Isolation and identification of Salmonella spp. from red foxes (Vulpes vulpes) and badgers (Meles meles) in northern Italy

Background

Salmonella spp. have been isolated from a wide range of wild animals. Opportunistic wild carnivores such as red foxes (Vulpes vulpes) and badgers (Meles meles) may act as environmental indicators or as potential sources of salmonellosis in humans. The present study characterizes Salmonella spp. isolated from the intestinal contents of hunted or dead red foxes (n = 509) and badgers (n = 17) in northern Italy.

Findings

Thirty-one strains of Salmonella belonging to 3 Salmonella enterica subspecies were isolated. Fourteen different serovars of S. enterica subsp. enterica were identified, among which were serovars often associated with human illness.

Conclusions

Wild opportunistic predators can influence the probability of infection of both domestic animals and humans through active shedding of the pathogen to the environment. The epidemiological role of wild carnivores in the spread of salmonellosis needs to be further studied.     

Spirocerca lupi isolated from gastric lesions in foxes (Vulpes vulpes) in Sicily (Italy)

Spirocerca lupi (Rudolphi 1809) is a cosmopolitan nematode of dogs and wild carnivores. In the past it has been reported in Italy, mainly in southern regions and in Sicily, where the parasite was observed in foxes in 2005. The parasite typically produces nodular masses in the oesophagus and thoracic aorta. During the 2003-2004 hunting season, the authors investigated a total of 55 foxes (Vulpes vulpes) hunted or killed by car accidents in the provinces of Palermo and Agrigento. All the foxes were subjected to necropsy and 6 (9.16%) had S. lupi nodules located exclusively in the gastric wall. The nature of the nodules was determined by opening them and detecting the nematodes inside, which were identified as S. lupi. Some of the nodules were characterized anatomopathologically and histopathologically. The formation of the parasitic nodule in the stomach only suggests a deviation from the route commonly followed by the nematode to reach the oesophagus, the elective anatomical site for completion of its lifecycle. This survey gives a contribution to the epidemiology of this parasite which is severely outdated in Italy and highlights some distinctive features of the life cycle and parasite migration.     

A serosurvey of Hepatozoon canis and Ehrlichia canis antibodies in wild red foxes (Vulpes vulpes) from Israel

A seroepidemiological survey was conducted to investigate the prevalence of antibodies reactive with Ehrlichia canis and Hepatozoon canis antigens in free-ranging red foxes (Vulpes vulpes) in Israel. Of 84 fox sera assayed, 36% were seropositive for E. canis by the indirect fluorescent antibody (IFA) test and 24% were positive for H. canis using an enzyme-linked immunosrbent assay (ELISA). Canine ehrlichiosis and hepatozoonosis appear to be endemic in the wild red fox populations in Israel, and foxes may serve as a reservoir for infection of domestic dogs and other wild canine species.     

Occurrence of Giardia and Cryptosporidium in Norwegian red foxes (Vulpes vulpes)

Faecal samples from 269 Norwegian wild red foxes (Vulpes vulpes) shot during the hunting season (October-April) in 2002-2004 were examined for the presence of Giardia and Cryptosporidium. Cryptosporidium oocysts were detected in samples from 6 (2.2%) of the foxes, and Giardia cysts in 13 (4.8%) of the foxes. The prevalence of Giardia infection was significantly higher in juvenile male foxes than in adult male foxes, but no other significant differences between age and sex were found. No significant differences in prevalence related to geographical origin of animals were found. Insufficient nucleated Cryptosporidium oocysts were isolated for successful PCR, but genotyping of Giardia duodenalis isolates from seven foxes demonstrated a high degree of heterogeneity amongst them, with all isolates belonging to the zoonotic Assemblages A and B.     

