Helminths of South American fur seals (Arctocephalus australis) from the Subtropical Convergence Zone of the Southwestern Atlantic.

Parasites are important components of ecosystems and may contribute to the ecological aspects of their hosts and indicate the integrity of their environment. To identify the gastrointestinal helminths of the South American fur seal, Arctocephalus australis, 52 animals found dead on the Rio Grande do Sul coast, Southern Brazil, were necropsied. All studied animals were parasitized, and 104,670 specimens of helminths from three phyla and 14 taxa were collected. Adult specimens represented five of the identified species: Contracaecum ogmorhini, Adenocephalus pacificus, Stephanoprora uruguayense, Ascocotyle (Phagicola) longa, and Corynosoma australe; and one of the identified genera: Strongyloides sp. Immature forms represented the other eight taxa: Anisakidae gen. sp., Anisakis sp., Pseudoterranova sp., Contracaecum sp., Tetrabothriidae gen. sp., Cestoda gen. sp., Corynosoma cetaceum, and Bolbosoma turbinella. The acanthocephalan C. australe was the most prevalent and abundant parasite, whereas Strongyloides sp. had the highest intensity. This is the first record of the nematode Anisakis sp., digenean S. uruguayense, and acanthocephalan B. turbinella in this host. Trophic generalist species such as A. australis can be good indicators of the composition of the helminth fauna of their ecosystems, indicating the presence of zoonotic parasites transmitted by the consumption of fish.

Verminotic pneumonia in South American fur seal (Arctocephalus australis) in Southern Brazil.

Verminotic pneumonia caused by Parafilaroides spp. nematodes is an underreported disease in beached South American fur seals, with scant literature available on the characteristics of parafilaroidiasis, the nematode itself, as well as its occurrence in pinnipeds in Brazil. The present work aims to identify, describe and detail the histological features of the infection and molecular characteristics of verminotic pneumonia in the South American fur seal. Twenty-six specimens of Arctocephalus australis, found dead on the northern coast of Rio Grande do Sul in 2021, were analysed. These animals were identified and submitted to necropsy and histology. For the molecular identification of metastrongylids, lung fragments were subjected to DNA extraction, polymerase chain reaction targeting the Internal transcribed spacer 2 (ITS-2) gene and subsequent sequencing. In total, 12 animals presented with parasites in the lung parenchyma on histological evaluation, and only 1 showed a granulomatous lung lesion at necropsy. Microscopically, the nematodes were found mainly in the alveoli, associated with little or no inflammatory response, and they had morphological characteristics compatible with metastrongylids. Six ITS-2 gene quality sequences were obtained; after comparative analysis via BLAST, they showed similarity with sequences obtained from Parafilaroides sp. Therefore, verminotic pneumonia caused by Parafilaroides represents an important differential diagnosis of lung disease in South American fur seals found on the northern coast of Rio Grande do Sul.

New host and geographical records for Parafilaroides normani (Nematoda: Filaroididae) Dailey, 2009 in South American fur seal, Arctocephalus australis, from southern Brazil.

Lungworms are a common finding in seals and fur seals around the world. However, from existing records, the biogeographical distribution of filaroid helminths appears to be restricted, and these parasites are endemic in only certain areas and species, mainly in the Northern Hemisphere. The occurrence of infection in pinniped species in the Southern Hemisphere is scarce. The objective of this work is to verify the prevalence of lungworms in Arctocephalus australis in waters off the southern coast of Brazil. Twenty subadult specimens of A. australis found recently dead on the southern coast of Brazil were necropsied and their lungs were examined. Parasitic cysts were found in only one specimen (prevalence of 5%). The helminths were morphologically identified as Parafilaroides normani (Metastrongyloidea: Filaroididae). This helminth species has been reported in pinnipeds from Australia, New Zealand and South Africa. This is the first record of P. normani in A. australis and for the western South Atlantic, providing additional data regarding the biogeographic distribution of the parasite.

Fatal pulmonary parafilaroidiasis in a free-ranging subantarctic fur seal (Arctocephalus tropicalis) coinfected with two gammaherpesviruses and Sarcocystis sp.

