Parasitic worms of fishes in tributaries of Otsego Lake F. B. Reyda 1 and D. D. Willsey 2 INTRODUCTION This particular study is part of an ongoing survey of the parasites of fishes of Otsego Lake that began in September 2008. The ultimate goal of this survey is to identify as many species of parasitic worms as possible from the diversity of fishes that occur in Otsego Lake, which is home to over 35 species of fish (MacWatters 1980), and to provide a host-parasite checklist for the watershed. Understanding the parasite diversity in Otsego Lake allows for insights to the ecology of Otsego Lake, and it facilitates future research by Reyda and his students. Results of earlier phases of this survey were included in previous annual reports (Hendricks & Reyda 2010; Reyda 2009; 2010; Szmygiel & Reyda 2011). Throughout this ongoing survey the focus has been on finding and identifying parasitic worms that occur within the digestive system, in particular the intestine, of the fishes examined, though other organs of the fish have also been examined. Prior to 2012, the fish that were collected and examined for parasites were predominantly centrarchids (e.g., smallmouth bass) and the percid, Perca flavescens (yellow perch), owing to the emphasis on angling as the sampling technique. Collection efforts in 2012 emphasized other fish groups, namely cyprinids, many of which cannot be caught via angling. In 2012 collection efforts were focused on Otsego Lake tributaries, as well as the Susquehanna River, and nearby Oaks Creek, which flows from Canadarago Lake to the Susquehanna below Cooperstown. These stream collections, emphasizing different fish groups than those previously studied, were possible because of the recent acquisition of a backpack electro-fisher. The present study summarizes the fish species that were collected as part of this fish parasite survey in 2012, and provides comments on the parasite species encountered, several of which await species-specific identification. METHODS A Hallteck HT-2000 battery backpack electrofisher and accessories were used to stun fish at several sites in the vicinity of Otsego Lake between June and November 2012. Fish collections were done in compliance with a NY State DEC permit to F. Reyda (License No. 1647). Fish that were stunned were preliminarily identified while in the field and subsequently kept or released, based on desirability for the fish parasite survey. Fish that were kept were transported back to the BFS in an aerated portable container and stabilized in aquaria according to collecting site. Fish were necropsied 0 14 days after collection as follows: Fish were initially anesthetized in FinQuil and/or were double-pithed with a knife incision. Fish were then identified to species with the aid of an identification key and reference website (Smith 1985; http://fish.dnr.cornell.edu/nyfish/fish.html). 1 Assistant Professor, SUNY Oneonta Biology Department. 1 BFS Intern, summer 2012. Current affiliation: SUNY College at Oneonta.
A subset of collected fish were preserved in 10% formalin for the BFS reference collection. All fish were assigned a field code and were photographed with digital camera, and tissue samples from nearly all fish were preserved in vials containing molecular-grade 100% ethyl alcohol and stored in a -80 C freezer to enable future molecular studies. The necropsy was initiated with a ventral incision starting at the anus, and the viscera were subsequently removed and isolated in a petri dish filled with tap water. The stomach and the intestine of all fish dissected were examined with the aid of a dissecting microscope. In addition, in most fish examined, the external surface, including the eyes and fins, as well as the gills, liver, heart and body cavity, were also examined. All parasites encountered were preserved in vials of either 4 or 10% formalin or 70% ethyl alcohol (for future morphological study) or in molecular-grade 100% ethyl alcohol (for future DNA sequence analysis). A subset of the parasitic worms encountered and preserved for morphological study were subsequently prepared as whole-mount microscope slides as follows: worms were hydrated and stained in Delafield s hematoxylin, and subsequently dehydrated in a graded ethanol series, cleared in methyl salicylate, and mounted in Canada balsam. Once dried, whole-mount microscope slides were examined with a compound microscope and initial identifications were made with the aid of a general fish-parasite identification reference (Hoffman 1999). In addition, a subset of the worms saved in molecular grade ethanol, mainly metacercaria of digenetic trematodes encountered in the body cavity or in other organs, were sent to colleagues at Environment Canada (Montreal, Canada) who intend to obtain DNA sequence data for the mitochondrial cytochrome oxidase I gene, and make that data publicly available, as part of the Barcode of Life Initiative (http://www.barcodeoflife.org/). RESULTS Table 1 summarizes the number of fish species encountered at each of the collecting sites, which included: Oaks Creek (site 7), Susquehanna River (near Cooperstown), Hayden Creek; Shadow Brook, and Leatherstocking Creek. In total, 138 specimens representing 26 species of fish, including 16 cyprinid species, were collected and examined for parasites. More than 10 specimens of each of the following fish species were collected: blacknose dace, brown trout, creek chub, longnose dace, margined madtom, and white sucker. Ten of the 26 species of fish that were collected were previously sampled relatively well during the Otsego Lake portion of the survey, such as the smallmouth bass and rock bass. Sixteen of the 26 species encountered had not been previously collected as part of the fish parasite survey. Among the field sites chosen, Oaks Creek was by far the most diverse; we collected 18 species from that site. Preliminary parasitological data are also included in Table 1. The number of species of parasitic worms that were encountered in the digestive system of each fish species is indicated, as is the number of parasite species that were encountered in sites other than the digestive system. Parasites encountered outside the digestive system included parasitic worms, as well as copepods, leeches and protists. Although the longnose dace was the most abundant fish encountered (21 individuals), it did not have the highest overall parasite diversity, with a total of four parasite species. The smallmouth bass was infected with the greatest diversity of parasites, with four species of intestinal worms, all of which were previously encountered in smallmouth bass in Otsego Lake, and with three species of parasites outside of the intestine, including an
unidentified leech externally, the copepod Ergasilis centrarchidarum on the gills, and an identified species of larval tapeworm on the gonads. Most of the species noted in Table 1 as occurring in non-intestinal sites were metacercaria, a larval stage of digenetic trematodes. Metacercaria were not identified to species, so the total number of digenetic trematodes obtained in this stream fish survey is not currently known.
