Verminous bronchitis and pneumonia by nasal trematodes in Greater Caribbean manatees from Puerto Rico.

Five adult Greater Caribbean manatees Trichechus manatus manatus were found stranded on various coasts of Puerto Rico; 2 stranded alive and 3 stranded dead. Clinical signs observed in live-stranded manatees included emaciation, weakness, bradypnea, arrhythmia, and nasal mucus discharge. Postmortem examinations revealed serosanguinous, mucohemorrhagic, or suppurative exudate in bronchi associated with luminal adult Pulmonicola cochleotrema (range: 18-182 trematodes), accompanied by pulmonary abscesses in 2 cases. Histologically, we observed eosinophilic bronchopneumonia of varying severity (n = 4) and chronic erosive to eosinophilic tracheobronchitis (n = 4) with squamous metaplasia (n = 3) and intralesional trematodes and eggs. The trematode identity was confirmed and compared through molecular analysis for the amplified 18S rDNA fragment. Comorbidities included enteric chiorchosis (n = 5), gastric heterocheilosis (n = 4), malnutrition (n = 4), trauma related to watercraft collision (n = 3), systemic toxoplasmosis (n = 1), acute bacterial peritonitis (n = 1), and interstitial nephritis (n = 1), suggesting that immunosuppression was a predisposing factor for lower respiratory tract pulmonicolosis. Based on lesion severity, clinical signs, and the presence and absence of other findings to explain death, this condition was considered the primary cause of death in 1 manatee, a contributory cause of death in 3 manatees, and an incidental finding in 1 individual. These clinicopathological descriptions will facilitate the diagnosis and clinical management of pulmonicolosis in T. manatus, a species endangered with extinction.

A combined diagnostic approach for the investigation of lesions resembling aspergillosis in Caribbean sea fans (Gorgonia spp.).

Aspergillosis of gorgonian sea fans is a Caribbean-wide disease characterized by focal, annular purple pigmentation with central tissue loss. We applied a holistic diagnostic approach including histopathology and a combination of culture and direct molecular identification of fungi to evaluate these lesions with the goal of determining the diversity of associated micro-organisms and pathology. Biopsies were collected from 14 sea fans without gross lesions and 44 sea fans with lesions grossly consistent with aspergillosis in shallow fringing reefs of St. Kitts. Histologically, the tissue loss margin had exposure of the axis and amoebocyte encapsulation with abundant mixed micro-organisms. Polyp loss, gastrodermal necrosis, and coenenchymal amoebocytosis were at the lesion interface (purpled area transitioning to grossly normal tissue) with algae (n = 21), fungus-like hyphae (n = 20), ciliate protists (n = 16), cyanobacteria (n = 15), labyrinthulomycetes (n = 5), or no micro-organisms (n = 8). Slender, septate hyaline hyphae predominated over other morphological categories, but were confined to the axis with little host response other than periaxial melanization. Hyphae were absent in 6 lesioned sea fans and present in 5 control biopsies, questioning their pathogenicity and necessary role in lesion causation. From cultivation, different fungi were isolated and identified by sequencing of the nuclear ribosomal internal transcribed spacer region. In addition, 2 primer pairs were used in a nested format to increase the sensitivity for direct amplification and identification of fungi from lesions, thereby circumventing cultivation. Results suggest mixed and opportunistic infections in sea fans with these lesions, requiring longitudinal or experimental studies to better determine the pathogenesis.

First molecular identification of the trematode Pulmonicola cochleotrema (Platyhelminthes: Opisthotrematidae) in West Indian manatees (Trichechus manatus, Sirenia: Trichechidae) from Puerto Rico and Florida.

