Release of Angiostrongylus cantonensis larvae from live intermediate hosts under stress.

The metastrongyloid nematode Angiostrongylus cantonensis causes eosinophilic meningitis in a variety of homeothermic hosts including humans. Third-stage infectious larvae develop in gastropods as intermediate hosts. Humans are usually infected by intentional or incidental ingestion of an infected mollusk or paratenic host (poikilothermic vertebrates and invertebrates). The infection may also hypothetically occur through ingestion of food or water contaminated by third-stage larvae spontaneously released from gastropods. Larvae are thought to be released in greater numbers from the intermediate host exposed to stress. This study aimed to compare larval release from stressed with unstressed gastropods. Experimentally infected Limax maximus and Lissachatina fulica were exposed to a stress stimulus (shaking on an orbital shaker). The mucus was collected before and after the stress and examined microscopically and by qPCR for the presence of A. cantonensis larvae and their DNA. In the case of L. maximus, no larvae were detected microscopically in the mucus, but qPCR analysis confirmed the presence of A. cantonensis DNA in all experimental replicates, without clear differences between stressed and non-stressed individuals. In contrast, individual larvae of A. cantonensis were found in mucus from Li. fulica after stress exposure, which also reflects an increased number of DNA-positive mucus samples after stress. Stress stimuli of intensity similar to the transport or handling of mollusks can stimulate the release of larvae from highly infected intermediate hosts. However, these larvae are released in small numbers. The exact number of larvae required to trigger neuroangiostrongyliasis is unknown. Therefore, caution is essential when interacting with potential intermediate hosts in regions where A. cantonensis is endemic.

Rat lungworm survives winter: experimental overwintering of Angiostrongylus cantonensis larvae in European slugs.

The rat lungworm Angiostrongylus cantonensis is a metastrongyloid nematode that causes neurological disorders in its accidental hosts, including humans. This invasive pathogen is native to Southeast Asia and adjacent regions and is gradually expanding its distribution to tropical and subtropical areas with new foci discovered near temperate regions. The parasite has a complex life cycle with a range of gastropods serving as intermediate hosts. A broad spectrum of poikilotherm vertebrates and invertebrates can serve as paratenic hosts. Since it has already been demonstrated that other, non-zoonotic metastrongyloids can survive in their intermediate hosts during the winter, the aim of our study was to evaluate the survival of A. cantonensis third-stage larvae in experimentally infected slugs (Limax maximus) kept at 4.5­7°C for 60 days. Third-stage larvae of A. cantonensis survived the period of low temperature and remained capable of infecting definitive hosts (laboratory rats) afterwards, even though their numbers dropped significantly. These results suggest that further spread to higher latitudes or altitudes is possible in areas with sufficient abundance of definitive hosts, since low winter temperatures are not necessarily an obstacle to the spread of the parasite.

Range of metastrongylids (superfamily Metastrongyloidea) of public health and veterinary concern present in livers of the endemic lizard Gallotia galloti of Tenerife, Canary Islands, Spain.

BACKGROUND: Endemic lizards of the genus Gallotia are of high ecological value to the terrestrial ecosystem of the archipelago of the Canary Islands, being potent seed spreaders as well as an important component of the diet of other vertebrates. The endemic lizard Gallotia galloti in Tenerife has recently been reported to be a paratenic host of Angiostrongylus cantonensis, an invasive metastrongylid with zoonotic potential that is associated with rats as definitive hosts. However, microscopic examination of G. galloti tissue samples also revealed the presence of other metastrongylid larvae inside granulomas on the liver of this reptile. The aim of this study was to investigate the presence of helminths other than A. cantonensis in tissues of G. galloti from Tenerife. METHODS: A multiplex-nested PCR targeting the internal transcribed spacer 1 was designed that enabled the species-specific detection of A. cantonensis, Angiostrongylus vasorum, Aelurostrongylus abstrusus, Crenosoma striatum and Crenosoma vulpis. Liver samples from 39 G. galloti were analysed. RESULTS: Five metastrongylids were detected: A. cantonensis (15.4% of samples analysed), A. vasorum (5.1%), Ae. abstrusus (30.8%), C. striatum (30.8%) and undetermined metastrongylid sequences (12.8%). Co-infection was highly prevalent among the lizards which tested positive. CONCLUSIONS: The study provides a new specific tool for the simultaneous detection of a range of metastrongylids of veterinary importance as well as new data on the circulation of metastrongylids in an ecosystem dominated by lizards.

