Host Locating Abilities of L. vulgaris Drag Duration 25.0 Min. Drag Distance 1.3 Km. Drag Speed 2.0 Knots/Hr. Depth of Drag 20 - 28 Meters 6.0 Meters Net Width Catch 120 Fish 7800 Meters“ Area Covered Fish Density 65 Meters*/Fish POPULATION SAMPLING DATA table 2 Host Locating Abilities of L. vulgaris (1) The marine parasitic isopod Lironeca vulgaris (Stimpson) is frequently found on several species of fish inhabiting Pacific Ocean coastal waters of the United States. I have observed the isopod in the gill chambers of the flatfishes Parophrys vetulus, Pleuronicthys coenosus, and Citharicthys sp., and the sculpin Leptocottus armatus from Monterey Bay, California. It also occurred in the rockfish Sebastes sp., the Ling Ophiodon elongatus, and the salmon Oncorhyncus kisutch in coastal waters off Southern Oregon. It is likely the parasite infests other fishes aside from those listed. Its distribution is from Baja California to Washington State (Schultz, 1969). The objective of this study was to determine behavior patterns and sensory capabilities utilized by larval Lironeca vulgaris to locate hosts after release from the marsupium of the adult female. The mechanisms of host finding looked for were (1) active use of sense organs of the isopod to locate host fish from a distance. followed by pursuit by the larval isopod, (2) ingestion of the isopod by a host fish, with subsequent attachment of the isopod to the interior of the mouth and movement by the isopod from the mouth into the gill chamber, and (3) random movement by the isopod until chance contact with the host fish occurred. Host Locating Abilities of L. vulgaris BEHAVIOR OF FREE-SWIMMING LARVAL LIRONECA VULGARIS Adult female L. vulgaris containing well-developed brood were isolated individually in aerated containers. Brood released from the marsupium and placed in a larger aquarium moved to the upper and lighted part of an aquarium, generally for the first four to eight hours. They then began swimming near the bottom of the aquarium, touching it at relatively regular intervals. Average distance between touchings was 4.1 cm. (values ranged from two to twelve centimeters) in 26 measurements. Swimming speed during this activity was between 3 cm. and 4 cm. per second. Little variation in swimming speed was noted during the first four days after release from the marsupium. No systematic pattern of swimming was noted. Occasionally the isopod would skim along the bottom, appearing to maintain continuous contact with it. When larval isopods contacted a host they immediately attached using the anterior peraeopods. The contact-recognition- attachment sequence was rapid, requiring less than four seconds. The isopods used were released from the marsupium in vitro; consequently they had never been in contact with fish before. The immediacy of the contact-recognition-attachment sequence therefore indicates a genetically programed behavioral pattern. (2) Host Locating Abilities of L. vulgaris (3) The host fish generally attempted to dislodge newly attached isopods. The Pointed-nosed Sole Parophrys vetulus rapidly quivered the entire body, while the Curl-fin Sole Pleuronicthys coenosus vigorously contorted the whole body. Although P. coenosus was relatively effective in dislodging isopods, P. vetulus rarely dislodged them. For this reason only P. vetulus was used in experiments described later. This behavior of L. vulgaris and host fish was observed throughout the entire six week study, indicating both host and parisite have highly stereotyped behavioral patterns. SENSORY CAPABILITIES OF L. VULGARIS IN HOST LOCATION A 20 gallon aquarium containing a single uninfested P. vetulus was placed in complete darkness. Twenty larval L. vulgaris were then introduced. The number of isopods infesting the fish after six hours was counted. An identical experiment was run concurrently in constant light as a control. The results from four runs are shown in figure 1. The data in figure 1 indicate light, and therefore visual stimulation, is not essential in host location by larval L. vulgaris. (4) Host Locating Abilities of L. vulgaris The following tests for chemoreception were done. A styro- foam tank was fitted with 125 ml. flasks opening into either end (figure 2). A piece of P. vetulus approximately l cm2 was placed in one flask. It had previously been observed that Lironeca vulgaris fed readily on