Thomas G. Rosser

Characterization of the Life Cycle of a Fish Eye Fluke, Austrodiplostomum ostrowskiae (Digenea: Diplostomidae), with Notes on Two Other Diplostomids Infecting Biomphalaria havanensis (Mollusca: Planorbidae) from Catfish Aquaculture Ponds in Mississippi, USA

Abstract Ocular diplostomiasis is caused by trematode species in the family Diplostomidae, specifically those in the genera Austrodiplostomum, Diplostomum, and Tylodelphys. Diplostomid trematodes are globally distributed parasites of fish. Heavy infections of diplostomids that parasitize the eyes of fish can result in acute mortality while chronic infections are often characterized by impaired vision or blindness. In the southeastern United States, commercial catfish production is threatened by piscivorous birds and the many trematode species that parasitize them. The life cycles typically involve a piscivorous avian definitive host, a mollusk first intermediate host, and a fish second intermediate host. A survey of parasites infecting the snail host Biomphalaria havanensis (= B. obstructa) in catfish production ponds was undertaken. Snails were collected from 2 separate ponds during the summer of 2014 and observed for the release of trematode cercariae. A total of 1,740 snails were collected. Three distinct longifurcate pharyngeate cercariae were observed and these cercariae were characterized morphologically and molecularly. Sequencing of ∼4,200 base pairs (bp) of the nuclear ribosomal genes and ∼450 bp of the mitochondrial cytochrome c oxidase gene revealed 3 genetically distinct species. One morphotype shared 99–100% sequence identity with metacercariae from the aqueous and vitreous humors of gizzard shad Dorosoma cepedianum and channel catfish Ictalurus punctatus as well as an adult trematode, Austrodiplostomum ostrowskiae, a parasite of the double-crested cormorant Nannopterum auritus. The remaining 2 cercariae morphotypes shared 99–100% sequence identity with an unidentified Tylodelphys sp. and Austrodiplostomum sp. metacercaria from the brain and eyes of several freshwater fish. Herein we molecularly link the cercaria, metacercaria, and adult stage of the life cycle of A. ostrowskiae, identifying the snail host for this parasite, in addition to providing notes on 2 cercariae representing 2 other diplostomids.

Molecular and Morphological Characterization of Myxozoan Actinospore Types from a Commercial Catfish Pond in the Mississippi Delta

Abstract:  The actinospore diversity of infected Dero digitata was surveyed (May 2011) from a channel catfish (Ictalurus punctatus) production pond in the Mississippi Delta region for the elucidation of unknown myxozoan life cycles. At present, only 2 myxozoan life cycles have been molecularly confirmed in channel catfish, linking the actinospore stage from an aquatic oligochaete (D. digitata) and the myxospore stage from the catfish. In this study D. digitata (n = 2,592) were isolated from oligochaetes collected from the bottom sediment of a channel catfish production pond. After 1 wk of daily observation, a total of 6 genetically different actinospore types were observed. The collective groups were classified as 2 aurantiactinomyxons, 2 helioactinomyxons, 1 raabeia, and 1 triactinomyxon. Overall prevalence of myxozoan infections in the isolated oligochaetes was 4.4%. Actinospores were photographed and measured for morphological characterization. Four previously undescribed actinospore types were identified and characterized molecularly and morphologically. Phylogenetic analysis revealed the raabeia and one of the helioactinomyxon (type 1) actinospores were closely related to the group of myxozoans known to parasitize ictalurids in North America. To date, no myxospores have been linked to the newly sequenced actinospores reported in this survey. The morphological and molecular data generated from this study will assist in the identification of myxospore counterparts for these actinospore stages and aid in the elucidation of unknown myxozoan life cycles in closed production systems.

Clinostomum album n. sp. and Clinostomum marginatum (Rudolphi, 1819), parasites of the great egret Ardea alba L. from Mississippi, USA