Zoonotic Bacteria in Anolis sp., an Invasive Species Introduced to the Canary Islands (Spain).

Lizards belonging to the genus Anolis are native to America and have been introduced in many parts of the world. In this work, a gastrointestinal microbiological analysis from Anolis sp. introduced to Tenerife, Canary Island, was carried out. A total of 74 individuals were analyzed by culture and molecular tools. Pseudomonas spp. was the most prevalent bacteria isolated (64.3%), followed by enteropathogenic Escherichia coli with at least one of the investigated virulent genes (stx1, stx2, and eae) (44.6%). The stx2 gene was more prevalent which differs to that reported in other reptiles, probably due to wastewater transmission. Campylobacter spp. was detected in 32.4% of the animals, highlighting the detection of C. jejuni and C. fetus by their relevance to public health. The zoonotic Staphylococcus lugdunensis, found in 14.9% of the animals, was firstly detected in reptiles. Vibrio sp. which is more associated with aquatic environments was found in 10.8% of the lizards in this study, with Vibrio cholerae being found in two of the animals. The prevalence of Salmonella sp. (5.4%) was low, compared with other studies carried out in reptiles. These results indicate that Anolis sp. in Tenerife could be playing a role in the maintenance and spread of the pathogens detected, being a possible risk factor for public health and biodiversity conservation.

New Data on Pterygodermatites (Pterygodermatites) plagiostoma Wedl, 1861 (Nematoda, Rictulariidae) Parasite of the Algerian Hedgehog Atelerix algirus Linnaeus, 1758 (Eulipotyphla: Erinaceidae) from the Canary Islands.

A redescription of the rictulariid nematode Pterygodermatites (Pterygodermatites) plagiostoma Wedl, 1861, is made by means of light and scanning electron microscopy, including the first data on male specimens. The morphologic study was based on specimens recovered from two Algerian hedgehogs (Atelerix algirus) from Tenerife and Gran Canaria islands (Canary Archipelago, Spain). The main characteristics of P. (P.) plagiostoma males are the presence of 49-53 pairs of cuticular projections, the presence of one or two midventral precloacal fans (generally one), and the size of two unequal spicules, measuring 98-123 µm (right spicule) and 185-236 µm (left spicule). The cloacal papillae are arranged according to the Ascaridida type. They include two precloacal pairs, an unpaired precloacal papilla, one pair lateral to the cloaca, six postcloacal pairs, and a pair of phasmids near the tail tip. Females are mainly characterized by the presence of 71-77 pairs of cuticular projections, with 43-46 pairs of prevulvar combs, by the differentiation from combs to spines at the level of or slightly posterior to the vulva and by the position of the vulva, located posteriorly to the esophagus-intestine junction. Clear differences were found between P. (P.) plagiostoma and related species of the subgenus P. (Pterygodermatites).

First Coronavirus Active Survey in Rodents From the Canary Islands.

Since the beginning of the 21st century five new coronaviruses inducing respiratory diseases in humans have been reported. These emergences has promoted research on coronaviruses in wildlife. We started the first eco-epidemiological study to screen the presence of coronaviruses circulating in mice and rats of four Canary Islands. Between 2015 and 2019, we obtained fecal samples of three rodent species (150 Mus musculus, 109 Rattus rattus and 1 Rattus norvegicus) captured in urban and rural areas. Fecal samples were analyzed by nRT-PCR and the resulting sequences were compared to known diversity using Bayesian phylogenetic methods. We only found coronavirus RNA in house mice from El Hierro (10.53%), Tenerife (7.02%) and Lanzarote (5.26%) islands. All coronaviruses detected belong to the species Murine coronavirus belonging to the genus Betacoronavirus and subgenus Embecovirus, being all positive house mice captured in anthropogenic environment. The phylogenetic analysis shows that murine coronaviruses from the Canary Islands are related to European murine coronaviruses. Albeit data are still scarce in the region, the most probable origin of M. coronavirus present in the Canary Islands is continental Europe. According to temporal Bayesian phylogenetics, the differentiation between Canary and continental viruses seems to be quite recent. Moreover, murine coronaviruses from El Hierro, Tenerife and Lanzarote islands tend to segregate in different clades. This enlightens the potential role of rodents or other possibly invasive species in disseminating infectious diseases to remote places through exchanges with the continent. It is important to consider these aspects in the sanitary control of islands, for health and biodiversity preservation concerns.

