ICES CM 2008/D:12

Extended Abstract for Theme Session D

Dichelesthium oblongum (Copepoda:Dichelesthiidae) infection in wild caught Atlantic , oxyrinchus .

Mark S. Sokolowski1, Keith J. Dunton1, Paul R. Bowser2 and Mark D. Fast1.

School of Marine and Atmospheric Sciences, Stony Brook University, Stony Brook, NY 11790-50001: Aquatic Health Program, Department of Microbiology and Immunology, College of Veterinary Medicine Cornell University Ithaca, NY 14853-64012

Atlantic sturgeon were sampled off Rockaway Beach, Long Island, in October and November 2007 to initiate a study on the population dynamics of this threatened species. During the tagging and release of fish, they were examined for external parasites and non-invasive samples of blood and gill were taken. Of the 43 fish examined for external parasites, the prevalence was 0.05% for an unknown species of isopod, 26% for Caligus elongatus, 65% for Nitzschia spp., and 91% for Dichelesthium oblongum. During the course of these examinations, macroscopic lesions associated with the attachment and feeding of D. oblongum were observed (Figure 1).

A B C

Figure 1. Juvenile Dichelesthium oblongum infection on the operculum (A), base of

the fins (B) and adult copepods within the oercular cavity (C) of

(scale bar in mm). Note the petechial hemorrhaging of the host tissue associated B

with adult egg-producing female copepods (C).

The juvenile Dichelesthium oblongum lesions (Figs. 1A, B) consisted of focal areas of ulceration with a raised border. Histologically the lesions were characterized by necrosis or complete loss of both epidermis and dermis as well as focal necrosis of the underlying musculature. Focal epidermal spongiosis was observed in close proximity to lesions where the epidermis was still intact (Figure 2).

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Figure 3. Atlantic sturgeon epithelium adjacent to juvenile Dichelesthium

oblongum infection (left) and away from infection (right). Note the intercellular

edema and erythrocyte infiltration into the epithelium on the left (arrows).

Abundances of D. oblongum ranged from 0-40 per individual sturgeon. The parasites present consisted mainly of gravid females that were attached to opercula cavities and occasionally on the gill or gill arch (119/252, 47%). A further 32 D. oblongum, all of which were juvenile stages, were observed attached to other external body surfaces (i.e. mouth and fins). Due to the large size of the lesions (i.e. diameter larger than the length of the parasite in all cases; 7-12 mm) associated with juvenile D. oblongum and the overall high parasite prevalence and abundance, an investigation was undertaken to determine if any physiological changes might be occurring in the most heavily infected individuals. We utilized blood chemistry analysis to determine if such changes were present. Serum chemistry of the most heavily infected indicated ion loading (i.e. sodium, calcium, magnesium) most likely due to the stress related to parasite load but may also be a result of water leakage through the epithelial breach. Infected individuals were classified into three groups according to their stage- specific parasite abundances: (1) 6 or greater gravid females (>6 GF); (2) 1-3 gravid females (1-3 GF); and (3) no gravid females (0 GF). These groupings were chosen due to, (1) the extremely high prevalence (91%) of D. oblongum resulting in a very small sample size of non-infected individuals, (2) the unknown history prior to sampling making it impossible to determine whether uninfected individuals had previously been exposed to the parasite or not, (3) the lack of affect/host response often associated with juvenile copepod parasite infections (Johnson and Fast, 2004; Jones et al., 1990; Tully and Nolan, 2000; Wagner et al., 2008), (4) the effects of mobile/adult life stages of parasitic copepods particularly at high abundances (Bjorn and Finstad, 1997; Bowers et al., 2000; Grimnes and Jakobsen, 1996; Wagner et al., 2008), and (5) the normal and rather equal distribution of D. oblongum gravid females. Using the number of gravid females to identify different exposure groups also strongly correlated with total D. oblongum abundances (Pearson correlation 0.83), such that mean abundances ±SEM for the three groups were 22.7 ± 3.98 (>6 GF), 8.38 ± 0.95 (1-3 GF), and 5.0 ± 1.68 (0 GF). The life stages of the parasite present on the host are not only indicative of exposure ‘time’ to infective copepods but can also indicate site of infection (i.e. body location) and the general composition of the parasite meal. As mentioned earlier, in the ICES CM 2008/D:12

