed deer from Northern Mexico. Sera from 532 white-tailed deer (Odocoileus virginianus) from three Northern states of Mexico were assayed for antibodies to T. gondii by ELISA and western blot. From these samples, 368 were available to test for N. caninum antibodies by ELISA. The overall prevalence for T. gondii antibodies was 13.9% (74/532; CI(95) 11-17) and for N. caninum 8.4% (31/368; CI(95) 6-12). There was a significant association between positive ELISA results for T. gondii, with management factors within ranches, such number of deer per hectare and geographic location of deer, but none for N. caninum. T. gondii infection in the deer from Guerrero, Coahuila had an increased risk than those from Nuevo Laredo, Tamaulipas (OR, 8.3; CI(95) 1.9-35.4; P<0.05) and ranches with one deer in 15ha had increased risk of positive association (OR, 2.61; CI(95) 1.5-4.4; P<0.05). These findings may have environmental or public health implications because venison can be an important meat source of T. gondii infections for humans and feral cats.     

Red foxes (Vulpes vulpes) are a natural intermediate host of Neospora caninum

The present study was undertaken to determine if red foxes are natural intermediate and/or definitive host for Neospora caninum and to study the importance of infection of N. caninum in this species in North-eastern Spain. Faecal samples and brain tissues were obtained from 122 foxes from 21 rural areas of Catalonia. Faeces collected were examined for parasite eggs and coccidian oocysts using sucrose flotation. For PCR-based diagnosis of N. caninum in brain tissues, the specific genomic Nc5 region was selected as the target sequence for DNA amplification. To control for PCR failure and facilitate identification of truly negative samples, the competitor pNc5C molecule was added to all negative samples in a second round of PCR reactions. Of the 122 foxes analysed, 13 (10.7%) were positive by PCR for N. caninum. Signal intensities of all positive samples were relatively weak with the exception of one sample from a 3-month male animal, that also showed the highest repeatability. No differences were observed by sex, age or area of sampling analysis. Detection of stages of N. caninum in brain from naturally infected red foxes demonstrated that red foxes are a natural intermediate host for N. caninum. Faecal samples were analysed for the presence of N. caninum oocysts, however, no oocysts compatible with N. caninum were found. A widespread latent infection of red foxes in North-eastern Spain found in the present study indicates that red foxes could have a very important role in the epidemiology of neosporosis in our area.     

Antibodies to Neospora caninum in European brown hare (Lepus europaeus)

Neospora caninum antibodies in European brown hares (Lepus europaeus) were reported for the first time in sera collected from hares imported in Italy from East Europe. Sera from 93 hares coming from Hungary and 44 hares coming from Slovakia were tested using a direct agglutination test (DAT). Seroprevalence was 8.0% at the 1:40 dilution and 1.5% at 1:320 dilution. Seroprevalence in females and males and in different countries of origin did not differ significantly. Our results indicate that European brown hares are exposed to N. caninum. The low prevalence observed suggests a hares minimal exposure, but, considering that hares are common prey for foxes, it could be supposed that, also with a low prevalence, hares can be an important source of N. caninum infection in foxes.     

Isolation of Neospora caninum from a calf in Malaysia

In order to attempt isolate the protozoan parasite Neospora caninum, an N. caninum seropositive pregnant Sahiwal Friesian cross heifer from a large-scale dairy farm in Malaysia was kept for observation until parturition at the Veterinary Research Institute, Ipoh. The heifer gave birth to a female calf that was weak, underweight and unable to rise. Precolostral serum from the calf had an N. caninum indirect fluorescent antibody test titre of 1:3200. It died 12 h after birth and necropsy was performed. Brain homogenate from the calf was inoculated into 10 BALB/c mice that were kept for 3 months after which brain tissue from the mice was inoculated onto 24 h fresh monolayer Vero cell lines. The cell cultures were examined daily until growth of intracellular protozoa was observed. DNA of the organisms from the cell cultures was analyzed by PCR and DNA sequencing. DNA fragments of the expected size were amplified from the isolate using N. caninum-specific primers, and sequence analysis of ITS1 clearly identified the isolate as N. caninum. This is the first successful isolation of N. caninum from a bovine in Malaysia, and the isolate is designated Nc-MalB1.     

Finding of Neospora caninum in the wild brown rat (Rattus norvegicus)

Nine rats (16.4%) out of 55 (Rattus norvegicus) from cattle farms were seropositive to Neospora caninum. Two of the seropositive rats were also PCR positive but all were negative by immunohistochemistry and PAS staining. The brains of all the captured rats were homogenized and initially inoculated intraperitoneally into nude mice or into SPF ICR mice, which had been immunosuppressed with prednisolone. One mouse that was inoculated with brain material from a seropositive rat became infected with N. caninum, as demonstrated by the presence of a tissue cyst in the brain and confirmed by immunohistochemistry and PCR. This is the first finding of N. caninum in naturally infected farm rats. The findings show that natural N. caninum infection occurs in wild brown rats and thus rats may serve as a reservoir for the protozoan on the cattle farm.