Contributions to Zoolog)>, 69 (1/2) 9-20 (2000)

SPB Academic Publishing bv, Amsterdam

The distribution of exocrine glands in Lepeophtheirus salmonis and Caligus elongatus (Copepoda: Caligidae)

S. Bell,J.E. Bron & C. Sommerville

Institute ofAquaculture, University of Stirling, Stirling, Scotland FK9 4LA

Keywords: gland, sea lice, diaminobenzidine, peroxidase, Caligidae, Copepoda

Abstract Adults 15

Caligus elongatus 16

Other species 17 The morphology, function and distribution of exocrine glands Discussion 17 of copepods have rarely been studied in detail and almost not- References 19 is known hing about them in the sea lice species L. salmonis

andC. elongatus. This study utilised a novel application of a of light-microscopy staining technique to reveal a variety glands

m nauplius, copepodid, chalimus, preadult and adult stages. Introduction The stain, 3’,3-diaminobenzidinetetrahydrochloride (DAB),

applied to fresh exocrine material, differentiated a population of Descriptions of glandular structures of copepods glands and stain was enabled a study of their distribution. The have been a feature of the literature for successful with many in highlighting the ducts and pores associated the decades larger glands. The locations of gland sub-populations were (Richards, 1891; Yonge, 1932; Clarke,

conserved in all life-stages, although glands in the swimming Conover, David, & Nicol, 1962; Fahrcnbach, 1962;

legs were found better in mobile and free- to be represented Park, 1966; Mauchline, 1977; Briggs, 1978; swimming stages i.e. copepodids, preadults and adults. Glands Gharagozlou-van Ginnekcn, 1979; Arnaud, Bru- associated with the mouth-tube were also located. Other, non- net, & Mazza, 1988a; Arnaud, Brunet, & Mazza glandular, peroxidase-positive regions were also highlighted by Dumont & Silva-Briano, Unfortu- the stain. These and 1988b; 1997). regions were foundmainly in preadult adult the few detailed studies which stages where they formed characteristic bi-symmetrical nately have been patterns on the cuticle of the dorsal surface. A of some study undertaken serve only to highlight our lack of calanoid copepods suggested that peroxidase-positive glands understanding of these structures, their secretions are a feature peculiar to caligid copepods. This staining technique and their the and has significance to biology ecology proven useful for elucidating the ontogeny of gland of these animals. populations in caligids. the fact that different Despite many types of

have glands been located in various parts of the

Contents copepod body, few of them have had their secre-

tions characterised and their biological significance

Introduction • 9 determined conclusively. Those best studied in- Material and methods 10 clude the glands of the buccal cavity (Boxshall,

DAB 10 1982; Arnaud et ah, 1988ab; Zeni & Zaffagnini, Light microscopy 11 Klein & the lumi- Results 1992; Vaupcl Koomen, 1994) 11

nous glands of the Calanoida (Clarke et ah, 1962; Lepeophtheirus salmonis II & Nauplii 11 Herring, 1988; Bannister Herring, 1989) and the Copcpodid II mucous glands of the capsule-dwelling copepods Chalimus 13 (Hicks & Grahame, 1979). Glandular complexes Preadults 14 have been in the urosome described previously by

Downloaded from Brill.com10/07/2021 11:30:31AM via free access 10 S. Bell et al. - Exocrine glands in Lepeophtheirus and Caligus

(Gharagozlou-van Ginneken, 1979) who suggested Peroxidase enzymes have been reported to be in- they produced chemicals involved in sexual activ- volved in many different functions within animal ity. Fahrenbach (1962) and Boxshall (1982) also tissues, including: free radical neutralisation (Salin

& described glands in the urosomes of copepods but Brown-Peterson, 1993), prostaglandin produc-

tion & did not suggest a function for them. Glands in the (Bowman, Dillwith, Sauer, 1996) and io- thoracic limbs have also been widely reported dine binding as part of protothyroid activity in

(Fahrenbach, 1962; Park, 1966; Dumont & Silva- tunicates (Fredriksson, Ofverholm, & Ericson,

Briano, 1997). 1988).