Members of the genus Clinostomum Leidy, 1856, colloquially known as yellow grubs, are cosmopolitan parasites of piscivorous birds, freshwater snails, fish and amphibians. In the southeastern United States, piscivorous birds present a continuous challenge for producers of farm-raised catfish. Ciconiiform birds are common hosts of Clinostomum spp. in North America and are endemic on most commercial catfish operations. The great egret Ardea alba L. is an avian predator often found foraging on commercial catfish operations, but to date the trematode fauna of great egrets preying on catfish ponds remains mostly understudied. Thirteen great egrets were captured from commercial catfish ponds in northeast Mississippi, and examined for trematode infections. Two morphologically distinct Clinostomum spp. were observed in the great egrets sampled, one morphologically consistent with Clinostomum marginatum (Rudolphi, 1819) and one morphologically unique species. These morphological descriptions were supplemented with molecular sequence data (c.4,800 bp of ribosomal DNA and c.600 bp of mitochondrial DNA). Gene sequences confirmed the identification of C. marginatum. However, the second species differed significantly from its congeners in both morphology and DNA sequence. Given these distinct morphological and molecular characters we propose this second species as Clinostomum album n. sp.

Biomphalaria havanensis is a Natural First Intermediate Host for the Trematode Bolbophorus damnificus in Commercial Catfish Production in Mississippi

AbstractThe digenetic trematode Bolbophorus damnificus is a significant hindrance to the production of farm-raised catfish in the southeastern USA. Severe infections with Bolbophorus damnificus can result in death, but the real damage lies in mild to moderate infections, which can go unnoticed by producers. Research has shown even mild infections can inhibit production to the point of operating at a net loss. The ramshorn snail Planorbella trivolvis is the first intermediate host for Bolbophorus damnificus and is prevalent in most commercial catfish ponds in the Southeast. Management practices aimed at controlling Bolbophorus damnificus infections primarily focus on reducing snail populations in ponds. Other aquatic snail species are also commonly associated with commercial catfish ponds, although little is known about their contributions to trematode infections in catfish. In the summer of 2014, 804 snails Biomphalaria havanensis (synonym: Biomphalaria obstructa) were collected from a single commercial c...

Myxobolus lepomis n. sp. (Cnidaria: Myxobolidae), a gill myxozoan infecting Lepomis marginatus Holbrook and Lepomis miniatus Jordan (Perciformes: Centrarchidae), in the Big Thicket National Preserve, Texas, USA

A parasitological survey of freshwater fishes in the Big Thicket National Preserve in southeast Texas revealed myxozoan infections in two species of sunfish, Lepomis marginatus Holbrook and Lepomis miniatus Jordan (Perciformes: Centrarchidae). Pseudocysts were elongate-oval, 988 × 485 µm (ex L. marginatus) and 800 × 606 µm (ex L. miniatus) and demonstrated a predilection to the edge of the primary gill lamellae. Myxospores consistent with the genus Myxobolus were oblong, 16.8–21.3 (19.0 ± 0.9) µm long, 7.0–8.8 (7.9 ± 0.5) µm wide and 5.3–6.1 (5.8 ± 0.3) µm thick (ex L. marginatus) and 17.2–20.3 (18.8 ± 0.7) µm long, 7.5–9.9 (8.7 ± 0.6) µm wide, and 6.8–7.2 (7.0 ± 0.2) µm thick (ex L. miniatus); with 2 pyriform polar capsules 8.3–9.8 (9.0 ± 0.5) µm long, 2.2–2.7 (2.5 ± 0.2) µm wide (ex L. marginatus) and 9.2–10.5 (10.0 ± 0.4) µm long, 2.2–3.0 (2.8 ± 0.2) µm wide (ex L. miniatus). Statistically, the measurements of spore body width, polar capsule length, and polar capsule width were significantly different between myxospores from L. marginatus and L. miniatus. However, intraspecific genetic variability between isolates at the 18S rRNA gene was negligible, with < 0.8% variability across > 2,000 bp of sequence. The isolates shared no significant sequence similarity with any myxozoan deposited in the GenBank nucleotide database. Phylogenetic analysis inferred from the 18S rRNA gene from both L. marginatus and L. miniatus placed the isolates within a clade of myxozoan parasites of perciform fishes. Based on shared tissue and host family tropism, overlapping morphological characters and high degrees of sequence conservation at the 18S rRNA gene, we propose these isolates as morphologically distinct, genetically conspecific representatives of M. lepomis n. sp. from the gills of L. marginatus and L. miniatus in the Big Thicket National Preserve in Texas, USA.