dead P. vetulus. Water was introduced into the tank slowly to establish an unmixed gradient of fish scent. Ten larval L. vulgaris were introduced and observed approximately every two hours during daylight for two days. The experiment was repeated using 20 larval isopods. At no time during the experiments were isopods observed in either flask. In another experiment a live P. vetulus was confined to a single area on a large piece of plankton netting for two hours and then removed, leaving some of the protective slime coating on the mesh. The slime area was outlined in pencil and the netting was stapled into an aquarium parallel to the bottom. The aquarium was filled and approximately 15 larval isopods were introduced. Normal larval swimming behavior was observed outside the slime area. No larvae stopped swimming outside the slime area during three hours of observations. At least five isopods attached to the netting within the slime area for longer than 20 seconds following actual contact with the netting. Swimming activity was resumed after halting, however, indicating the Host Locating Abilities of L. vulgar (5) slime was not sufficient stimulus for permanent attachment. Isopods swimming over the slime area without actually contacting the mesh did not stop. A choice experiment was run utilizing a Y-joint and a gentle current which forced larvae to choose between a tank containing a live fish and an empty control tank (figure 3). The two tanks were filled with an equal volume of water. A single P. vetulus was placed in one tank. Water dripping from the excurrent capillary tube caused a slight current. All tubing and the entire Y-joint was wrapped with black electrical tape to eliminate light gradients. Larval isopods were inserted in pairs into the introduction tube using an eye dropper, and were removed when they emerged from the tubing into the tank. The choice was then recorded. Current flow from each tank was shown to be equal after the experiment by putting methylene blue in one tank. A clear division between blue water from one tank and colorless water from the other was established down the exact center of the choice chamber (figure 3). Data on choice of 74 larvae is shown in table 1. The presence of the fish had no clear effect on the choice of tube. Host Locating Abilities of L. vulgari (6) Throughout the six week study larval Lironeca vulgaris were observed on at least 30 occasions to pass within ten centimeters of the host fish without any visible reaction. The above work indicates L. vulgaris does not use vision or chemoreception as a means of locating hosts beyond a few centimeters distant. It is conceivable that flatfish are infested by random movement of the isopods, as indicated by the following model. Flatfish population data was taken off Hopkins Marine Station in Monterey Bay, using an otter trawl at depths between 20 and 28 meters. The data is shown in table 2. A conservative estimate of the surface area of an average sized adult flatfish is 100 cm2. A strip 10 cm. in width rep- resents a mean between maximum probability of contact (fish oriented perpendicular to the path of the isopod) and minimum probability of contact (fish oriented parallel to the path of the isopod). The width of a swath formed by a swimming isopod is therefore 10 cm. Undamaged one to four day old larvae were rarely observed resting during the six week study. For the purposes of this model the larvae are considered to be constantly swimming. Larval speed is constant at a conservative three centimeters per second in this model. Isopod larvae in aquaria generally swam in straight lines until diverted by obstructions. Isopods will therefore swim in straight lines for model purposes. Hence the area an isopod covers in four days is 1036 me, calculated Host locating Abilities of L. vulgaris (7) by (speed) X (time) X (swath width). Since the flatfish popu- lation density was calculated to be roughly one fish per 65 me the isopod covers approximately 15 times the area necessary to find a single host. The chances of finding a host are undoubtedly reduced by such factors as predation or environmental stress. However, this rough model indicates that out of a brood of 300 larvae there is an excellent chance that one will infest a host through random swimming activity, thereby increasing the isopod population or at least keeping the population