A Peculiar Distribution of the Emerging Nematode Angiostrongylus cantonensis in the Canary Islands (Spain): Recent Introduction or Isolation Effect?

Angiostrongylus cantonensis is an emerging zoonotic nematode recognized as the leading cause of eosinophilic meningitis in the word. After its discovery in China, it was recorded in 30 countries worldwide. Recently, it has expanded to new areas such as South America and it has been recently found in the Atlantic island of Tenerife (Canary Islands). In order to characterize the distribution of A. cantonensis in the Canary Islands, the lungs of 1462 rodents were sampled in eight islands of the archipelago over 13 years and were then analyzed for A. cantonensis. Remarkably, the parasite was detected only in Tenerife, in Rattus rattus (19.7%) and Rattus norvegicus (7.14%). They were concretely in the northern part of the island, which had a warmer and more humid climate than the south and main cities. The absence of this nematode in other islands with similar environmental conditions could be explained by an isolation effect or by a recent introduction of the parasite in the islands. Besides, the presence in Tenerife of the most invasive lineage of A. cantonensis reinforced the hypothesis of a recent introduction on this island. This study highlights the need to implement control measures to prevent the expansion to other areas in order to avoid the transmission to humans and other animals.

Bartonella species and haplotypes in rodents and their fleas in Lanzarote and El Hierro in the Canary Islands, Spain.

Because isolated ecosystems contribute to species variability, especially oceanic island ecosystems, the present work focused on the study of the Bartonella species and haplotypes in Lanzarote and El Hierro, two Canary islands with evident bioclimatic differences between them. A total of 123 rodents and 110 fleas from two islands were screened for the presence of Bartonella by PCR analysis of the gltA and nuoG genes. The overall prevalence was 5.7% in rodents and 20.4% in fleas. A total of seven gltA-haplotypes was found in both rodents and fleas, belonging to the species Bartonella mastomydis and Bartonella tribocorum in Lanzarote, and to Bartonella rochalimae and Bartonella elizabethae in El Hierro, as well as recently described species Bartonella kosoyi in both islands. Besides, potential co-infections were detected based on the nuoG analysis. Further, Xenopsylla cheopis was the only flea species identified. Our study shows that isolated ecosystems such as the Canary Islands lead to the appearance of new Bartonella haplotypes along different biotopes, with diverse flea species involved in the spreading of the pathogen being of great relevance due to the zoonotic potential of the species found.

Study of Zoonotic Enteric Pathogens of Atelerix algirus in Tenerife, Canary Islands, Spain.

Atelerix algirus is an invasive species in the Canary Islands (Spain). There are few studies about the zoonotic pathogens this species could be hosting; therefore, this study was focused on analyzing causative agents of diarrhea in humans in feces from hedgehogs. A total of 45 fecal samples obtained in Tenerife (Canary Islands) were analyzed in this study using Biofire FilmArray gastrointestinal panel with an integrated Biofire FilmArray system. Forty-two (93.33%) of the samples presented at least one of the pathogens detected by the panel. The prevalence of four bacteria stands out as for enteropathogenic Escherichia coli (71.11%), Salmonella (66.67%), Clostridioides difficile (33.33%), and Campylobacter sp. (22.22%), all of which were widely distributed along Tenerife. Besides, other pathogens were found, Cryptosporidium sp. and enterotoxigenic E. coli lt/st in 6.66% of the animals, Shigella/enteroinvasive E. coli in 4.44%, and Norovirus GI/GII, Plesiomonas shigelloides, and Vibrio sp. in 2.22%. Of the hedgehogs, 26.66% were hosting just one pathogen, and the others showed coinfection: 24.44% hosted two, 31.11% hosted three, and 11.11% hosted four or more. The close contact with hedgehogs may imply the transmission of not only one causative agent of diarrhea but also multiple agents, since coinfection is highly prevalent. The lack of management measurements for this animal in the Canary Islands, the common habit of adopting hedgehogs from wildlife without veterinary control, and the fact that most of the hedgehogs studied belonged to highly populated areas imply a high risk of transmission of pathogens to humans.