Alligator wrestling: morphological, molecular, and phylogenetic data on Odhneriotrema incommodum (Leidy, 1856) (Digenea: Clinostomidae) from Alligator mississippiensis Daudin, 1801 in Mississippi, USA

Based on specimens collected from harvested American alligator Alligator mississippiensis Daudin, 1801 in Mississippi, USA, novel molecular data for both nuclear ribosomal genes (18S, ITS1-5.8S, ITS2, and 28S) and mitochondrial genes (cytochrome c oxidase subunit 1 and nicotinamide adenine dinucleotide dehydrogenase subunit 1) are provided for Odhneriotrema incommodum (Leidy, 1856), a trematode of the family Clinostomidae Lühe, 1901 infecting A. mississippiensis and the Florida spotted gar Lepisosteus platyrhincus DeKay, 1842. This represents the first sequencing data available for the genus Odhneriotrema and the subfamily Nephrocephalinae Travassos, 1928. Additionally, the results of phylogenetic analyses, additional morphometric data, a photomicrograph, and a line drawing supporting the present identification of O. incommodum are provided. These data will aid in elucidating the life cycle of O. incommodum through molecular identification of larval stages as well as understanding the evolutionary history of Clinostomidae and its subfamilies. Implications for the currently accepted organization of the Clinostomidae are discussed.

North and South American Haplotypes of Drepanocephalus auritus (Digenea: Echinostomatidae) Are Released from Biomphalaria havanensis (Mollusca: Planorbidae) Inhabiting Catfish Aquaculture Ponds in Mississippi, U.S.A.

ABSTRACT: Trematodes of the subclass Digenea are a major impediment to catfish aquaculture in the southeastern United States. Catfish production ponds are ideal feeding grounds for piscivorous birds, and given the variety of aquatic snails that inhabit these ponds, they offer a prime environment for the propagation of digenetic life cycles. In July 2014, snails morphologically consistent with Biomphalaria havanensis L. Pfeiffer, 1839, were collected from 2 different catfish production ponds in the catfish farming region of eastern Mississippi. These snails were refractory to Schistosoma mansoni Sambon, 1907, infection, and phylogenetic analyses based on internal transcribed spacer (ITS)1 and ITS2 fragments placed the snails within the B. havanensis/obstructa complex. Some (1.5%) of these snails shed cercariae morphologically and molecularly consistent with Drepanocephalus auritus Kudlai, Kostadinova, Pulis, and Tkach, 2015 (formerly reported as Drepanocephalus spathans Dietz, 1909), an echinostomatid parasite of the double-crested cormorant Phalacrocorax auritus Lesson, 1831, and channel catfish Ictalurus punctatus Rafinesque, 1818. Juvenile channel catfish were challenged individually with ∼275 cercariae of D. auritus per fish, resulting in limited mortality (2 out of 10 fish). Surviving fish were euthanatized 7 d postchallenge, and the presence of metacercariae was confirmed histologically. Parasite-associated pathology was consistent with previous studies. Subsequent molecular analysis of 5 different gene targets confirmed morphological identifications. Previous research has suggested the existence of discrete North American and South American haplotypes of D. auritus. Mitochondrial cytochrome c oxidase subunit 1 and nicotinamide adenine dinucleotide dehydrogenase subunit 1 sequences demonstrated that both haplotypes were released by snails in this study, suggesting a broader distribution of these genetic variants than previously thought. Along with Planorbella trivolvis Say, 1817, this work identifies a second snail species common to catfish aquaculture ponds that releases D. auritus infective to juvenile channel catfish and is the first report of B. havanensis as an intermediate host for D. auritus in North America.

Morphological, Histological, and Molecular Description of Unicauda fimbrethilae n. sp. (Cnidaria: Myxosporea: Myxobolidae) from the Intestinal Tract of Channel Catfish Ictalurus punctatus.

Abstract:  The channel catfish Ictalurus punctatus is a known host for 10 species of Henneguya, but few other myxozoan genera are described from this species. Unicauda is a genus of myxozoan parasites within the family Myxobolidae that consists of 10 valid species from freshwater fish. Herein, we describe a novel species of Unicauda from the intestinal tract of farm-raised channel catfish in Mississippi. Myxospores were consistent with the genus Unicauda but exhibited a unique branching at the terminal end of the caudal process that has not previously been reported. Myxospores measured 90.39 ± 14.97 μm (mean ± SD; range = 70.88–126.02 μm) in total length. The spherical spore body measured 7.31 ± 0.26 μm (6.75–7.84 μm) in length and 7.01 ± 0.63 μm (6.1–8.01 μm) in width. The 2 polar capsules measured 3.45 ± 0.33 μm (3.02–4.03 μm) in length and 2.65 ± 0.32 μm (2.18–3.11 μm) in width. The single caudal process measured 82.98 ± 14.97 μm (63.39–118.63 μm) in length from the base of the spore body to the end of the most terminal projection. Terminal projections measured 26.83 ± 8.8 μm (12.34–42.29 μm) in length and 0.95 ± 0.23 μm (0.52–1.6 μm) in width. The 18S rRNA gene sequence obtained did not match any published sequences. Given the uniqueness of the myxospore morphology, histological presentation, and gene sequence data, we describe this as an unreported species, Unicauda fimbrethilae n. sp.

