Susceptibility of carnivore hosts to strains of canine distemper virus from distinct genetic lineages

Using the complete haemagglutinin (HA) gene and partial phosphoprotein (P) gene we investigated the genotype of canine distemper virus (CDV) strains recovered from two wildlife species in Mecklenburg-Vorpommern, Germany. Phylogenetic analyses demonstrated significant differences between the strains from raccoons Procyon lotor (family Procyonidae) obtained in 2007 and strains from red foxes Vulpes vulpes (family Canidae) obtained in 2008. The raccoon strains belonged to the CDV European wildlife lineage whereas the red fox strains belonged to the CDV Europe lineage. We combined our genetic sequence data with published data from 138 CDV stains worldwide to investigate the proposed importance of amino acid substitutions in the SLAM binding region of the CDV HA protein at position 530 (G/E to R/D/N) and 549 (Y to H) to the spread of domestic dog-adapted CDV strains to other carnivores. We found no evidence that amino acid 530 was strongly affected by host species. Rather, site 530 was conserved within CDV lineages, regardless of host species. Contrary to expectation, strains from non-dog hosts did not exhibit a bias towards the predicted substitution Y549H. Wild canid hosts were more frequently infected by strains with 549Y, a pattern similar to domestic dogs. Non-canid strains showed no significant bias towards either H or Y at site 549, although there was a trend towards 549H. Significant differences between the prevalence of 549Y and 549H in wild canid strains and non-canid strains suggests a degree of virus adaptation to these categories of host.     

Relative condition factor and parasitism in anostomid fishes from the floodplain of the Upper Paraná River, Brazil

Individuals of four species of Leporinus were captured in the floodplain of the Upper Paraná River and their metazoan parasites were collected. Fifty-eight taxa of ectoparasites and endoparasites were recorded: 31 in Leporinus lacustris, 32 in Leporinus friderici, 28 in Leporinus obtusidens and 25 in Leporinus elongatus. The aim of this paper was to study the relationship between the host's condition and the parameters of infrapopulations and infracommunities. The health of the host was represented by the relative condition factor (Kn). Richness and number of individuals in the infracommunities of ectoparasites covariated with the Kn of the hosts in one species of fish. Some infrapopulations of ectoparasites covariated negatively and/or the mean Kn of parasitized individuals was lower than those without parasites. The abundance in some infrapopulations of endoparasites covariated positively with the Kn and/or the mean Kn of parasitized fish was better. These results may be related to different transmission strategies used by ecto and endoparasites.     

Paragonimus westermani and some rare intestinal trematodes recovered from raccoon dogs (Nyctereutes procyonoides viverrinus) introduced recently on Yakushima Island, Japan

Helminth parasites were collected from 9 raccoon dogs (Nyctereutes procyonoides viverrinus) and 2 Japanese weasels (Mustela itatsi sho) on Yakushima Island, Japan. The former carnivore was introduced to this World Natural Heritage Area presumably within the last two decades, expanding its population thence, although detailed process(es) of the introduction is unknown. The collected trematodes from raccoon dogs included the triploid form of Paragonimus westermani, Brachylaima tokudai, Maritrema eroliae, and Pseudocryptotropa sp. Simultaneously, Paragonimus ohirai was found in one weasel killed by a traffic accident. Although the triploid form of P. westermani and P. ohirai are known to be distributed in some river-mouth areas of Yakushima Island based on previous surveys on crab hosts, natural infection was detected for the first time in wild final hosts. Particularly, the raccoon dog infected with P. westermani was caught in a mountainous area, distant from human residence or river-mouth areas. Although it is possible that the infected raccoon dog moved from a river-mouth area endemic with P. westermani after infection, the alternative scenario remains to be pursued; the endemic area of this zoonosis is expanding along with the recent expansion of raccoon dogs or feral cats (Felis catus), that became prevalent recently on this island including the mountainous areas. Maritrema eroliae taking a variety of shorebirds as its natural final hosts, and a minute trematode, Pseudocryptotropa sp., taking unknown natural final host(s) were recorded for the first time in raccoon dogs.     

Host range relationships and the evolution of canine parvovirus.