West Indian manatees (Trichechus manatus) harbor a variety of endoparasites, including the nasal trematode Pulmonicola cochleotrema, which infects the respiratory tract, especially the nasal passages. Previous studies have described and identified this digenean using morphological data only. This study presents the first molecular identification of P. cochleotrema in West Indian manatees from Puerto Rico and Florida. Samples of the trematode were collected from seven manatees found stranded dead at both locations. The small subunit ribosomal DNA (18S rDNA) was amplified from each sample using universal primers for different regions of the gene, resulting in a consensus sequence of 1871 base pairs. The phylogenetic reconstruction was carried out using DNA sequences of other species of digenean parasites from other hosts, including a trematode of the same taxonomic family from another sirenian species. Specimens collected from both locations show the same molecular identity using SSU rDNA sequence data. The identity of P. cochleotrema was confirmed using the Basic Local Alignment Search Tool from the National Center for Biotechnology Information database, yielding a high similarity of 98.8 % with Opisthotrema dujonis and 98.2 % with Lankatrema mannarense located in the same clade in our analysis. The latter two digeneans belong to the Opisthotrematidae as does P. cochleotrema and previous studies reported them infecting the Eustachian tubes, esophagus, and digestive tract in dugongs (Dugong dugon). These findings evidence that the nasal trematode of manatees in Florida and Puerto Rico and the dugong, all inhabiting and feeding in marine environments, will have a marine mollusk as an intermediate host, probably a gastropod. The question remains, which species of nasal trematode are harbored by lotic-dwelling manatees in other parts of their distribution like South America.

A case report and statewide surveillance of "weak meat" condition of Alaska weathervane scallops, Patinopecten caurinus, linked to a recently identified pathogenic parasite, Merocystis kathae (Apicomplexa: Aggregatidae).

Weathervane scallop, Patinopecten caurinus, the largest scallop species in the world, is distributed from northern California, U.S.A., to the Bering Sea, and is only commercially harvested in Alaska. The fishery is considered well managed by the State of Alaska (U.S.A) Department of Fish and Game (ADF&G) and federal government, with many precautionary measures in place to avoid overharvest. There have been episodic declines in some management areas due to unknown causes. Fishermen also encounter scallops with abnormal adductor muscles, a condition colloquially termed "weak meat", characterized by the retention of muscle when shucked, an obvious darkened discoloration, and/or an abnormal texture making the product unacceptable for marketing. A similar syndrome in Atlantic sea scallops, Placopecten magellanicus, described as "gray meat", occurs in the eastern U.S. and Canada, and proposed causes include senescence, loss of bioenergetics due to chronic infestations, or a synergism of these factors. Recently a severe apicomplexan infection was found to cause a gray meat condition in Iceland scallops, Chlamys islandica, and the collapse of that stock. This parasite was subsequently detected in Atlantic sea scallops with the gray meat condition off the U.S. East Coast. Studies that followed identified the parasite as Merocystis kathae, previously described from the common whelk, Buccinum undatum, more than 100 years ago. In 2015 Bering Sea fishermen reported weak meat in their catch, so samples were submitted to ADF&G for diagnosis. Adductor muscles from all affected scallops had many large foci of an apicomplexan associated with necrosis, fibrosis, and muscular atrophy. Given the reduced quality, marketability, and possibly fitness of affected scallops, we performed a survey to estimate prevalence, intensity, and geographic distribution of this apicomplexan in Alaskan weathervane scallops. We sampled 180 scallops, from individual beds within each of the three major geographically broad scallop areas in Alaska. Overall prevalence was about 82%, ranging from 69 to 100% by district. Overall mean infection intensity, based on the number of parasite foci/section, was about 9 (range of 5-29, by location), with scallops from the Bering Sea and Southwest Kodiak being most severely infected. Molecular analyses confirmed that the Alaskan parasite is M. kathae, i.e., the same apicomplexan that caused the collapse of Icelandic scallops and a suspected cause for gray meat and mass mortality of Atlantic sea scallops in northeast North America.

Gross and microscopic pathology of West Indian sea eggs (Tripneustes ventricosus).

