Proterometra macrostonia Fishes — Rosen et al. 



103 



with all three egg stages as Type III (18 days 

 or older; Figure Id). 



Individual snails were separated into plastic 

 boxes divided into compartments containing 

 filtered stream water, held in an environmental 

 chamber at 20°C with a 12-hr light: 12-hr dark 

 cycle, and fed lettuce ad libitum. When cer- 

 cariae were required for experiments, any pre- 

 viously emerged larvae were removed and the 

 host snails held for 12 to 24 hr under contin- 

 uous light. The light source was switched off, 

 promoting a copious release of new cercariae 

 within 2 hr for experimental infections of 

 young, hatchery-reared bluegill (Farley and 

 Farley Farms, Cash, Arkansas). The following 

 experiments were designed to establish an 

 equal mix of different age/type worms in in- 

 dividual fish to confirm the use of egg stages 

 for differentiating between new vs. older 

 infections. 



In the first experiment, each of four bluegill 

 was individually placed in separate 1 -gallon 

 tanks with three cercariae in each tank so par- 

 asite ingestion by each host could be visually 

 confirmed. Infections with three additional 

 cercariae were repeated with the same fish 7 

 days later (i.e., each fish was exposed to a 

 total of six cercariae resulting in 8- and 15- 

 day old worms at the termination of the 15- 

 day experiment). The same protocol was fol- 

 lowed in a second experiment with 14 

 bluegill, but additional infections were con- 

 ducted at 7 and 14 days following initial ex- 

 posure (i.e., each fish was exposed to a total 

 of nine cercariae resulting in 6-, 13-, and 20- 

 day old worms at the end of the 20-day study). 

 Following ingestion, fish were maintained 

 within their respective groups in aerated 38- 

 liter aquaria at 24°C and fed Tetramin (Tetra 

 Sales, Blacksburg, VA). At 15- and 20-days 

 following exposure to the initial set of infec- 

 tions, all fish were necropsied and their worm 

 types determined. A Student's ?-test (15-day 

 experiment) and a one-way ANOVA (20-day 

 experiment) were used to assess possible dif- 

 ferences in the mean number of worm types 

 recovered. In addition, the distribution of egg 

 developmental stages (I to III) in each worm 

 type was determined in the 20-day study of 

 bluegill. 



Twenty-one additional hatchery-reared 

 bluegill were experimentally infected as pre- 

 viously described with four cercariae each for 



BWarmouth 



II III 

 Adult Worm Types 



Figure 2. Relative frequency of Type I to III Protero- 

 metra macrostonia adults from natural infections of war- 

 mouth (N = 18 fish and 1524 worms), bluegill (N = 32 

 fish and 503 worms), and longear sunfish (N = 30 fish 

 and 220 worms). 



assessment of histopathology. Seven fish were 

 dissected on days 7, 14, or 24 post-infection 

 (PI), respectively. The esophagus and stomach 

 of each fish were removed, fixed in buffered 

 formalin, routinely processed for paraffin em- 

 bedding, and serially sectioned with a rotary 

 microtome. Slides were stained with hematox- 

 ylin-eosin or hematoxylin in combination with 

 Gomori's trichrome. Several esophagi and 

 stomachs from naturally infected bluegill and 

 warmouth were fixed and assessed in a similar 

 manner. 



RESULTS 



The mean intensity and prevalence of P. 

 macrostonia in naturally infected warmouth, 

 bluegill, and longear sunfish from North Elk- 

 horn Creek were 85.8 ± 16.2 (prevalence = 

 100.0%), 15.7 ± 2.4 (prevalence = 88.9%), 

 and 7.3 ± 3.1 (prevalence = 67.0%), respec- 

 tively. Between 50.5 and 74.4% of the P. ma- 

 crostoma populations in these fish were dom- 

 inated by Type I worms (Figure 2). Smaller 

 numbers of Type II (11.4 to 20.5%) and III 

 (14.6 to 29.7%) worms were also present (Fig- 

 ure 2). 



In our experimental infections of bluegill, 

 95.8% (23/24; 15-day experiment) and 87.3% 

 (1 10/126; 20-day experiment) of the cercariae 

 originally ingested were recovered as adult 

 worms. The results of a Student's r-test 

 showed no significant difference (t = 0.522; 

 6 df; sig. = 0.620) in the average number of 

 Type I (N = 12) and Type II (N = 11) worms 