A juvenile subantarctic fur seal (Arctocephalus tropicalis) found dead in Santa Catarina state, southern Brazil, presented with disseminated verminous pneumonia due to Parafilaroides sp. A concomitant infection with two different gammaherpesviruses was identified by PCR in different tissues; one of them possibly a novel species (tentatively named Otariid herpesvirus 7). Sarcocystis sp. DNA was identified molecularly in skeletal muscle samples with intrasarcoplasmic bradyzoites and no apparent tissue response. All analyzed samples (mandibular, laryngeal, tracheal, and mesenteric lymph nodes, and lung) were PCR-negative for Brucella spp. The most likely cause of death was severe pulmonary parafilaroidiasis. The pathogenic role of the gammaherpesviruses in several of the tissues was not evident. This study describes the pathogenicity of Parafilaroides sp. in a subantarctic fur seal, widens the host range of herpesvirus in pinnipeds, and reports the first molecular identification of Sarcocystis sp. in this species.

Fatal pulmonary parafilaroidiasis in a free-ranging subantarctic fur seal (Arctocephalus tropicalis) coinfected with two gammaherpesviruses and Sarcocystis sp / Parafilaroidose pulmonar fatal em um lobo-marinho-subantártico (Arctocephalus tropicalis) de vida livre coinfectado com dois gammaherpesvírus e Sarcocystis sp

Abstract A juvenile subantarctic fur seal (Arctocephalus tropicalis) found dead in Santa Catarina state, southern Brazil, presented with disseminated verminous pneumonia due to Parafilaroides sp. A concomitant infection with two different gammaherpesviruses was identified by PCR in different tissues; one of them possibly a novel species (tentatively named Otariid herpesvirus 7). Sarcocystis sp. DNA was identified molecularly in skeletal muscle samples with intrasarcoplasmic bradyzoites and no apparent tissue response. All analyzed samples (mandibular, laryngeal, tracheal, and mesenteric lymph nodes, and lung) were PCR-negative for Brucella spp. The most likely cause of death was severe pulmonary parafilaroidiasis. The pathogenic role of the gammaherpesviruses in several of the tissues was not evident. This study describes the pathogenicity of Parafilaroides sp. in a subantarctic fur seal, widens the host range of herpesvirus in pinnipeds, and reports the first molecular identification of Sarcocystis sp. in this species.

Resumo Um lobo-marinho-subantártico (Arctocephalus tropicalis) juvenil foi achado morto no Estado de Santa Catarina, sul do Brasil, apresentando pneumonia parasitária disseminada por Parafilaroides sp. Infecção concomitante por dois gammaherpesvírus diferentes foi identificada pela PCR em diversos tecidos, um desses herpesvírus possivelmente uma nova espécie (denominada provisoriamente Otariid herpesvirus 7). DNA de Sarcocystis sp. foi identificado molecularmente em amostras de músculo esquelético que apresentavam bradizoítos intra-sarcoplasmáticos sem aparente resposta tecidual. Todas as amostras analisadas (linfonodo mandibular, laríngeo, traqueal e mesentérico, e pulmão) pela PCR para Brucella spp. foram negativas. A causa mais provável da morte do animal foi parafilaroidose pulmonar severa. O papel patogénico dos gammaherpesvírus em vários tecidos não foi evidente. Este estudo descreve a patogenicidade de Parafilaroides sp. em um lobo-marinho-subantártico, amplia a variedade de hospedeiros de herpesvírus em pinípedes e reporta a primeira identificação molecular de Sarcocystis sp. para essa espécie.

Two Bacterial Genera, Sodalis and Rickettsia, Associated with the Seal Louse Proechinophthirus fluctus (Phthiraptera: Anoplura).