In this study, we performed comprehensive pathology examinations on 83 Tripneustes ventricosus from 11 locations on St. Kitts to build baseline data necessary for disease diagnosis in this species. Gross abnormalities were observed in 23/83 (28%) urchins and included spine loss, visceral hyperpigmentation, test discoloration, and test ulceration. Ciliates were the only protists identified in this study via examination of tissue wet mounts and histology, documented in 50/83 (60%) urchins. Microscopic observations associated with visibly abnormal status included muscle necrosis, test and appendage inflammation, appendage (tube feet, spines, and pedicellariae) degeneration, severe coelomocytosis, and generalized hypermelanosis. Enterocyte intranuclear inclusion bodies, microbial aggregates, nerve pigmentation, enteric pigmentation, integument-associated crustaceans, and encysted metazoan parasites were of uncertain pathological significance. The etiology for any lesion was not microscopically apparent, contrasting literature implicating common marine bacteria in urchin diseases. This study highlights the importance of histopathology in urchin disease investigations and facilitates the recognition of disease in T. ventricosus.

A novel dimorphic microsporidian Ameson iseebi sp. nov. infecting muscle of the Japanese spiny lobster, Panulirus japonicus, in western Japan.

Japanese spiny lobsters (Panulirus japonicus) exhibiting white opaque abdominal muscle were found in Mie and Wakayama prefectures, in mid-Western Japan. Microscopically, two types of microsporidian spores, ovoid and rod-shaped, were observed infecting the muscle. Histologically, both types of spore were detected inside myofibers of the abdomen, appendages, and cardiac muscles and were often both observed in a single myofiber simultaneously. Transmission electron microscopy revealed that ovoid spores have villous projections on the surface, and that ovoid and rod-shaped spores have a polar filament with 12 coils and 6 to 8 coils respectively. Merogonic and sporogonic stages were observed around ovoid spores, but rarely around rod-shaped spores. The small subunit ribosomal DNA sequences obtained from both spore types were identical to each other, indicating that this microsporidian exhibits a clear spore dimorphism. Phylogenetic analysis based on the rDNA sequences indicates that this microsporidian is part of a clade consisting of the genera Ameson and Nadelspora, with the most closely related species being A. herrnkindi found in the Caribbean spiny lobster P. argus. Based on ultrastructural features, molecular phylogenetic data, host type and geographical differences among known species in these genera, the species found in whitened abdominal muscles of the Japanese spiny lobster is described as Ameson iseebi sp. nov.

Pathology of multifocal purple spots, a nonspecific lesion morphology of Caribbean sea fans Gorgonia spp.

Disease is contributing to the decline of coral reefs globally, but the cause and pathogenesis of most coral diseases are poorly understood. Using Gorgonia ventalina and G. flabellum as a model for coral disease diagnosis, we histologically and microbiologically examined 45 biopsies of lesions resembling Gorgonia multifocal purple spots (MFPS) with the aim of forming a comprehensive case definition based on gross and microscopic morphologic descriptions and associated etiologies. Macroscopically, all lesions were small circular areas of purple pigmentation. Gross morphologies included pigmentation only (4/45, 9%), or pigmentation with branchlet expansion and fusion (19/45, 22%), sessile masses (17/45, 38%), or hard nodules (5/45, 9%). Histological morphologic diagnoses included amoebocyte encapsulation (9/45, 20%), coenenchymal amoebocytosis (6/45, 13%), melanin (17/45, 38%), and gorgonin deposition (13/45, 29%). Sixty-four percent of instances of fungi and 86% of labyrinthulomycetes were localized to grossly normal portions of the biopsy, whereas barnacles were only within lesions, and 87% of instances of algae and 82% of cyanobacteria were within lesioned area of the biopsy. Penicillium (n = 12) was the predominant genus of fungi isolated from biopsies. Barnacles were identified as Conopea sp. using molecular techniques. The pathology and etiology underlying MFPS lesions are diverse, consistent with a highly nonspecific lesion pattern rather than a specific disease. This study demonstrates the importance of microscopic examination of tissues for accurate classification of coral diseases and lesion patterns.