Canine parvovirus (CPV) is an example of an unusual class of emerging virus-those that gain an altered host range through genetic variation and subsequently become widespread pathogens of their new and previously resistant host species. CPV was first detected in 1978 as the cause of new diseases in dogs throughout the world, when it rapidly spread throughout domestic populations, as well as becoming widespread in wild dogs. CPV was soon shown to be a variant of the long recognized feline panleukopenia virus (FPV), from which it differed in less than 1% at the nucleotide sequence level. Genetic analysis showed that virtually all of the biological differences between CPV and FPV, including the canine host range, were determined by three or four sequence differences in the viral capsid protein gene. Analysis of the atomic structures of the CPV and FPV capsids showed that the differences controlling host range were located within two different structural regions and were exposed on the capsid surface. The CPV which first emerged in 1978 appeared to be derived from a single ancestral sequence, which has allowed the ready analysis of the subsequent evolution of the virus in nature. Sequence analysis has also revealed that CPV strains have undergone a series of evolutionary selections in nature which have resulted in the global distribution of new virus variants. This was first seen in the global replacement between 1979 and 1981 of the original (1978) strain of the virus by a genetically and antigenically variant strain, and the subsequent widespread selection of other variants which have also become globally distributed. The genetic and antigenic variation in the virus strains was also correlated with changes in the host range of the virus, in particular in the ability to replicate in cats, and in canine host range differences seen in tissue culture cells.     

犬流感的流行病学研究进展

近年来,不同宿主来源的流感病毒感染犬的病例报道使得犬成为流感病毒生态循环中的重要中间宿主。介于人与犬之间的亲密关系,其公共卫生意义越来越值得人们关注。本文从流感病毒由马—犬、禽—犬、猪—犬和人—犬等跨宿主传播方面阐述不同宿主流感病毒在犬中的流行现状及其重配现象,这对今后全面掌握其流行特征和切断其传播途径具有重要的借鉴和指导意义。     

Echinococcus granulosus in wildlife in and around the Kosciuszko National Park, south-eastern Australia

OBJECTIVE: To investigate the distribution of Echinococcus granulosus in wild dogs and foxes and hydatidosis in wildlife coexisting with foxes and wild dogs in and around Kosciuszko National Park. DESIGN: Prospective and ad hoc surveys by necropsy of definitive and intermediate hosts. PROCEDURE: Wild dogs and foxes were trapped at one location in the Kosciuszko National Park and at 7 locations around the periphery of the Park. Feral pigs, macropodid marsupials, wombats, and feral goats were collected at some of the same locations. The animals were humanely killed, their small intestines removed in the field, the contents collected, preserved and examined microscopically. All internal organs of intermediate hosts were examined for hydatid cysts. Unidentified lesions were examined histologically. RESULTS: Echinococcus granulosus tapeworms were found in wild dogs from all locations. Prevalence ranged up to 100% with worm burdens up to 300,000 worms. Prevalence in foxes ranged up to 50% in animals recovered from 5 locations. The worm burdens were usually less than 50 E. granulosus per fox. Hydatid cysts were found in all macropodid species. Prevalence (69%) and cyst fertility (100%) were highest in swamp wallabies (Wallabia bicolour). Prevalence of cysts in feral pigs ranged up to 49%. Less than 22% of the cysts were fertile. No cysts were found in any of the wombats or feral goats. CONCLUSION: Echinococcus granulosus occurs commonly in wildlife in and around the Kosciuszko National Park. High numbers of fertile cysts in swamp wallabies, a favoured dietary item for wild dogs in this region, suggests swamp wallabies are pivotal in maintaining transmission. Physical contact with wild dogs and foxes or accidental contact with wild canid faeces is a public health risk.     

Evaluation of a genetic assay for canine transmissible venereal tumour diagnosis in Brazil

The canine transmissible venereal tumour (CTVT) is a transmissible cancer that is spread between dogs by the allogeneic transfer of living cancer cells. The infectious agents in CTVT are the living cancer cells themselves, which are transmitted between dogs during coitus. CTVT first arose several thousand years ago and the disease has a global distribution and is frequently observed in dogs from Brazil. We evaluated the utility of a LINE‐MYC quantitative polymerase chain reaction for diagnosis of CTVT cases in Brazil. Our analysis indicated that the LINE‐MYC rearrangement was detectable in all CTVT samples but not in their corresponding hosts. This genetic assay proves to be a useful tool for providing a definitive molecular diagnosis of CTVT, which presents with varying degrees of aggressiveness and invasiveness in different host dogs and can therefore be a diagnostic challenge in some specific cases.     