New data on Neodiplostomum americanum Chandler and Rausch, 1947 (Digenea: Diplostomidae), in the Great Horned Owl Bubo virginianus Gmelin, 1788 and the Eastern Screech Owl Megascops asio Linnaeus, 1758 in Mississippi, USA

Neodiplostomum americanum Chandler and Rausch, 1947 has been reported from six species of owls in North America. At present, there are no molecular data for this species and gene sequence data from Neodiplostomum Railliet, 1919 are limited. A freshly deceased specimen of the Great Horned Owl Bubo virginianus Gmelin, 1788 and a freshly deceased specimen of the Eastern Screech Owl Megascops asio Linnaeus, 1758 were collected in Oktibbeha County, Mississippi in 2014 and 2016, respectively. Neodiplostomum americanum were recovered from both hosts. Herein, updated morphological descriptions are supplemented with gene sequence data from conserved (18S, ITS1-5.8S, ITS2, and 28S rRNA) and fast-evolving (cytochrome c oxidase subunit 1 mtDNA) regions. Preliminary phylogenetic analysis of the genus based on cytochrome c oxidase subunit 1 gene sequence data supports the placement of N. americanum within a discrete phylogroup of the family Diplostomidae. The life history of N. americanum is unknown and currently limited to the description of the adult stage in avian hosts. The molecular data generated in this study offer insight into the phylogenetic placement of N. americanum within the Diplostomatidae and will aid in identifying different life stages in putative intermediate hosts.

Myxobolus ictiobus n. sp. and Myxobolus minutus n. sp. (Cnidaria: Myxobolidae) from the gills of the smallmouth buffalo Ictiobus bubalus Rafinesque (Cypriniformes: Catostomidae)

The smallmouth buffalo Ictiobus bubalus Rafinesque (Catostomidae) is native to North American waterways and occasionally grown in pond aquaculture. Species of Myxobolus Bütschli, 1882 have been reported from the gills, integument, and intestinal tract of buffalo fish, although there is ambiguity in some host records. In the summer of 2013, thirteen adult smallmouth buffalo were seined from a 0.1-acre (0.04-hectare) experimental research pond at the Thad Cochran National Warmwater Aquaculture Center in Stoneville, Mississippi, USA, and examined for the presence of parasitic infection. Two previously unknown species of Myxobolus were observed parasitising the gills. Plasmodia of the two species differed from each other in both size and shape. Morphologically the two species were distinct from one another and from other Myxobolus spp. previously reported from buffalo fish. Myxospores of Myxobolus ictiobus n. sp. were spherical and measured 12.7–14.5 (13.9 ± 0.4) µm in length and 10.7–13.6 (12.5 ± 0.7) µm in width with a thickness of 10.3–14.8 (12.6 ± 2.3) µm. Polar capsules measured 5.6–7.4 (6.6 ± 0.4) µm in length and 3.7–4.9 (4.5 ± 0.8) µm in width and each contained a coiled polar filament with 5–6 turns. Myxospores of Myxobolus minutus n. sp. were circular in shape and measured 7.4–9.6 (8.6 ± 0.7) µm in length and 7.5–9.9 (8.8 ± 0.7) µm in width with a thickness of 6.5–7.3 (6.7 ± 0.3) µm. Polar capsules measured 3.6–4.9 (4.3 ± 0.3) µm in length and 2.8–3.8 (3.3 ± 0.3) µm and each contained a coiled polar filament with 5–6 turns. Supplemental 18S rRNA gene sequencing identified unique sequences for each isolate. Phylogenetic analysis of 18S rRNA sequences demonstrated a strong clustering of both isolates with other species of Myxobolus from cypriniform fish.