Extraintestinal Acanthocephalan Oncicola venezuelensis (Oligacanthorhynchidae) in Small Indian Mongooses (Herpestes auropunctatus) and African Green Monkeys (Chlorocebus aethiops sabaeus).

We identified multiple extraintestinal cystacanths during routine postmortem examination of 3 small Indian mongooses and 2 African green monkeys from the Caribbean island of St. Kitts. In mongooses, cystacanths were encysted or free in the subcutaneous tissue, skeletal muscle, or peritoneal or pericardial cavities, whereas in the monkeys, they were in the cavity and parietal layer of the, tunica vaginalis, skeletal muscle, and peritoneal cavity. Morphological, histological, and molecular characterization identified these cystacanths as Oncicola venezuelensis (Acanthocephala: Oligacanthorhynchidae). There was minimal to mild lymphoplasmacytic inflammation associated with the parasite in the mongooses and moderate inflammation, mineralization, hemorrhage, and fibrosis in the connective tissue between the testis and epididymis in 1 monkey. We identified a mature male O. venezuelensis attached in the aboral jejunum of a feral cat, confirming it as the definitive host. Termites serve as intermediate hosts and lizards as paratenic hosts. This report emphasizes the role of the small Indian mongoose and African green monkey as paratenic hosts for O. venezuelensis.

Scaphanocephalus-associated dermatitis as the basis for black spot disease in Acanthuridae of St. Kitts, West Indies.

Acanthurus spp. of St. Kitts and other Caribbean islands, including ocean surgeonfish A. bahianus, doctorfish A. chirurgus, and blue tang A. coeruleus, frequently show multifocal cutaneous pigmentation. Initial reports from the Leeward Antilles raised suspicion of a parasitic etiology. The aim of this study was to quantify the prevalence of the disease in St. Kitts' Acanthuridae and describe its pathology and etiology. Visual surveys demonstrated consistently high adjusted mean prevalence at 3 shallow reefs in St. Kitts in 2017 (38.9%, 95% CI: 33.8-43.9) and 2018 (51.5%; 95% CI: 46.2-56.9). There were no differences in prevalence across species or reefs, but juvenile fish were less commonly affected than adults. A total of 29 dermatopathy-affected acanthurids were sampled by spearfishing for comprehensive postmortem examination. Digenean metacercariae were dissected from <1 mm cysts within pigmented lesions. Using partial 28S rDNA sequence data they were classified as Family Heterophyidae, members of which are commonly implicated in black spot disease of other fishes. Morphological features of the parasite were most typical of Scaphanocephalus spp. (Creplin, 1842), and 2 genetic profiles were obtained suggesting more than 1 digenean species. Histologically, pigmented lesions had mild chronic perivascular dermatitis and increased melanophores and melanin density, often centered on encysted digenean metacercariae. In 1 affected A. chirurgus, similar metacercariae were histologically identified in skeletal and cardiac muscle. Further research is needed to clarify impact on host fitness, establish the number of heterophyid digenean species that cause black spots on Caribbean fishes and to determine the intermediate and definitive host species.

Pathology and reproductive health of queen conch (Lobatus gigas) in St. Kitts.