Presence of Trichinella spiralis in free-living red foxes (Vulpes vulpes) in Sweden related to trichinella infection in swine and man

One thousand one hundred and fifty-one free-living foxes (Vulpes vulpes) from different parts of Sweden were investigated. Totally 19.6 % were infected with trichinella. Infected foxes were found in all counties except the geographically isolated island of Gotland. In the different counties 6–48 % of investigated foxes were infected.Trichinella was more common in old foxes than in young, 40 % and 11 %, respectively. Regarding male and female, however, the frequency was the same.The number of trichinella per g of muscle varied between 0.05 and 200. Less than 1.0 trichinella larva per g muscle was established in 27.3 % of the foxes, between 1 and 49.9 trichinella larvae in 69.3 % and 50 or more trichinella larvae per g muscle in 3.4 % of the foxes. The number of trichinella larvae per infected fox was roughly the same in both sexes as well as in different age groups.The potential danger of transmitting trichinella from foxes and other carnivores to swine and man is pointed out.The high frequency of trichinella in foxes, 19.6 %, was compared to the very low frequency in swine, 0.00018 %, and in man, 0.00003 %.     

First molecular detection of Pneumocystis spp. in red foxes (Vulpes vulpes linnaeus, 1758) and raccoon dogs (Nyctereutes procyonoides gray, 1834)

Fungal organisms of the genus Pneumocystis may cause Pneumocystis pneumonia (PCP) in humans, but also domestic and wild mammals. Almost every animal species hosts its own genetically distinct Pneumocystis species, however information is sparse. In this study, 62 red foxes (Vulpes vulpes) and 37 raccoon dogs (Nyctereutes procyonoides) were collected in North-East Germany. The lung tissues of the animals were analysed by a new designed specific pan-Pneumocystis mtLSU rRNA gene PCR and sequencing. With this PCR, detection and discrimination of all known Pneumocystis spp. in a single step should be possible. This first detection of Pneumocystis spp. in 29/62 (46.8%) red foxes and 29/37 (78.4%) raccoon dogs indicated, that they harbour two dissimilar strains, as seen by specific single nucleotide position changes (SNPs). Nevertheless, five samples with contrary SNPs showed a probable inter-species transmission.     

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.     

Evaluation of a serological test (indirect ELISA) for the diagnosis of sarcoptic mange in red foxes (Vulpes vulpes)

Sarcoptic mange occurs in many parts of the world and is common in populations of domestic and wild canids, including red foxes (Vulpes vulpes). In recent years, an indirect antibody enzyme-linked immunosorbent assay (ELISA), with higher sensitivity and specificity than traditional diagnostic methods, has been successfully applied in the diagnosis of sarcoptic mange in dogs. The same ELISA has also demonstrated specific antibodies to Sarcoptes scabiei in experimentally infected red foxes. The aim of this study was to evaluate the indirect ELISA when used to detect antibodies to S. scabiei in field sera from Swedish red foxes. One cohort of both infected and non-infected red foxes (cohort 1; n = 88), and one cohort of apparently non-infected foxes (cohort 2; n = 67) were examined for skin lesions and presence of S. scabiei by thorough visual examination at autopsy and skin scrapings. Samples of blood-tinted body liquid from the abdomen or thorax cavity were collected and analysed by the indirect ELISA. The relative sensitivity and specificity of the ELISA at different cut-offs (OD values) were estimated by comparing the test results to the infection status as determined by examination and skin scrapings. The highest combination of relative sensitivity and specificity, calculated based on cohort 1, was 95.4 and 100.0%, respectively. These estimates were constant for cut-offs 0.150-0.225, which included the cut-off based on the mean plus three standard deviations of test results from cohort 2 (0.165). It is concluded that this test can be useful in diagnosis and epidemiological studies of S. scabiei infection in red foxes.     

Prevalence of Trichinella larvae and extra-intestinal nematodes in Norwegian red foxes (Vulpes vulpes)

