Jonas Malmsten

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.

Ninety-nine de novo assembled genomes from the moose ( Alces alces ) rumen microbiome provide new insights into microbial plant biomass degradation

The moose (Alces alces) is a ruminant that harvests energy from fiber-rich lignocellulose material through carbohydrate-active enzymes (CAZymes) produced by its rumen microbes. We applied shotgun metagenomics to rumen contents from six moose to obtain insights into this microbiome. Following binning, 99 metagenome-assembled genomes (MAGs) belonging to 11 prokaryotic phyla were reconstructed and characterized based on phylogeny and CAZyme profile. The taxonomy of these MAGs reflected the overall composition of the metagenome, with dominance of the phyla Bacteroidetes and Firmicutes. Unlike in other ruminants, Spirochaetes constituted a significant proportion of the community and our analyses indicate that the corresponding strains are primarily pectin digesters. Pectin-degrading genes were also common in MAGs of Ruminococcus, Fibrobacteres and Bacteroidetes and were overall overrepresented in the moose microbiome compared with other ruminants. Phylogenomic analyses revealed several clades within the Bacteriodetes without previously characterized genomes. Several of these MAGs encoded a large numbers of dockerins, a module usually associated with cellulosomes. The Bacteroidetes dockerins were often linked to CAZymes and sometimes encoded inside polysaccharide utilization loci, which has never been reported before. The almost 100 CAZyme-annotated genomes reconstructed in this study provide an in-depth view of an efficient lignocellulose-degrading microbiome and prospects for developing enzyme technology for biorefineries.

HAIR-LOSS EPIZOOTIC IN MOOSE (ALCES ALCES) ASSOCIATED WITH MASSIVE DEER KED (LIPOPTENA CERVI) INFESTATION

Deer keds (Lipoptena cervi) are blood-sucking flies in the family Hippoboscidae; moose (Alces alces) are their main host in Scandinavia. There are no detailed reports of the negative impacts of deer keds on moose. In 2006 and 2007, hunters in southeastern Norway and midwestern Sweden found several moose cadavers with severe alopecia; numerous moose had extensive hair loss. Between February 2006 and June 2007, materials from 23 moose were submitted for laboratory examination and large numbers of deer keds were found in the coat of most animals. The body condition of the moose varied but was poor in animals with severe alopecia. The findings of enormous numbers of deer keds in the coat of the majority of the affected animals and a consistent histologic image (acute to chronic, multifocal to coalescing, eosinophilic to lymphocytic dermatitis), concurrent with the absence of any other lesions, trace element deficiencies, or dermal infections which are known to cause alopecia, suggest that the hair-loss epizootic was linked to massive infestations with deer keds. The emergence of this hair-loss syndrome implies that the dynamics between parasite and host have been disrupted by a currently unknown environmental or ecological factor. A high moose density, combined with extraordinarily mild weather June 2006–June 2007 and a particularly long period with the absence of night-frost in autumn of 2006, may have been ideal for deer ked development, survival, and optimal host acquisition.

High Prevalence of Hepatitis E Virus in Swedish Moose – A Phylogenetic Characterization and Comparison of the Virus from Different Regions

Background Hepatitis E virus (HEV) infects a range of species, including humans, pigs, wild boars and deer. Zoonotic transmission may contribute to the high HEV seroprevalence in the human population of many countries. A novel divergent HEV from moose (Alces alces) in Sweden was recently identified by partial genome sequencing. Since only one strain was found, its classification within the HEV family, prevalence in moose and zoonotic potential was unclear. We therefore investigated samples from 231 moose in seven Swedish counties for HEV, and sequenced a near complete moose HEV genome. Phylogenetic analysis to classify this virus within the family Hepeviridae and to explore potential host specific determinants was performed. Methods and Findings The HEV prevalence of moose was determined by PCR (marker for active infection) and serological assays (marker of past infection) of sera and 51 fecal samples from 231 Swedish moose. Markers of active and past infection were found in 67 (29%) animals, while 34 (15%) were positive for HEV RNA, 43 (19%) were seropositive for anti-HEV antibodies, and 10 (4%) had both markers. The number of young individuals positive for HEV RNA was larger than for older individuals, and the number of anti-HEV antibody positive individuals increased with age. The high throughput sequenced moose HEV genome was 35-60% identical to existing HEVs. Partial ORF1 sequences from 13 moose strains showed high similarity among them, forming a distinct monophyletic clade with a common ancestor to HEV genotype 1-6 group, which includes members known for zoonotic transmission. Conclusions This study demonstrates a high frequency of HEV in moose in Sweden, with markers of current and past infection demonstrated in 30% of the animals. Moose is thus an important animal reservoir of HEV. The phylogenetic relationship demonstrated that the moose HEV belonged to the genotype 1-6 group, which includes strains that also infect humans, and therefore may signify a potential for zoonotic transmission of this HEV.