Queen conch, Lobatus gigas, are one of the largest commercial fisheries in the Caribbean and are severely depleted due to overharvesting. Dwindling and fragmented populations are at high risk to stochastic events such as disease epidemics; however, there is a paucity of literature regarding queen conch disease. A histopathological survey was conducted to assess the disease status of St. Kitts' queen conch populations, and contribute to the little information known about L. gigas pathology. Using a standard dissection and sampling protocol, pathology status was assessed in 61 L. gigas sampled opportunistically from fishermen in St. Kitts from October 2015 to November 2016. Pathology was observed in 13.1% (8/61) of the study population, mostly comprising internal parasitism (n = 5), especially encysted metazoan parasites (likely digenean metacercaria), observed in the gill, mantle, digestive gland and large intestine. Parasitism appeared incidental, having little impact on the host's health and minimal host response to the infection. Additionally, aspects of L. gigas reproductive health were analyzed: reproductive season in St. Kitts was confirmed from May to September; lip thickness is suggested as a better regulation measure for protecting juvenile populations compared to shell length; and variation in gonad color is found to be an unreliable predictor of reproductive activity. We conclude that disease appears to be uncommon in fisherman-caught L. gigas from St. Kitts. Pathologies observed elsewhere, including imposex and apicomplexan infection of the digestive gland, could not be substantiated in the study population. The cryptic nature of sick gastropods, as well as high predation rate and selection pressure for diseased queen conch may yield underrepresentation of pathology in such surveys. Future disease surveys could benefit from targeting stunted individuals, those showing unusual behavior, or those grown in an aquaculture environment.

Digestive gland inclusion bodies in queen conch (Lobatus gigas) are non-parasitic.

Unusual inclusion bodies occur within the epithelial cells of the digestive gland of queen conch, Lobatus gigas, and have previously been described as apicomplexan parasites. The aim of this study was to investigate the parasitic features of these inclusion bodies in queen conch. L. gigas from St. Kitts (Caribbean Sea) consistently (100% of n = 61) showed large numbers of ovoid to tri-bulbous dark brown inclusion bodies (15 × 30 µm) within vacuolar cells. Histochemical stains demonstrated iron, melanin, and glycoprotein and/or mucopolysaccharide within the inclusion bodies. Microscopic features indicative of a host response to injury were lacking in every case, as were consistent morphological forms to indicate distinct parasitic stages. Transmission electron microscopy failed to reveal cellular organelles of parasitic organisms and DNA extractions of purified inclusion bodies did not yield sufficient concentrations for successful PCR amplification. Scanning electron microscopy with energy dispersive X-ray analysis revealed a number of elements, particularly iron, within the inclusion bodies. We conclude that the inclusion bodies are not an infectious agent, and hypothesize that they represent a storage form for iron, and potentially other elements, within a protein matrix. Similar structures have been described in the digestive glands of other invertebrates, including prosobranchs.

A Recent Whole-Genome Duplication Divides Populations of a Globally Distributed Microsporidian.

The Microsporidia are a major group of intracellular fungi and important parasites of animals including insects, fish, and immunocompromised humans. Microsporidian genomes have undergone extreme reductive evolution but there are major differences in genome size and structure within the group: some are prokaryote-like in size and organisation (<3 Mb of gene-dense sequence) while others have more typically eukaryotic genome architectures. To gain fine-scale, population-level insight into the evolutionary dynamics of these tiny eukaryotic genomes, we performed the broadest microsporidian population genomic study to date, sequencing geographically isolated strains of Spraguea, a marine microsporidian infecting goosefish worldwide. Our analysis revealed that population structure across the Atlantic Ocean is associated with a conserved difference in ploidy, with American and Canadian isolates sharing an ancestral whole genome duplication that was followed by widespread pseudogenisation and sorting-out of paralogue pairs. While past analyses have suggested de novo gene formation of microsporidian-specific genes, we found evidence for the origin of new genes from noncoding sequence since the divergence of these populations. Some of these genes experience selective constraint, suggesting the evolution of new functions and local host adaptation. Combining our data with published microsporidian genomes, we show that nucleotide composition across the phylum is shaped by a mutational bias favoring A and T nucleotides, which is opposed by an evolutionary force favoring an increase in genomic GC content. This study reveals ongoing dramatic reorganization of genome structure and the evolution of new gene functions in modern microsporidians despite extensive genomic streamlining in their common ancestor.

Gray meat in the Atlantic sea scallop, Placopecten magellanicus, and the identification of a known pathogenic scallop apicomplexan.