UNLABELLED: Roughly 10% to 15% of insect species host heritable symbiotic bacteria known as endosymbionts. The lice parasitizing mammals rely on endosymbionts to provide essential vitamins absent in their blood meals. Here, we describe two bacterial associates from a louse, Proechinophthirus fluctus, which is an obligate ectoparasite of a marine mammal. One of these is a heritable endosymbiont that is not closely related to endosymbionts of other mammalian lice. Rather, it is more closely related to endosymbionts of the genus Sodalis associated with spittlebugs and feather-chewing bird lice. Localization and vertical transmission of this endosymbiont are also more similar to those of bird lice than to those of other mammalian lice. The endosymbiont genome appears to be degrading in symbiosis; however, it is considerably larger than the genomes of other mammalian louse endosymbionts. These patterns suggest the possibility that this Sodalis endosymbiont might be recently acquired, replacing a now-extinct, ancient endosymbiont. From the same lice, we also identified an abundant bacterium belonging to the genus Rickettsia that is closely related to Rickettsia ricketsii, a human pathogen vectored by ticks. No obvious masses of the Rickettsia bacterium were observed in louse tissues, nor did we find any evidence of vertical transmission, so the nature of its association remains unclear. IMPORTANCE: Many insects are host to heritable symbiotic bacteria. These heritable bacteria have been identified from numerous species of parasitic lice. It appears that novel symbioses have formed between lice and bacteria many times, with new bacterial symbionts potentially replacing existing ones. However, little was known about the symbionts of lice parasitizing marine mammals. Here, we identified a heritable bacterial symbiont in lice parasitizing northern fur seals. This bacterial symbiont appears to have been recently acquired by the lice. The findings reported here provide insights into how new symbioses form and how this lifestyle is shaping the symbiont genome.

The utility of diversity profiling using Illumina 18S rRNA gene amplicon deep sequencing to detect and discriminate Toxoplasma gondii among the cyst-forming coccidia.

Next-generation sequencing (NGS) has the capacity to screen a single DNA sample and detect pathogen DNA from thousands of host DNA sequence reads, making it a versatile and informative tool for investigation of pathogens in diseased animals. The technique is effective and labor saving in the initial identification of pathogens, and will complement conventional diagnostic tests to associate the candidate pathogen with a disease process. In this report, we investigated the utility of the diversity profiling NGS approach using Illumina small subunit ribosomal RNA (18S rRNA) gene amplicon deep sequencing to detect Toxoplasma gondii in previously confirmed cases of toxoplasmosis. We then tested the diagnostic approach with species-specific PCR genotyping, histopathology and immunohistochemistry of toxoplasmosis in a Risso's dolphin (Grampus griseus) to systematically characterise the disease and associate causality. We show that the Euk7A/Euk570R primer set targeting the V1-V3 hypervariable region of the 18S rRNA gene can be used as a species-specific assay for cyst-forming coccidia and discriminate T. gondii. Overall, the approach is cost-effective and improves diagnostic decision support by narrowing the differential diagnosis list with more certainty than was previously possible. Furthermore, it supplements the limitations of cryptic protozoan morphology and surpasses the need for species-specific PCR primer combinations.

High morphological plasticity and global geographical distribution of the Pacific broad tapeworm Adenocephalus pacificus (syn. Diphyllobothrium pacificum): molecular and morphological survey.

The most important causative agent of human diphyllobothriosis in South America, Diphyllobothrium pacificum, is transferred to the original genus Adenocephalus Nybelin, 1931; revised and redescribed on the basis of the evaluation of an extensive material collected mainly from northern fur seal, Callorhinus ursinus, from St. Paul Island, Alaska. Detailed analysis of morphological and morphometrical data shows a high variability in most of the characteristics traditionally used in diagnosis of diphyllobothriid tapeworms. Phylogenetic analyses based on newly characterised sequences of mitochondrial cytochrome c oxidase subunit 1 and nuclear large subunit ribosomal RNA genes consistently reveal Adenocephalus pacificus as a sister lineage to the clade formed of the remaining Diphyllobothrium species and other genera (Digramma, Diplogonoporus, Ligula). Despite the generally similar morphology, A. pacificus can be differentiated from the closely related taxa in the presence of transverse papilla-like tegumental protuberances distributed anteriorly, separated by narrow semicircular grooves on the ventral surface of proglottids between their anterior margin and the anterior edge of the male gonopore, and relatively small eggs. A. pacificus displays a relatively low host specificity (found in 9 of 16 otariids, and in accidental hosts such as man, dog and jackal, the latter representing a new host) and a uniquely wide geographical distribution on both hemispheres. In addition, suitability of morphological criteria used in diagnostics of diphyllobothriid cestodes is discussed.