REVERSIBLE IMMOBILIZATION OF FREE-RANGING SNOW LEOPARDS (PANTHERA UNCIA) WITH A COMBINATION OF MEDETOMIDINE AND TILETAMINE-ZOLAZEPAM

Conservation and research of the elusive snow leopard (Panthera uncia) have been hampered by inadequate knowledge about its basic life history. Global positioning system (GPS) collars can provide useful information, but there has been limited information available on safe capture methods, drug doses, and efficacy for effective immobilization of free-ranging snow leopards. We describe a drug protocol using a combination of medetomidine and tiletamine-zolazepam for the chemical immobilization of free-ranging snow leopards. We also describe physiologic responses to immobilization drugs, including rectal temperature, heart rate, respiratory rate, and relative hemoglobin oxygen saturation (SpO2) recorded every 10 min. Our study was carried out in the Tost Mountains adjacent to the Great Gobi Desert, in southern Mongolia, between August 2008 and April 2012. Eighteen snow leopards were captured or recaptured with foot-snares on 42 occasions and anesthetized for marking with GPS collars. The snow leopards received on average (±SD) 0.020±0.04 mg/kg body mass medetomidine and 2.17±0.45 mg/kg tiletamine-zolazepam. The duration of ensuing anesthesia was 69±13 min, including an induction period of 10 (±4) min. Anesthesia was reversed with 4 mg (0.10±0.04 mg/kg) atipamezole administered intramuscularly. The mean value for SpO2 for the 37 captures where we could record physiologic values was 91±4. The SpO2 increased significantly during anesthesia (+0.06±0.02%/min), whereas rectal temperature (average 38.1±0.7 C/min, change −0.04±0.003 C/min), heart rate (average 97±9 beats/min, change −0.20±0.03 beats/min), and respiratory rate (average 26±6 breaths/min, change −0.11±0.03 breaths/min) decreased significantly. A dose of 80 mg tiletamine-zolazepam (2 mg/kg body weight) and 0.72 mg medetomidine (0.02 mg/kg body weight) safely immobilized all adult and subadult snow leopards (weight 25–45 kg) in our study. All measured physiologic values remained within clinically acceptable limits.

Temporal and spatial variation in Anaplasma phagocytophilum infection in Swedish moose (Alces alces)

SUMMARY The occurrence of Anaplasma phagocytophilum was investigated in spleen and serum samples from Swedish moose (Alces alces) in southern Sweden (island and mainland). Samples were analysed for presence of A. phagocytophilum DNA by real-time PCR (n = 263), and for Anaplasma antibodies with ELISA serology (n = 234). All serum samples had antibodies against A. phagocytophilum. The mean DNA-based prevalence was 26·3%, and significant (P < 0·01) temporal, and spatial variation was found. Island moose had significantly (P < 0·001) higher prevalence of A. phagocytophilum DNA than moose from the mainland areas. Two samples were sequenced to determine genetic variation in the 16S rRNA and groESL genes. Genetic sequence similarity with the human granulocytic anaplasmosis agent, equine granulocytic ehrlichiosis agent, and different wildlife-associated A. phagocytophilum variants were observed in the 16S rRNA and groESL genes. Our study shows that moose are exposed to A. phagocytophilum in Sweden, and represent a potential wildlife reservoir of the pathogen.

Seroprevalence of Neospora caninum in gray wolves in Scandinavia

Transmission of the protozoan parasite Neospora caninum between wild and domestic animals has gained some interest during recent years. Because of the close relationship between gray wolf (Canis lupus) and dog it has been suggested that gray wolf is a definitive host for the parasite. The aim of this study was to estimate the seroprevalence of N. caninum in Scandinavian gray wolves and to investigate any geographical patterns of the infection. The investigation was based on blood samples collected from 109 wolves between 1998 and 2009 within the Scandinavian wolf project Skandulv. They were analysed by N. caninum iscom ELISA and those with absorbance values exceeding 0.20 were also analysed by immunoblotting. Samples that were positive in both tests were deemed positive. Four (3.7%) wolves were positive at the first sampling. They were all sampled 2005 at different locations, and were both females and males. From one male wolf three samples were collected over a 7-year period. No antibodies were detected at the first sampling in 1998 when he was approximately 8 months old but when he was sampled again 5 and 7 years later the ELISA and immunoblotting were positive. The results indicate that N. caninum infection is present in Scandinavian wolves. It is unclear how the wolves acquired the infection.