The occurrence of Contracaecum sp. larvae (Nematoda: Anisakidae) in the catfish Clarias gariepinus (Burchell) from Lake Chivero, Zimbabwe

Clarias gariepinus were collected from Lake Chivero, Zimbabwe, and examined for nematode parasites from November 2000 to May 2002. Of the 202 specimens collected, 42.6% were infected with third-stage larvae of Contracaecum sp. in the body cavity. The intensity of the infection was 1-7 worms per fish (mean intensity = 2.2). Seasonal variation in the prevalence of the parasite was not obvious and there was no significant difference in the prevalence of infection between males and females (chi 2 = 2.228; P > 0.05). No significant relationship between host size and prevalence was established. There was also no significant relationship between intensity and the body condition factor (r = 0.11; P > 0.05). The low parasite prevalence may have been caused by the disruption of the infection cycle since piscivorous birds, which are the final hosts of the parasite, do not feed on C. garieplnus in Lake Chivero.     

Egg trait variation in a large hawk‐cuckoo (Hierococcyx sparverioides) host population of Chinese babax (Babax lanceolatus)

Mutual interaction between brood parasites and their hosts is a well‐known model system for studying host–parasite coevolution. Both parties have acted reciprocally, resembling an evolutionary arms race, in which adaptations and counter‐adaptations have evolved as a result of host–parasite dynamics, such as the classical cuckoo–host system. Discrimination among parasite and cuckoo eggs and rejection of foreign eggs is regarded as an important anti‐parasitism strategy. The Chinese babax (Babax lanceolatus) is a large hawk‐cuckoo (Hierococcyx sparverioides) host distributed in southwest China. A previous study shows that the babax is an intermediate egg rejector, and most cuckoo eggs are accepted by the Chinese babax, although a small proportion of hosts reject cuckoo eggs. Interestingly, the large hawk‐cuckoo lays non‐mimetic eggs in contrast to the uniform blue eggs of babaxes. Because egg coloration is a critical cue used by host species in favor of the recognition of parasitic eggs by hosts, we used a spectrometer to quantify egg color variation to understand the differentiation in discrimination ability between the egg rejectors and acceptors. We found that the chroma of intra‐clutch variation of babax eggs was more consistent in egg rejectors than in acceptors. However, no statistical significance was found in inter‐clutch variation between these two types of hosts. Our results suggest that hosts lay eggs with a low level of intra‐clutch variation without the necessity of a high level of inter‐clutch variation simultaneously as predicted by the egg signature hypothesis. This study may further indicate that selection pressures from evolutionarily recent parasites can drive individual‐based differences in an anti‐parasitism strategy.     

Genetic analysis of Trichuris suis and Trichuris trichiura recovered from humans and pigs in a sympatric setting in Uganda

The whipworms Trichuris trichiura and Trichuris suis in humans and pigs, respectively, are believed to be two different species yet closely related. Morphologically, adult worms, eggs and larvae of the two species are indistinguishable. The aim of this study was to examine the genetic variation of Trichuris sp. mainly recovered from natural infected pigs and humans. Worm material isolated from humans and pigs living in the same geographical region in Uganda were analyzed by PCR, cloning and sequencing. Measurements of morphometric characters were also performed. The analysis of the ITS-2 (internal transcribed spacer) region showed a high genetic variation in the human-derived worms with two sequence types, designated type 1 and type 2, differing with up to 45%, the type 2 being identical to the sequence found in pig-derived worms. A single human-derived worm showed exclusively the type 2-genotype (T. suis-type) and three cases of 'heterozygote' worms in humans were identified. However, the analysis showed that sympatric Trichuris primarily assorted with host origin. Sequence analysis of a part of the genetically conserved β-tubulin gene confirmed two separate populations/species but also showed that the 'heterozygote' worms had a T. suis-like β-tubulin gene. A PCR-RFLP on the ITS-2 region was developed, that could distinguish between worms of the pig, human and 'heterozygote' type. The data suggest that Trichuris in pigs and humans belong to two different populations (i.e. are two different species). However, the data presented also suggest that cross-infections of humans with T. suis takes place. Further studies on sympatric Trichuris populations are highly warranted in order to explore transmission dynamics and unravel the zoonotic potential of T. suis.     