Atlantic sea scallop (Placopecten magellanicus) meats are normally firm and creamy white. However, scallops with small, darkened and stringy adductor muscle (gray meat) episodically occur along the Eastern Seaboard, most recently in the rotational management areas of Georges Bank after extended fishing closures. These gray meat scallops are associated with reduced harvestable biomass and mass mortality events. We tested age, nutritional stress and disease as causative agents for this condition. Adult scallops of different shell heights (SH) ranging from (90-145mm) were collected from Georges Bank and analyzed for meat quality and the presence of pathogens using biochemical, histopathological and molecular methods. Gray meat occurrence was weakly correlated with shell height only explaining 8.49% of the variance in a generalized additive model (GAMS). Gray meat weights were lower than white meat (p<0.001) and there was a dramatic reduction in protein content (p<0.05) in gray meat scallops associated with extensive myodegeneration. Amino acid profiles confirmed the breakdown of muscle tissue with an increase in free hydroxyproline in gray meat scallops. Infection by an apicomplexan parasite was detected in the muscle tissue of all gray meat scallops tested. An intermediate pathology stage (brown meat) was also identified. As the parasitic infection increased, meat quality decreased. Numerous developmental stages of the parasite were present in various organs of the scallops. This apicomplexan has an identical SSU rDNA sequence to a novel parasite occurring in the Iceland scallop during a recent mass mortality event. The range of this parasite in Atlantic sea scallops and the effect of abiotic/biotic stressors on pathogenicity are currently unknown. Results from this study link an apicomplexan species, known to be highly pathogenic in scallops, to gray meat occurrence with a potentially high impact on the fishery.

Morphological, ultrastructural, and molecular description of Unicapsula fatimae n. sp. (Myxosporea: Trilosporidae) of whitespotted rabbitfish (Siganus canaliculatus) in Omani waters.

Investigations regarding the parasite fauna of wild whitespotted rabbitfish (Siganus canaliculatus) Park, 1797 revealed white, spherical, loosely attached cysts measuring 896 (375-1406) µm in diameter in the inner endothelial wall of the esophagus and stomach. Mature spores inside these cysts corresponded to the original description of spores belonging to the genus Unicapsula Davis, 1924. Unicapsula fatimae n. sp. spores were 6.23 (5.60-6.60) µm in length and 6.80 (6.12-7.39) µm in width. The length of large polar capsule was 2.62 (2.18-2.97) µm and width was 2.65 (2.32-2.90) µm, and the extended large polar capsule filament length was 15.50 (11.71-19.99) µm. Transmission electron microscope images of the plasmodia revealed a complex cyst structure that was unique among other Unicapsula spp. Ultrastructural details of the host-parasite interface and developmental stages of a species from the Unicapsula genus are described for the first time. Histology of an infected esophagus revealed some abnormalities and changes in the host tissue around the infection site, including hypertrophy of host esophagus epithelial cells and hyperplasia of host glandular tubules. The parasite presented here has been added to the genus Unicapsula using comparative morphological analysis and ultrastructural investigations supported by 18S small subunit ribosomal DNA molecular analysis.

Evolutionary origin of Ceratonova shasta and phylogeny of the marine myxosporean lineage.

In order to clarify the phylogenetic relationships among the main marine myxosporean clades including newly established Ceratonova clade and scrutinizing their evolutionary origins, we performed large-scale phylogenetic analysis of all myxosporean species from the marine myxosporean lineage based on three gene analyses and statistical topology tests. Furthermore, we obtained new molecular data for Ceratonova shasta, C. gasterostea, eight Ceratomyxa species and one Myxodavisia species. We described five new species: Ceratomyxa ayami n. sp., C. leatherjacketi n. sp., C. synaphobranchi n. sp., C. verudaensis n. sp. and Myxodavisia bulani n. sp.; two of these formed a new, basal Ceratomyxa subclade. We identified that the Ceratomyxa clade is basal to all other marine myxosporean lineages, and Kudoa with Enteromyxum are the most recently branching clades. Topologies were least stable at the nodes connecting the marine urinary clade, the marine gall bladder clade and the Ceratonova clade. Bayesian inference analysis of SSU rDNA and the statistical tree topology tests suggested that Ceratonova is closely related to the Enteromyxum and Kudoa clades, which represent a large group of histozoic species. A close relationship between Ceratomyxa and Ceratonova was not supported, despite their similar myxospore morphologies. Overall, the site of sporulation in the vertebrate host is a more accurate predictor of phylogenetic relationships than the morphology of the myxospore.