DISCUSSION The use of the backpack electrofisher and emphasis on stream sites made possible the collection of a diversity of fishes not previously encountered in the ongoing fish parasite survey, especially species of cyprinids. The resulting inclusion of stream fishes, however, did little to increase our list of intestinal parasitic worm species known from this area. Nearly all of the parasitic worms encountered in the digestive system of streams associated with Otsego Lake were already known to occur in fishes in Otsego Lake, such as the acanthocephalans Leptorhynchoides thecatus and Neoechinorhynchus rutili, the nematodes Spinitectus carolini and Spinitectus gracilis, and the cestode Proteocephalus cf ambloplites. The only intestinal parasite not previously encountered in the survey was a species of digenetic trematode currently being identified to species from the northern hogsucker. Cyprinids are generally poor hosts of intestinal worms. In other words, cyprinids do not typically serve as definitive hosts for parasitic worms that mature and complete their life cycle in the intestine. Ten of the 16 species of cyprinids examined had no parasites occurring in their digestive system, and the other 6 cyprinid species had only one or two species of parasitic worms in the intestine (Table 1). On the other hand, cyprinids serve an important role as intermediate hosts of parasites, parasites that occur in locations other than the digestive system (Hoffman 1999). Eleven of the 16 cyprinid species examined were infected with at least one species of non-intestinal parasite, typically the metacercaria stage of digenetic trematodes, in the body cavity, the liver, the eyes, or some other non-intestinal site. Digenetic trematodes, such as the well-known yellow grub a worm encountered throughout our survey work in both Otsego Lake and the streams utilize the fish as the second intermediate host in the life cycle, which can only be completed when the intermediate fish host is preyed upon by a piscivorous bird, such as a heron, where the worm will develop to the adult stage, mature, and mate (Hoffman 1999). Although five species of cyprinids were found not to host any parasites outside of the digestive system, it is assumed that these species also serve as intermediate hosts of metacercaria in these streams but that we failed to encounter infections owing to small sample sizes. For example, only one specimen of eastern silvery minnow was examined (see Table 1). Our understanding of the diversity of digenetic trematodes awaits species-specific identifications of the plethora of metacercaria that were encountered in this survey. Because metacercaria are immature worms, there are relatively few morphological features to facilitate their identification, and morphology-based identification of metacercaria to species is virtually impossible (Hoffman 1999). It is our hope that our collaboration with colleagues at the Barcode of Life Initiative in Canada will result in available sequence data of some of these unknown species and shed light on the species diversity of fish trematodes in Otsego Lake and its tributaries. In addition to the stream sampling summarized here, fish were collected directly from Otsego Lake in 2012 as part of the fish parasite survey. Those data, however, are not reported here. Rather, those data will be included in a more comprehensive summary of the parasitic worms of fishes of Otsego Lake and its tributaries that will be provided in the future.
ACKNOWLEDGEMENTS Thanks to Mark Cornwell for providing training on using the electro-fisher, and thanks to him and Justin Hurlbert, both of SUNY Cobleskill, and to Matt Albright, Holly Waterfield and Jeff Heilveil of SUNY Oneonta, for assistance with electro-fishing. The following students contributed to this study: Austin Borden, Rebecca Russell, Amanda Sendkewitz and Joe Westenberger. This study was funded in part by the SUNY Oneonta Research Foundation and an NSF FSML grant to W. Harman (DBI 1034744). REFERENCES Hendricks, L. & F. B. Reyda. 2010. A survey of the acanthocephalan parasites of fish species of Otsego Lake, NY. In 41 st Annual Report of the SUNY Oneonta Biological Field Station. Pp. 272 275. Hoffman, G.L..1999. Parasites of North American Freshwater Fishes. Cornell University. Second Edition. 576 p. MacWatters, R. C. 1980. The fishes of Otsego Lake, Biological Field Station, Cooperstown, New York. Occasional Paper No. 7. 52 p. Reyda, F. B. 2009. Fish parasite survey, 2008. In 41 st Annual Report of the SUNY Oneonta Biological Field Station. Pp. 153 157. Reyda, F. B. 2010. Parasitic worms of fishes of Otsego Lake and nearby water bodies, 2009. In 42 nd Annual Report of the SUNY Oneonta Biological Field Station. Pp. 276 281. Smith, C. L. 1985. The Inland Fishes of New York State. The New York State Department of Environmental Conservation. 287-309. Szmygiel, C. & F. B. Reyda. 2011. A survey of the parasites of Smallmouth bass (Micropterus dolomieu) In 43 rd Annual Report of the SUNY Oneonta Biological Field Station. Pp. 235 240.