Anisakids (Nematoda: Anisakidae) from stomachs of northern fur seals (Callorhinus ursinus) on St. Paul Island, Alaska: parasitological and pathological analysis.

Anisakid nematodes parasitize the alimentary tracts of aquatic vertebrates, including northern fur seals (Callorhinus ursinus) (NFS). The main purpose of this study was to estimate the prevalence, intensity, and species composition of anisakids in NFSs on St. Paul Island, Alaska, and to analyze changes in NFS infection with these nematodes during the last decades. The study was carried out on St. Paul Island, Alaska, in July-August 2011-2013. Stomachs of 443 humanely harvested NFS males were collected from five separate haul-out areas. A total of 4,460 anisakid specimens were collected and identified by morphological criteria. Gross and histological examinations of stomachs were also performed. The overall prevalence of NFS infection was 91.2%; overall mean intensity 10.9, and overall median intensity 6. Five species, Anisakis simplex s. l., Contracaecum osculatum s. l., Pseudoterranova decipiens s. l., P. azarazi and Phocascaris cystophorae, were found. The prevalence of Anisakis was 46.5%, its intensity 7.5. The prevalence and intensity of Contracaecum, Pseudoterranova, and Phocascaris were 33.6% and 3.5, 81.9% and 6.5, and 5.2% and 1.7, respectively. Significant differences in the prevalence of NFS infection were observed between separate haul-outs. Comparison of the present data with the results of studies performed in the North Pacific in the 1960s, showed a significant decrease in the intensity of NFS infection with anisakids during the last decades. The prevalence of mucosal lesions in NFS stomachs caused by anisakids also decreased from 92 to 20.9%. Possible reasons for the changes in NFS infection with anisakids are discussed.

Multisystemic toxoplasmosis associated with a type II-like Toxoplasma gondii strain in a New Zealand fur seal (Arctocephalus forsteri) from New South Wales, Australia.

We report the first confirmed case of toxoplasmosis in an Australian pinniped. Presence of Toxoplasma gondii DNA was detected in the brain of a free-ranging subadult New Zealand fur seal (Arctocephalus forsteri) with nonsuppurative meningoencephalitis, hypophysitis, posterior uveitis, retrobulbar cellulitis, and myocarditis associated with protozoan cysts and tachyzoites. The emaciated seal stranded moribund on a beach in northern Sydney in New South Wales. Histopathology coupled with specific immunohistochemistry and PCR assays confirmed the presence of T. gondii. The T. gondii sample (NZfs8825) identified in this study has an identical genotype as the type II (ToxoDB PCR-RFLP genotype #1) based on the direct sequencing and virtual RFLP of multilocus DNA markers including SAG1, 5'- and 3'-SAG2, alt.SAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico. Direct sequencing of T. gondii B1 DNA marker from the T. gondii sample (NZfs8825) identified a type II-like strain, based on presence of non-archetypal B1 gene polymorphisms previously reported as unique to Australia. This study suggests that T. gondii oocysts originating from mainland Australia, which has a large population of feral cats, may act as a disease threat to native marine fauna. Therefore, emerging toxoplasmosis in the Arctic has a relevant parallel in the Southern Ocean within Australian waters with yet unknown relevance to Antarctica.

Serum chemistry and antibodies against pathogens in antarctic fur seals, Weddell seals, crabeater seals, and Ross seals.