A survey of the parasitic fauna of the Norwegian red fox (Vulpes vulpes) population was carried out in 1994/1995 and 2002-2005. All foxes were killed during the licensed hunting season from October to April and, in total, 393 animals from all regions of the country were examined. The present study details the results of extra-intestinal nematode and Trichinella larvae examinations. All individuals were examined for Trichinella, using routine digestion methods. Parasitological examination of the internal organs of some of the foxes also identified a number of different extra-intestinal nematodes. The following prevalences were identified (number positive/number foxes examined): Trichinella larvae 19/393 (4.8%); Capillaria b?hmi (C. b?hmi) 88/174 (51%); Capillaria aerophila (C. aerophila) 160/181 (88%); Crenosoma vulpis (Cr. vulpis) 105/181 (58%) and Capillaria plica (C. plica) 81/154 (53%). No evidence of Angiostrongylus vasorum infection was found. The 19 different Trichinella isolates were species typed by PCR and sequence analysis; 18 isolates were identified as Trichinella nativa and one as Trichinella britovi. A wide geographical distribution of the parasites was seen. The following exceptions were recorded: C. b?hmi, the prevalence of which was significantly lower in northern Norway (6%) compared to other regions (central Norway, eastern Norway and southern and western Norway; 52-57%). There was a significantly higher prevalence of Trichinella infection in eastern Norway (8.1%), when compared with the rest of the country (0.6%). Cr. vulpis prevalence was significantly higher in central Norway (83%) than in other regions (41-56%). There were no significant differences in age and sex distribution of the parasites with the exception of Cr. vulpis where juvenile foxes had a greater likelihood of infection. The data also indicated that adult foxes were more commonly infected with Trichinella larvae (5.8%) than juveniles (3.3%) (no statistical significance).     

Direct high-resolution genotyping of Toxoplasma gondii in arctic foxes (Vulpes lagopus) in the remote arctic Svalbard archipelago reveals widespread clonal Type II lineage

Characterization of Toxoplasma gondii genotypes in hosts living in remote, isolated regions is important for elucidating the population structure and transmission mode of this parasite. Herein, we report the results of direct genotyping of T. gondii in brain tissue of arctic foxes (Vulpes lagopus) from the remote, virtually cat-free, high arctic islands of Svalbard. DNA extracts from brains of 167 seropositive arctic foxes (including four cases of fatal toxoplasmosis) and 11 seronegative arctic foxes were genotyped at 10 loci (SAG1, SAG2, SAG3, BTUB, GRA6, L358, c22-8, c29-2, PK1, and Apico) using the polymerase chain reaction-restriction fragment length polymorphism method. Of the 167 samples from seropositive foxes (including toxoplasmosis cases), 31 were genotyped at all 10 loci and 24 were genotyped at four to nine loci. To ensure confidence in T. gondii strain genotyping, samples for which less than four loci were genotyped were not considered positive. None of the 11 samples from seronegative foxes was positive for the 10 markers. Of the 55 samples that genotyped positively, 46 were of the Type II strain, 7 were of the Type III strain, and 2 were of atypical T. gondii strains. Five representative samples of the three genotypes were sequenced at loci SAG2, SAG3, GRA6, PK1, and UPRT-1. The DNA sequences confirmed the genotyping results. This study shows that the archetype Type II T. gondii strain, which is most widely distributed in North America and Europe, also predominates in arctic foxes on the Svalbard archipelago. This suggests that the T. gondii at this location originate from continental Europe and that transmission may be mediated by migrating birds. This study highlights the significance of long-distance transport of T. gondii and demonstrates that high-resolution genotyping protocols are useful for direct genetic studies of T. gondii when isolation of live parasites is infeasible.     

Potential application of serological tests on fluids from carcasses: detection of antibodies against Toxoplasma gondii and Sarcoptes scabiei in red foxes (Vulpes vulpes)

Background

Serological surveys for disease investigation of wild animal populations require obtaining blood samples for analysis, which has logistic, ethic and economic difficulties. Applying serological test to fluids collected from dead animals is an alternative. The aim of this study was to assess if antibodies could be detected in two types of fluids collected from 56 carcasses of red foxes (Vulpes vulpes): pleural fluid and lung extract.

Findings

In 22 (39%) foxes antibodies against Sarcoptes scabiei were detected in both fluid types by ELISA and Western blot. In 46 (82%) foxes, antibodies against Toxoplasma gondii were detected in pleural fluid and in 41 (73%) in lung extract applying a Toxo-screen test (DAT). Antibodies were still detectable in the same fluids kept at room temperature for 28 days, although in fewer foxes (16 and 14 foxes tested for T. gondii in lung extract and pleural fluid respectively; and 1 and 4 tested for S. scabiei in lung extract and pleural fluid respectively.

Conclusions

These results indicate the potential utility of using fluids from carcasses for antibody screening of wild animals at the population level.     