case of D. oblongum adult females were only observed within the opercula cavities and on the gills and gill arches, a site more accessible to host blood than the fins or other body attachment sites. Furthermore, production of eggs by parasitic is often associated with an increased requirement for host blood (i.e. mosquitoes, ticks, gill maggots, etc.). To our knowledge this is the first work on clinical serum chemistry of marine stage A. oxyrhynchus and therefore there is little to no comparative literature. Previous work on 2-yr old freshwater A. oxyrhynchus (Baker et al., 2005) and 2-yr old A. persicus maintained in seawater (Kazemi et al., 2006) suggests that serum sodium, calcium and magnesium levels are all much higher in the fish sampled here. However, recently we have obtained information that the serum ion levels in the 0 GF group were within physiological ranges for seawater adapted Atlantic sturgeon (S. McCormick Pers. Comm.). Regardless of whether the levels reported here are indicative of baseline or physiologically optimal levels for this species, there were significant differences between the differentially infected groups. The >6 GF group had significantly higher levels of serum sodium and magnesium when compared to the 0 GF group, whereas hosts infected with 1-3 GF had significantly higher serum sodium, calcium and magnesium, when compared to those sturgeon without gravid female (Figure 3).

180 13 L)

12

170 11

10 160

9 Serum Sodium (mEq/ Sodium Serum Serum Calcium (mg/dL) Calcium Serum 150 8 0 1-3 >6 0 1-3 >6 NumberNumber of of Gravid Adult Female Females Egg ers Number of of Adult Gravid Female Females Eggers

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5.5

4.5

3.5

2.5 Serum Magnesium (mEq/L) Magnesium Serum 0 1-3 >6 NumberNumber of ofAdult Gravid Female Females Eggers

Figure 3. Serum sodium, calcium, and magnesium levels in Atlantic sturgeon at differing infection levels with Dichelesthium oblongum. (0) signifies the lack of gravid female (GF) D. oblongum on the sturgeon host; (1-3) signifies the presence of 1, 2 or GF D. oblongum on the sturgeon host; (>6) signifies the presence of 6 or more GF D. oblongum on the sturgeon host. * signifies significant differences from the 0 AFE group (ANOVA, P<0.05).

Serum bicarbonate was lower in sturgeon infected with >6 GF and anion gap significantly higher, when compared to sturgeon without GF (0 GF). In both of these cases, the levels observed for sturgeon infected with 1-3 GF were intermediate to the two. Finally, aspartate amino transferase (AST) was observed to be elevated in both the sturgeon infected with 1-3 GF and >6 GF when compared to 0 GF, but only significantly in the 1-3 GF group. The increased serum ion concentrations were similar to findings in Atlantic salmon (Salmo salar) and sea trout (S. trutta) infected with the parasitic copepod, Lepeophtheirus salmonis (Bjorn and Finstad, 1997; Grimnes and Jakobsen, 1996). This may be a result of osmoregulatory imbalance (i.e. water loss) due to decreased osmotic competence from elevated cortisol, the extensive lesions and breached epithelial layer, as well as reduced function of Na+K+ATPase (important in NaCl regulation) in gill filaments damaged by parasitic copepod attachment and feeding (Bjorn and Finstad, 1997; Grimnes and Jakobsen, 1996; Nolan et al., 1999). The lower bicarbonate concentrations and elevated anion gap were also suggestive of metabolic acidosis (Kraut and Madias, 2007). This is due to a reduction in the buffering capacity of the serum through the loss of bicarbonate. Metabolic acidosis can be brought on by strenuous activity and lactic acid build up. Furthermore, the observed elevation in AST was suggestive of some tissue (i.e. liver or kidney) damage, which taken together with serum ion build up and metabolic acidosis, suggests possible osmoregulatory stress and subsequent tissue failure in the most heavily infected individuals over a much longer term. The effects of this parasite on host life history characteristics may have significant ecological impacts to this fish species of concern and requires further investigation.