Pathogenic Leptospira species in rodents from Corsica (France).

Leptospirosis is a worldwide emerging zoonotic disease caused by Leptospira species, that in some patients develop severe forms with high mortality. In France, Corsica is the area where the highest incidences have been reported. The present study was focused on the analysis of pathogenic Leptospira species in rodents of Corsica, as these micromammals are the main natural reservoirs of the bacteria, in order to identify the circulating species and to locate possible risk focuses of transmission, as no previous study on the presence of Leptospira species has been carried out in the island. Rattus rattus, Rattus norvegicus, Apodemus sylvaticus and Mus musculus domesticus were captured in the proximity of water sources along Corsica, the detection of pathogenic Leptospira species was carried out by amplification of the LipL32 gene. The bacteria were found in all the rodent species analyzed and widely. The general prevalence was 10.4%, reaching the maximum value in Bastia (45%). Leptospira interrogans and Leptospira borgpetersenii were identified by phylogenetic analysis, but also two sequences which corresponded to an unnamed Leptospira species, only previously found in rodents of New Caledonia. The high incidence of human leptospirosis in Corsica could be partially explained by the wide distribution of pathogenic Leptospira species identified in this study. Also, the presence of an unknown pathogenic species of Leptospira in an area with high prevalence, may be involved in the higher incidence of Leptospirosis in this island, however, the zoonotic capacity of this species remains unknown. The results obtained are interesting for public health since all positive samples were found near water sources and one of the routes of transmission of leptospirosis is contact with contaminated water. This information could help the competent entities to take preventive measures, reducing the incidence of human leptospirosis in Corsica.

Rodents as Reservoirs of the Zoonotic Pathogens Coxiella burnetii and Toxoplasma gondii in Corsica (France).

Coxiella burnetii and Toxoplasma gondii are intracellular pathogens that cause important reproductive disorders in humans and animals worldwide, which translates into high economic losses. The aim of this study was to analyze the possible role of peridomestic small mammals in the maintenance and transmission of C. burnetii and T. gondii in the Mediterranean island of Corsica, by the analysis of the seroprevalence in rodents. Our results showed a wide distribution of C. burnetii and T. gondii in Corsica in Rattus rattus and Mus musculus domesticus. The overall seroprevalence of anti-C. burnetii was 9.32%, being 8.73% from R. rattus and 13.33% from M. m. domesticus. With respect to T. gondii, seropositive rodents were observed in 12.96% of total animals, being 12.76% from rats and 14.29% from mice. Taking into consideration the large range of host these pathogens can infect, including humans and livestock, and their pathologies, the results provided by this study are relevant for public health and veterinary, and could help the governmental entities to prevent possible infectious focuses of Q fever and toxoplasmosis.

First ocular report of Gurltia paralysans (Wolffhügel, 1933) in cat.

A two-year-old cat from Tenerife, Canary Islands (Spain), presented with two months history of uveitis in the left eye. It had been treated for two months but still showed an active uveitis. After new treatment, eye examination showed a mobile worm in the anterior chamber. Following surgical removal, the worm was obtained. Morphological study revealed that it was a male metastrongyloid nematode (Nematoda: Metastrongyloidea) with caudal bursa and two similar spicules. Molecular tools based on the 18Sr RNA gene sequence identified the parasite as Gurltia paralysans, which is a neurotropic nematode previously found in South America. Therefore, this article describes the first report of ophthalmic case of parasitism by G. paralysans, and cites for first time this species out of South America.