Information on health parameters, such as antibody prevalences and serum chemistry that can reveal exposure to pathogens, disease, and abnormal physiologic conditions, is scarce for Antarctic seal species. Serum samples from Antarctic fur seals (Arctocephalus gazella, n=88) from Bouvetøya (2000-2001 and 2001-2002), and from Weddell seals (Leptonychotes weddellii, n=20), Ross seals (Ommatophoca rossii, n=20), and crabeater seals (Lobodon carcinophagus, n=9) from the pack-ice off Queen Maud Land, Antarctica (2001) were analyzed for enzyme activity, and concentrations of protein, metabolites, minerals, and cortisol. Adult Antarctic fur seal males had elevated levels of total protein (range 64-99 g/l) compared to adult females and pups (range 52-79 g/l). Antarctic fur seals had higher enzyme activities of creatine kinase, lactate dehydrogenase, and amylase, compared to Weddell, Ross, and crabeater seals. Antibodies against Brucella spp. were detected in Weddell seals (37%), Ross seals (5%), and crabeater seals (11%), but not in Antarctic fur seals. Antibodies against phocine herpesvirus 1 were detected in all species examined (Antarctic fur seals, 58%; Weddell seals, 100%; Ross seals, 15%; and crabeater seals, 44%). No antibodies against Trichinella spp., Toxoplasma, or phocine distemper virus (PDV) were detected (Antarctic fur seals were not tested for PDV antibodies). Antarctic seals are challenged by reduced sea ice and increasing temperatures due to climate change, and increased anthropogenic activity can introduce new pathogens to these vulnerable ecosystems and represent a threat for these animals. Our data provide a baseline for future monitoring of health parameters of these Antarctic seal species, for tracking the impact of environmental, climatic, and anthropogenic changes in Antarctica over time.

Detection of Toxoplasma gondii antibodies in Antarctic pinnipeds.

The presence of Toxoplasma gondii antibodies was investigated in Antarctic marine mammals. Two hundred and eleven sera from different species of pinnipeds collected in years 2007, 2010 and 2011 from different locations in the South Shetland Islands and Antarctic Peninsula were analysed using a commercially available agglutination test kit. The presence of antibodies (titres ≥ 1:25) against T. gondii was detected in a total of 28 animals (13.3%). Amongst animal species, percentages of detection were higher in Southern elephant seals (Mirounga leonina) (76.9%; 10/13) followed by Weddell seals (Leptonychotes weddellii) (41.9%; 13/31). Antibodies were also found in 4 of 165 (2.4%) Antarctic fur seals (Arctocephalus gazella) and 1 of 2 Crabeater seals (Lobodon carcinophaga). Highest titres (1:100-1:800) were also observed in Southern elephant seals and Weddell seals. To the best of our knowledge this is the first report on the detection of antibodies against T. gondii in Antarctic marine mammals.

Corynosoma australe Johnston, 1937 and C. cetaceum Johnston & Best, 1942 (Acanthocephala: Polymorphidae) from marine mammals and fishes in Argentinian waters: allozyme markers and taxonomic status.

Genetic and morphological studies were carried out on acanthocephalans belonging to Corynosoma Lühe, 1904 and referable to the species C. cetaceum Johnston & Best, 1942 and C. australe Johnston, 1937, which were recovered from both definitive and intermediate hosts in Argentinian waters. The aims were to estimate the level of genetic differentiation between the two taxa at any stage of their life-cycle, to provide genetic (allozyme) markers for their recognition and to analyse the systematic status of both taxa. Acanthocephalans were collected from the stomach and intestine of Arctocephalus australis (Zimmerman), the intestine of Mirounga leonina (Linnaeus) and the stomach of Pontoporia blainvillei Gervais & D'Orbigny (definitive hosts) in Argentinian waters. Alternative alleles at all the 13 enzymatic loci studied were observed for C. australe and C. cetaceum. The specimens from the stomach of both P. blainvillei and A. australis were identified, on the basis of the great number of diagnostic loci found, as C. cetaceum; those from intestine of both A. australis and M. leonina as C. australe. A high level of genetic differentiation (D(Nei)=infinity: I(Nei)=0.00) between the two taxa was found, suggesting a generic distinction between the two species. Cystacanths of the two species from the body-cavity of the fish Cynoscion guatucupa (Cuvier) collected from the same geographical area were identified genetically. Morphological patterns, such as the number of hooks and hook rows on the proboscis, the distribution of somatic and genital armature, and other morphometric and meristic differences, in addition to ecological data, enabled the identification of these two species at cystacanth, juvenile and adult stages. However, a number of morphological and morphometric features of the Argentinian material were different to those of C. australe and C. cetaceum described from other regions of the world.