First report of Angiostrongylus vasorum (French heartworm) in red foxes (Vulpes vulpes) on Prince Edward Island

Objective To identify first-stage nematode larvae (L1) recovered from a red fox scat sample and adult female worms recovered from 2 red fox lungs at necropsy, using published molecular methods to confirm a morphological diagnosis of Angiostrongylus vasorum (French heartworm).Animal Red fox (Vulpes vulpes).Procedure Nematode larvae recovered from a Baermann examination survey of wild canid scats (n = 101) conducted from January 2017 to August 2020, were identified by size and morphology and subjected to PCR and DNA sequencing of the small subunit (SSU) rRNA gene, the large subunit (LSU) rRNA gene, or the second internal transcribed spacer (ITS2). In addition, these techniques were applied to adult female worms recovered from the heart/lungs of 2 red foxes (obtained from PEI trappers and stored frozen at −20°C since December of 2018 and 2020).Results Size and morphology of L1 recovered by Baermann examination from a wild canid scat sample (presumed to be red fox) collected near Montague, PEI and adult female worms recovered at necropsy from 2 red fox carcasses were identified as A. vasorum. Molecular analysis confirmed the larvae and adult worms were A. vasorum.Conclusion These findings indicated that A. vasorum has become endemic in the red fox population on PEI.Clinical relevance Angiostrongylus vasorum infection is potentially fatal in dogs. Veterinarians and regional diagnostic laboratories in the Maritime provinces should consider the possibility of A. vasorum infection in dogs with clinical signs of cardiopulmonary and/or central nervous system disease or bleeding disorders.     

Life cycle of the tick Amblyomma parvum Aragão, 1908 (Acari: Ixodidae) and suitability of domestic hosts under laboratory conditions

Amblyomma parvum is a widespread Neotropical tick found on several domestic animals and is known to harbor a Rickettsia species of yet unknown pathogenicity. However its life cycle on, and suitability of, several of these hosts has not been described. In this work the biology of A. parvum is presented when fed on seven domestic hosts (chicken, dog, rabbit, horse, guinea pig, cattle and goat). The complete life cycle of the tick varied from 96.8 to 102 days. Highest engorgement weight of larvae was from ticks fed on horses and that of nymphs from guinea pigs. Highest larval yield was from guinea pigs and that of nymphs from horses. Engorged female and egg mass weights, yield and conversion of female weight to eggs rates were the highest in dog ticks and the lowest in goat ticks. The highest egg hatching rate was seen in ticks from dogs and the lowest in ticks from cattle. Overall it was seen that dogs were the best host for adult A. parvum ticks, and guinea pigs for immatures. Horses were also shown to be a good host for all tick stages. It can thus be affirmed that A. parvum is a host generalist tick, and its distribution is probably determined by environmental requirements rather than by hosts.     

A comparison of the protein constituents of the major body compartments of the dog roundworm, Toxocara canis

An analysis of the protein profiles of intact worms and isolated tissues of adult male and female Toxocara canis worms was conducted. Soluble proteins recovered from homogenized whole specimens and dissected tissues (body wall, reproductive tract, esophagus and intestine) of T. canis adults from several different canine hosts were separated by size using gradient sodium dodecyl sulfate electrophoresis (SDS-PAGE) and visualized with silver staining. SDS-PAGE profiles of worms from different hosts were found to be virtually identical irrespective of sex or tissue type. Recovered proteins ranged in size from 3.4 to 325 kDa. As expected, variations existed between the protein profiles of different body tissues, with only slight variations between the sexes. The largest number of recovered proteins was present in the female reproductive tract extracts.     

Raccoon dogs (Nyctereutes procyonoides) are the new natural definitive hosts of Metagonimus hakubaensis

The definitive hosts of Metagonimus hakubaensis are reported to be hamsters, rats, mice, dogs, cats, chickens, and quails in experimental infection and Japanese water shrews in natural infection. Here we report that raccoon dogs are new natural definitive hosts of M. hakubaensis, based on morphological and molecular analyses of Metagonimus flukes collected from the host species from Aomori Prefecture, Japan. Moreover, M. hakubaensis recovered from raccoon dogs showed higher fecundity than those recovered from Japanese water shrews. Therefore, raccoon dogs were considered as a more suitable natural definitive host of M. hakubaensis than Japanese water shrews.     