Extraintestinal nematode infections of red foxes (Vulpes vulpes) in Hungary

A survey was carried out to investigate the prevalence and worm burden of extraintestinal nematodes in 100 red foxes (Vulpes vulpes) of Hungary. The overall prevalence of nematode infections of the respiratory tract was 76%. Eucoleus aerophilus (Capillaria aerophila) was the predominant species (66%), followed by Crenosoma vulpis (24%), Eucoleus (Capillaria) b?hmi (8%) and Angiostrongylus vasorum (5%). Pearsonema (Capillaria) plica was found in 52% of the urinary bladders. In 3% of the foxes, Trichinella britovi was present in muscle samples. The high prevalence of lungworms and P. plica and the fox colonisation in urban areas may enhance the prevalence of these nematode infections in domestic dogs and cats, and the flow of T. britovi from the sylvatic cycle to the domestic cycle, enhancing the risk of infections in humans.     

Prevalence of antibodies against Toxoplasma gondii and Neospora caninum in Hungarian red foxes (Vulpes vulpes)

In the present study we have investigated the seroprevalence to the protozoan parasites Toxoplasma gondii and Neospora caninum in 337 red foxes (Vulpes vulpes) from 16 out of 19 counties in Hungary. The foxes were originally collected within a National vaccination program against rabies. Antibodies to T. gondii were detected in as many as 228 (68%) of the foxes using a commercial direct agglutination test (DAT). In an indirect iscom ELISA, five foxes (1.5%) were positive for antibodies against N. caninum. The high prevalence of foxes positive for T. gondii might be explained by the widespread occurrence of the parasite in the diet of foxes. As a contrast, latent infections of N. caninum among red foxes in Hungary are much less common.     

A Study on the Predilection Sites of Trichinella spiralis Muscle Larvae in Experimentally Infected Foxes (Alopex lagopus,Vulpes vulpes)

Kapel, Chr. M., Sv. Aa. Henriksen, H. H. Dietz, P. Henriksen, P. Nansen: A study on the predilection sites of Trichinella spiralis muscle larvae in experimentally infected foxes (Alopex lagopus, Vulpes vulpes). Acta vet. scand. 1994, 35, 125-132.–Studies were carried out on the predilection sites of Trichinella spiralis muscle larvae in experimentally infected arctic foxes (Alopex lagopus) and silver foxes (Vulpes vulpes) reared in cages. The highest number of larvae per gramme tissue was found in the muscles of the legs, eyes, diaphragm, and tongue. The 2 fox species showed no significant differences with regard to predilection sites.     

Characterization of widespread canine leishmaniasis among wild carnivores from Spain

Visceral Leishmaniasis (VL) is an emerging zoonotic parasitic disease caused by Leishmania infantum in Mediterranean countries, with sand flies (Phlebotomus spp.) as vectors and dogs as the main domestic reservoir. The role of wild carnivores in the epidemiology of leishmaniasis is still controversial. In order to determine the prevalence of natural infection with L. infantum in wild carnivores from Spain, we analyzed 217 samples by PCR and western blotting and used restriction fragment length polymorphism (RFLP) to compare the patterns present in wild carnivores with those of domestic dogs from the same areas. DNA of the parasite was detected in spleen or blood samples from 35 (16.12%) analyzed wild carnivores, including 8 of 39 (20.5%) wolves (Canis lupus), 23 of 162 (14.1%) foxes (Vulpes vulpes), 2 of 7 (28.6%) Egyptian mongooses (Herpestes ichneumon), 1 of 4 genets (Geneta geneta), and 1 of 4 Iberian lynxes (Lynx pardinus). No significant sex or age differences in prevalence were observed in wolves and foxes (P>0.05), but there was a significant difference among regions in foxes (P<0.05). A total of 12 PCR-RFLP patterns were found in foxes, 6 in wolves, 4 in dogs, 2 in Egyptian mongooses and 1 in lynx and genet. RFLP patterns differed between dogs and foxes in the two areas where they could be compared. This is the first study of canine leishmaniasis in wild canids and other carnivores from different regions of Spain by PCR. The prevalence of infection indicates the existence of natural infection in apparently healthy wild carnivore populations, and our results are suggestive of a sylvatic cycle independent of dogs.     

Seroprevalences of Toxoplasma gondii and Neospora caninum in Swedish red foxes (Vulpes vulpes).