constant. Random swimming cannot account for the infestation of the active bottom fish Ophiodon elongatus or the pelagic salmon Oncorhyncus kisutch. Both species are quick enough to easily avoid isopods yet are occasionally infested. Six juvenile 0. elongatus between 12 and 16 cm. in length were exposed to free swimming L. vulgaris. There was no reaction using three larvae between four and five millimeters in length. Three isopods between 1.0 and 1.5 cm. in length were ingested, but all three attached to the interior of the mouth and subsequently crawled into the gill chamber. An unidentified tidepool sculpin took a one centimeter long isopod, which then crawled out the mouth, over the operculum, and into the gill from the outside. In all four observations the isopod avoided being swallowed. Thus ingestion is a possible means by which these fish become infested. No flatfishes were seen feeding on isopods. Host Locating Abilities of L. vulgaris (8) SUMMARY AND CONCLUSION This investigation indicated larval L. vulgaris were inca- pable of sensory perception of host fish without physical contact between isopod and fish. Random activity as a means of infesting flatfish was shown a possibility using behavioral studies in combination with a brief study of flatfish population densities. Infestation by the feeding upon the isopod by fish was shown physically possible, and provided a logical explanation for the occurence of the parasitic isopod on pelagic fish. Much work remains to be done on this subject. REFERENCE Schultz, G.A., Marine Isopod Crustaceans, Wm. C. Brown Co. Publishers, 1969 Host Locating Abilities of L. vulgaris APPENDIX NOTES ON THE NATURAL HISTORY OF L. VULGARIS (1) Damage to gill filiments by isopods could be extensive. Three or more large isopods could prevent closing of the operculum of the host, resulting in death of the host. None of the approximately 300 flatfish examined carried more than three parasites. This suggests the existence of a mechanism or behavior which limits the number of isopods per fish. (2) Effective Lethal Time 50 was recorded for 30 starved larval isopods, with 30 fed isopods as a control. ELT-50 was defined as the time elapsed before half the isopods examined could not swim when stimulated by a current of water from an eye dropper. Fifteen of the thirty isopods were dead or could not swim at the end of six days. (3) Adult L. vulgaris of both sexes survived on a diet of dead fish for at least three weeks. (4) Adult L. vulgaris did not die after releasing larvae. Size variation of 100 brooding females ranged from approximately one gram to 3.25 grams. This suggests the possibility of multiple broods by females. The suggestion is reinforced by the observation of immature eggs in the ovary of a female already Host Locating Abilities of L. vulgaris carrying almost mature brood. (5) The largest L. vulgaris collected was a female (carrying 995 eggs) of length 3.4 cm., maximum width 2.0 cm., and a weight of 3.25 gm. The normal number of eggs for L. vulgaris was estimated at 250 to 400. t- kkt- ttttttttt ++++ ++ tt t ++ ttt ++++ t t +++ Host Locating Abilities +++++++++++ +++++++ vulgaris +++ + ++++ t + + ++++++++ ++ + ++ t ++++ +++ +++++++ ++ ++ ++++++++ +++ ++ +++ t +++++ ++++++ P + tt ++ + +++ +++++ + ++ + + + + + +tt — + + t t ++ +++++++ ++ +++ ++ + P + t + tt ++ + +++ ++++ ++ + + +++ ++++ +++++ ++ ++++ ++ + ++++ +++ ++ + 20 ++++ +++ ++ ++++ ++ ISOPODS 15 INFESTING 10 HOS FISH + + + Run +t Run 2 Run Total Run 3 ++ +++ + tt + +++++ +++ t ++ ++ + + + +++++++ +++ +++ ++++ +++++ ++ t 1 + +++ ++++ + + ++ ++ l t +++ + ++ ++++ ++ ++++ +++ 1 + + + ++ ++++ + + ++ +++ Dark - Control + +++ + + + ++ + + + DARK EXPERIMENT LIGHT + +++ tt +++ figure t + t t +++ +++ + ++ +++ ++ +++++ ++ ++ + + + + + +++ 1 + + ttt ++ +++ ++++++++ +++++++ + ++ +++++ ++++ t tt +++ tttt +++++ + ++ ++ +++ + 0 28.5 cm. Host Locating Abilities of L. vulgaris L 2 39 cm. BAIT FLASK CHEMORECEPTION TEST figure 2 Introduction Point 21m 116.5 em. CONTROL FLASK Host Locating Abilities of L. vulgaris Seawater Source 6.0 12.0 3/8" Tubing 7.0 Introduction Site 1.5 Glass Capillary Tube Y - TUBE EXPERIMENT figure 3 Introduction Tube —as Caplar, Tube L Host Locating Abilities of L. vulgaris RIGHT TANK LEFT TANK No Fish Present Fish Present 15 Isopods 22 Isopods Fish Present No Fish Present 21 Isopods 16 Isopods Y - TUBE EXPERIMENT RESULTS table 1