Antibody and blood leukocyte response in Rhipicephalus sanguineus (Latreille, 1806) tick-infested dogs and guinea pigs

The dog is considered to be the natural host of Rhipicephalus sanguineus and is unable to develop appreciable resistance even after repeated feedings. The guinea pig develops strong resistance after one infestation with adult ticks. Antibody (IgG) titres against tick salivary gland antigens (SGAs) and blood leukocyte numbers in dogs and guinea pigs undergoing experimental R. sanguineus tick infestations were measured to detect a possible correlation with susceptibility or resistance of hosts. Since infested dogs develop an immediate hypersensitivity reaction to R. sanguineus antigens, total and anti-R. sanguineus SGA IgE levels were also measured in this host species. IgG and IgE antibody levels were determined by an enzyme-linked immunosorbent assay (ELISA) along three consecutive infestations of both hosts. Most dogs and guinea pigs displayed low IgG levels against R. sanguineus SGAs, though marked differences in individual response were observed. Although dog's total serum IgE levels increased significantly after infestations, no change in the amount of anti-salivary gland IgE was detected. Total and differential blood cell counts were determined in dogs and guinea pigs during primary and secondary infestation. In dogs, a tertiary infestation and a subsequent higher infestation level were also evaluated. Infested dogs did not display any alteration in blood leukocyte counts throughout the experiment. Guinea pigs, on the other hand, developed a significant basophilia during primary infestation which increased further during secondary infestation. These data reveal similarities and differences in the reactions of resistant and non-resistant hosts to ticks. They contribute for the understanding of such host-parasite relationships and will hopefully aid in the development of immune control of ticks.     

Assessment of the qualitative variation in bacterial microflora among compartments of the intestinal tract of dogs by use of a molecular fingerprinting technique

OBJECTIVE: To evaluate the qualitative variation in bacterial microflora among compartments of the intestinal tract of dogs by use of a molecular fingerprinting technique. ANIMALS: 14 dogs (similarly housed and fed identical diets). PROCEDURE: Samples of intestinal contents were collected from the duodenum, jejunum, ileum, colon, and rectum of each dog. Bacterial DNA was extracted from the samples, and the variable V6 to V8 region of 16S ribosomal DNA (gene coding for 16S ribosomal RNA) was amplified by use of universal bacterial primers; polymerase chain reaction amplicons were separated via denaturing gradient gel electrophoresis (DGGE). Similarity indices of DGGE banding patterns were used to assess variation in the bacterial microflora among different compartments of the intestine within and among dogs. Bacterial diversity was assessed by calculating the Simpson diversity index, the Shannon-Weaver diversity index, and evenness. RESULTS: DGGE profiles indicated marked differences in bacterial composition of intestinal compartments among dogs (range of similarity, 25.6% to 36.6%) and considerable variation among compartments within individual dogs (range of similarity, 36.7% to 579%). Similarities between neighboring intestinal compartments were significantly greater than those between non-neighboring compartments. Diversity indices for the colon and rectum were significantly higher than those of the duodenum, jejunum, and ileum. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that the different intestinal compartments of individual dogs appear to host different bacterial populations, and these compartmental populations vary among dogs. In dogs, fecal sample analysis may not yield accurate information regarding the composition of bacterial populations in compartments of the gastrointestinal tract.     

A comparative histochemical and morphometric study of canine skeletal muscle.

The purpose of this study was to determine whether there were differences in skeletal muscle properties in the hindlimb muscles of different types of dogs. Muscle samples were obtained from the gracilis, sartorius cranial head, sartorius caudal head and tibialis anterior muscles of mixed-breed and hound-type dogs and Beagles. Fiber type, fiber size and capillary morphometry determinations of each muscle from each dog were made from sections stained for myofibrillar ATPase activity. Individual animals were bilaterally symmetric for all measured variables. Fiber type, fiber size and capillary geometry varied between dogs of a given type and muscles within a given dog. There were no differences between dog types for fiber type or fiber size; significant variation in log(muscle)/log(body) mass ratios between dog types was observed for all muscles. The results indicate that for a given muscle, significant variation can occur in skeletal muscle characteristics between different types of dogs and that these differences can be independent of differences in exercise history.     