Zoothamnium duplicatum infestation of cultured horseshoe crabs (Limulus polyphemus).

An outbreak of the sessile peritrich Zoothamnium duplicatum in a pilot, commercial-scale Limulus polyphemus hatchery resulted in the loss of ∼96% (40,000) second/third instar larvae over a 61day period. peritrich growth was heavy, leading to mechanical obstruction of the gills and physical damage. The peritrichs were controlled without resultant loss of juvenile crabs by administering 10ppm chlorine in freshwater for 1h and the addition of aquarium grade sand; a medium into which the crabs could burrow and facilitate cleaning of the carapace. Peritrich identity was confirmed from a partial SSU rDNA contiguous sequence of 1343bp (99.7% similarity to Z. duplicatum).

Occurrence and molecular identification of Anisakis Dujardin, 1845 from marine fish in southern Makassar Strait, Indonesia.

Anisakis spp. (Nematoda: Anisakidae) parasitize a wide range of marine animals, mammals serving as the definitive host and different fish species as intermediate or paratenic hosts. In this study, 18 fish species were investigated for Anisakis infection. Katsuwonus pelamis, Euthynnus affinis, Caranx sp., and Auxis thazard were infected with high prevalence of Anisakis type I, while Cephalopholis cyanostigma and Rastrelliger kanagurta revealed low prevalence. The mean intensity of Anisakis larvae in K. pelamis and A. thazard was 49.7 and 5.6, respectively. A total of 73 Anisakis type I larvae collected from K. pelamis and A. thazard were all identified as Anisakis typica by PCR-RFLP analysis. Five specimens of Anisakis from K. pelamis and 15 specimens from A. thazard were sequenced using ITS1-5.8S-ITS2 region and 6 specimens from A. thazard and 4 specimens from K. pelamis were sequenced in mtDNA cox2 region. Alignments of the samples in the ITS region showed 2 patterns of nucleotides. The first pattern (genotype) of Anisakis from A. thazard had 100% similarity with adult A. typica from dolphins from USA, whereas the second genotype from A. thazard and K. pelamis had 4 base pairs different in ITS1 region with adult A. typica from USA. In the mtDNA cox2 regions, Anisakis type I specimens from A. thazard and K. pelamis showed similarity range from 94% to 99% with A. typica AB517571/DQ116427. The difference of 4 bp nucleotides in ITS1 regions and divergence into 2 subgroups in mtDNA cox2 indicating the existence of A. typica sibling species in the Makassar Strait.

Case report: Disseminated Cladophialophora bantiana phaeohyphomycosis in a dog with hepatic dysfunction, and concurrent ehrlichiosis.

A 1-year-old mixed breed dog initially presented with marked ascites due to a low-protein transudate resulting from portal hypertension. Laboratory evaluation revealed non-regenerative anemia, lymphopenia, thrombocytopenia, evidence of hepatic insufficiency [hypoalbuminemia, decreased urea, increased post-prandial bile acids, prolonged activated partial thromboplastin time (aPTT)] and Ehrlichia canis infection. Approximately a week later, the dog was declining and was euthanized. On autopsy, multifocal hepatic granulomas and acquired portosystemic shunts (APSS) were seen. Imprint cytology revealed fungal hyphae and pyogranulomatous inflammation in the liver and brain. Disseminated Cladophialophora bantiana phaeohyphomycosis was diagnosed by histologic examination, culture and PCR. Immunosuppression due to ehrlichiosis is suspected to have predisposed this animal to fungal infection. To the authors' knowledge, this is the first report of C. bantiana in the West Indies.