The prevalences of antibodies to the protozoan parasites Toxoplasma gondii and Neospora caninum were investigated by the direct agglutination test (DAT) and ELISA, respectively, in 221 red foxes (Vulpes vulpes) from different parts of Sweden. A total of 84 (38%) of the analysed sera had antibodies to T. gondii, but none of the foxes had antibodies to N. caninum. The results indicate that T. gondii infection is fairly common in Swedish red foxes and that the infection is present in most parts of the country. They also show that N. caninum is not widespread as a latent infection among red foxes in Sweden.     

Faecal virome of red foxes from peri-urban areas

Red foxes (Vulpes vulpes) are the most abundant carnivore species in the Northern Hemisphere. Since their populations are well established in peri-urban and urban areas, they represent a potential reservoir of viruses that transmit from wildlife to humans or domestic animals. In this study, we evaluated the faecal virome of juvenile and adult foxes from peri-urban areas in central Croatia. The dominating mammalian viruses were fox picobirnavirus and parvovirus. The highest number of viral reads (N = 1412) was attributed to a new fox circovirus and complete viral genome was de novo assembled from the high-throughput sequencing data. Fox circovirus is highly similar to dog circoviruses identified in diseased dogs in USA and Italy, and to a recently discovered circovirus of foxes with neurologic disease from the United Kingdom. Our fox picobirnavirus was more closely related to the porcine and human picobirnaviruses than to known fox picobirnaviruses.     

Behavioural Changes in Free-ranging Red Foxes (Vulpes vulpes) due to Sarcoptic Mange

Sarcoptic mange (Sarcoptes scabiei var. vulpes) reached Scandinavia in the mid-1970’s and is mainly prevalent among red foxes (Vulpes vulpes) (Borg 1987). In the laboratory, foxes succumb 2–4 months after being infested, and it is commonly thought that carriers in the wild exhibit an abnormal behaviour and quickly die (Mörner & Christensson 1984, Holt & Berg 1990). However, there is a lack of comparative data from free-ranging animals to contribute to the ecology of the species in general and ta supplement the content of the above selected references in particular (e.g. Plowright 1988). Using telemetry studies, I have compared the behaviour in winter of 2 mange infested and 2 healthy red foxes. The work took place in 1987 and 1990 in a boreal area adjacent to farmland in central Norway (63° 20′N 10° 45′E).     

Natural and experimental Salmonella Typhimurium infections in foxes (Vulpes vulpes)

The red fox (Vulpes vulpes) can be considered as a relevant indicator species for Salmonella in the local environment and Salmonella faecal carriage was investigated in 215 red foxes in Norway shot during the winters 2002/2003 and 2003/2004. Fourteen (6.5%) of the foxes carried Salmonella. Four isolates were determined as serovars Kottbus (n=2) and Hessarek (n=2) of Salmonella enterica subspecies enterica, and one as S. enterica subspecies IIIb:61:k:1,5,(7). The remaining nine isolates were S. enterica subspecies enterica serovar Typhimurium 4,12:i:1,2 and all displayed the same pulsed-field gel electrophoresis (PFGE) profile designated A2. This serovar regularly causes disease outbreaks amongst small passerine birds during winter and most of these outbreaks are associated with the PFGE profile A2. The results strongly indicated that the Salmonella Typhimurium infections in red foxes were primarily acquired through ingestion of infected small passerines. To investigate the capability of the A2 strain to establish a true intestinal infection in the fox an inoculation experiment with an A2 isolate from small passerines was carried out in farmed silver foxes (V. vulpes). The experiment also included one strain with an uncommonly occurring profile (X201) from small passerines. To highlight possible differences in capability of the two inoculation strains to pass the acid gastric juice in the fox, in vitro studies of their acid tolerance was carried out. Also their catalase activity and biofilm production were studied. All three foxes inoculated with the A2 strain developed sub-clinical intestinal infection of 2 weeks duration, whereas none of the three foxes inoculated with the X201 strain shed this bacterium. The X201 strain displayed a much lower capability, than the A2 strain, to survive at pH 3 in vitro. The low acid tolerance probably made it difficult for the X201 strain to pass the stomach and establish an intestinal infection in the experimental foxes. Reduced catalase activity and biofilm production were found for the X201 strain, indicating that the low acid tolerance was caused by a defect in the stationary-phase stress response system.