No evidence for enzootic plague within black-tailed prairie dog (Cynomys ludovicianus) populations

Yersinia pestis, causative agent of plague, occurs throughout the western United States in rodent populations and periodically causes epizootics in susceptible species, including black-tailed prairie dogs (Cynomys ludovicianus). How Y. pestis persists long-term in the environment between these epizootics is poorly understood but multiple mechanisms have been proposed, including, among others, a separate enzootic transmission cycle that maintains Y. pestis without involvement of epizootic hosts and persistence of Y. pestis within epizootic host populations without causing high mortality within those populations. We live-trapped and collected fleas from black-tailed prairie dogs and other mammal species from sites with and without black-tailed prairie dogs in 2004 and 2005 and tested all fleas for presence of Y. pestis. Y. pestis was not detected in 2126 fleas collected in 2004 but was detected in 294 fleas collected from multiple sites in 2005, before and during a widespread epizootic that drastically reduced black-tailed prairie dog populations in the affected colonies. Temporal and spatial patterns of Y. pestis occurrence in fleas and genotyping of Y. pestis present in some infected fleas suggest Y. pestis was introduced multiple times from sources outside the study area and once introduced, was dispersed between several sites. We conclude Y. pestis likely was not present in these black-tailed prairie dog colonies prior to epizootic activity in these colonies. Although we did not identify likely enzootic hosts, we found evidence that deer mice (Peromyscus maniculatus) may serve as bridging hosts for Y. pestis between unknown enzootic hosts and black-tailed prairie dogs.     

Species spectrum, diversity profile and infection indices of helminth parasite fauna of Chirruh snowtrout, Schizothorax esocinus (Heckel) in lake ecosystems of Kashmir Himalayas-Do similarity and host-parasite associations arise?

In order to assess the species richness and diversity profile of helminth parasite fauna in an endemic fish, an investigation was carried out in two urban and two rural lakes of Kashmir. Overall nine species of helminth parasites were observed in four lakes. Of these three were autogenic and six were allogenic. Heteroxenous parasite species were more in number than monoxenous species. Results showed significant differences in heteroxenous / monoxenous ratio between different lakes. Core species (Prevalence?>?20) were only found in hypertrophic lake (Anchar Lake). Overall, majority of helminth species were either secondary or satellite species. Prevalence of some helminth parasites showed significant differences in different lakes. In addition mean intensity showed significant differences between autogenic and allogenic parasites (P?<?0.05). Principle Component Analysis based on prevalence showed that Anchar Lake was strongly associated with most of helminth parasites. Diversity indices showed significant variation between different lakes. Maximum helminth species per host was in Anchar Lake. Finally we concluded that helminth parasite fauna showed significant differences in species richness and infection indices between different lakes. Diversity profile was higher in Anchar Lake in comparison to other three lakes. The results clearly show that environmental features of lake ecosystems have got an impact on distribution pattern of helminth parasites in S. esocinus. We suggest comparative parasitological study should be taken between different species of fish in order to have a clear picture regarding the species composition of helminth species in this region. Also we need to characterize the species spectrum of parasitic worms in fish of freshwater bodies of this region as well as other similar type of climatic zones because parasite fauna is an integral part of the inventory of biodiversity and as possible regulators of host populations in aquatic ecosystems.     

Glycosylated haemoglobin assay of canine blood samples

A method for assaying canine glycosylated haemoglobin was evaluated. The method is a turbidimetric inhibition immunoassay and the final reaction is bichromatically measured using a multichannel automatic analyser. Within-run coefficients of variation (2.07 to 4.46 per cent) were permissible, but between-run coefficients of variation (2.10 to 8.25 per cent) were slightly more elevated. The detection limit of this assay is 0.052 per cent. A sample dilution of 10 microliters of sample and 500 microliters of haemolysing reagent is recommended for routine analysis of canine blood samples. A normal reference interval of 1.39 +/- 0.70 per cent was obtained from the glycosylated haemoglobin analysis in 82 healthy dogs and no statistically significant differences in relation to age or gender were observed. Some changes in glycosylated haemoglobin concentrations were noted throughout the ovarian cycle, although differences between dogs were evident. Since this assay specifically measures the glycosylated haemoglobin content in canine blood samples, it could be very useful for monitoring diabetic dogs.