Surveys on Coxiella burnetii infections in Swedish cattle, sheep, goats and moose

BackgroundQ fever is a zoonotic disease caused by the bacterium Coxiella burnetii. Prevalence data in ruminant species are important to support risk assessments regarding public and animal health. The aim was to investigate the presence of or exposure to C. burnetii in cattle, sheep, goats and moose, and to compare two enzyme-linked immunosorbent assays (ELISAs). National surveys of antibodies against C. burnetii were performed for dairy cattle (n=1537), dairy goats (n=58) and sheep (n=518). Bovine samples consisted of bulk milk, caprine of pooled milk, and ovine of pooled serum. Antibodies were investigated in moose samples (n=99) from three regions. A one-year regional cattle bulk milk survey was performed on the Isle of Gotland (n=119, four occasions). Cattle, sheep and goat samples were analysed with indirect ELISA and moose samples with complement fixation test. For the sheep, goat, and parts of the cattle survey, samples were run in parallel by ELISAs based on antigens from infected ruminants and ticks. Bulk milk samples from the regional cattle survey and vaginal swabs from a subset of the sheep herds (n=80) were analysed for the agent by polymerase chain reaction. Spatial clustering was investigated in the national cattle survey.ResultsThe prevalence of antibodies in dairy herds was 8.2% with large regional differences. High risk clusters were identified in the southern regions. The prevalence among dairy herds on the Isle of Gotland varied from 55.9% to 64.6% and 46.4% to 58.9.0% for antibodies and agent, respectively, overall agreement between agent and antibodies was 85.2%. The prevalence of antibodies in sheep was 0.6%, the agent was not detected the vaginal swabs. Antibodies were not detected in goats or moose, although parts of the moose samples were collected in an area with high prevalence in cattle. The overall agreement between the two ELISAs was 90.4%.ConclusionsThe prevalence of antibodies against C. burnetii in dairy cattle in Sweden shows large regional differences. The results suggest that C. burnetii is a rare pathogen among Swedish moose, dairy goat and sheep. ELISAs based on ruminant and tick antigen performed in a similar manner under Swedish conditions.

Opportunities and challenges with growing wildlife populations and zoonotic diseases in Sweden

In many parts of Europe and North America, populations of large mammals and birds have recovered during recent decades. In Sweden, this has resulted in more wildlife than was historically recorded. This positive development provides a number of opportunities for ecosystem services and for biodiversity. More wildlife also means more interactions with humans, as many birds and mammals may interfere with human interests in the landscape, such as natural resource use. Thus, more wildlife may shift the baseline for societal perception of wildlife. Wildlife species may host pathogens with potential for infecting humans and domestic animals. With increasing knowledge about zoonotic diseases and their dynamics, more scientific, media, and public attention is given to zoonotic processes. We are concerned with how the public image of the wild animals is affected, because many of the recent zoonotic outbreaks connect animal groups to diseases such as avian and swine influenza, lyme disease, and tick-borne encephalitis. The societal focus on zoonotic diseases may increase the fear of the wild and will separate the public further from the outdoors in general and wildlife in particular. Ultimately, we risk a juxtaposition of the overall acceptance of biological diversity and a shifting societal perception of wildlife that could be harmful for life on earth. We therefore suggest multidisciplinary research on societal awareness of zoonotic diseases and its implications for public acceptance for wildlife and biological diversity.

Differences in body mass, health status and genetic variation between insular and mainland brown hares (Lepus europaeus) in Sweden

Introduced populations can be affected by random processes such as genetic drift, deterministic processes given by the local environmental conditions and anthropogenic factors such as hunting and management. Geographically constrained populations are particularly exposed to these processes, and altogether, these factors may result in rapid differentiation from the ancestral populations. The introduced European brown hare (Lepus europaeus) population on the island Ven is isolated from the Swedish and Danish mainland. Undocumented observations suggest that the hares on the island have been increasingly diseased in recent years and also decreased in body size. To test the substance of these observations, as well as the potential for inbreeding depression in this geographically constrained population, a total of 321 hares from Ven and three reference populations on the Swedish mainland were analysed with respect to body mass, general health status and genetic variation. The results confirm that the hares on Ven have lower body mass than hares on the mainland, but there are no indications of health deficits. We argue that the difference in body mass primarily is an island effect of stress and/or nutritional shortage, possibly induced by high population density, anthropogenic selection regimes and absence of mammalian meso-predators. Further, the genetic data indicate that the insular population is substructured, and subadults from these two subpopulations differ in body mass. This apparent substructuring could be due to chance effects, but may also be related to assortative mating or presence of sustained populations with different ancestry.