As of a wider into the func- Other glands arc commonly found in close asso- part investigation tional ofexocrine in salmonis ciation with the integument (‘integumental glands’) significance glands L.

we undertook a locate all the but their role is still the subject of debate. Dennell study to DAB-posi- tive regions in all life-stages of that species and (1947) and Stevenson (1961) suggested that they the related might be involved in cuticle tanning, while Rybakov parasitic copepod Caligus elongatus. determine whether & To are a Dolmatov (1991) suggest a role in cuticle for- DAB-positive regions

widespread feature of the Copepoda, we stained mation. Other functions such as predator deter-

several of and also some rence (Pochon-Masson, Renaud-Mornant, & Cals, species free-living copepod other aquatic arthropods. 1975) immunity to host nematocysts (Briggs, 1978), cuticle anti-fouling (Boxshall, 1982; Bannister,

1993), drag reduction (Bannister, 1993) ‘secretion- Materials and methods trap’ for water-borne chemical cues (Hipeau-

Jacquotte, 1987) and production of pheromones DAB for mate attraction (Gharagozlou-van Ginneken,

1979) have all been suggested as functions of these One lOmg DAB tablet (Sigma D5905) was added glands. It is likely that there is variation in integu- to 15ml of Tris buffer to give a 0.66% w/v solu- mental gland products and functions between spe- tion which to 12 pi of H,0, was added (0.026% cies which probably represent evolutionary changes solution) to provide a substrate for the reaction. necessitated the environment in which by particular of The reaction was halted by the addition 70% the animal lives. ethanol. In contrast to the widely accepted idea that glands Fresh specimens were added to the prepared are actively involved in secretion, Chapman (1981) DAB solution and left to stain at room tempera- showed that the of Neocalanus ‘dermal’glands The ture for a range oftime periods. optimal staining involved in the of dissolved plumchrus were uptake found time was to be: =15min for nauplii and glucose from the sea water. surrounding =6h for chalimus and ~16h for copepodids, pre- Previous work (Bron, 1993; Andrade-Salas, 1997) adults and adults. After these periods had elapsed has shown that the immunohistochemical stain 3’,3- the reaction was halted by carefully pipetting off diaminobenzidine (DAB) high- tetrahydrochloride the stain solution and replacing it with 70% etha-

in L. salmonis larvae and lighted glandular regions nol. adults. DAB is a chromogenic compound used Chalimus, preadult and adult stages were stained routinely in immunohistochemical analyses. It is soon after collection (i.e. not exceeding 48h post- often used coupled to antibodies, as a marker, to removal) to minimise any effects on gland activ- of highlight areas antibody Used on its binding. ity that being removed from the salmon host may it own however in the of reacts, presence hydro- incur. gen peroxide to produce a characteristic (H,0,) The same procedures were followed for naup- brown product which is insoluble in alco- the water, lius, copepodid and adult specimens of caligid hol and This reaction xylene. positive to the DAB copcpod Caligus elongatus Nordmann, 1832. Adults stain generally occurs with endogenous peroxidases of the free-living marine copepods Eurytemora but also with may occur catalase, cytochrome affinis (Poppe, 1880) and Acartia tonsa Dana, 1849, oxidase of and possibly a number other the cladoceran and compounds. Daphnia magna Straus, 1820,

Downloaded from Brill.com10/07/2021 11:30:31AM via free access Contributions to Zoology, 69 (1/2) - 2000 11

larval stages of the anostracan Artemiasalina (L.), Nauplii and the palaemonid Macrobrachium rosenbergii

(de also stained in the for Both the first and second lar- Man) were same way (NI) (Nil) nauplius comparison. vae displayed the same general pattern of DAB-

positive staining, although some slight differences

were evident. In addition to two large dorsal and

Light microscopy ventral ‘median’ glands, both larval stages con-

sistently displayed three pairs of distinct glandu-

Small larval in stages (i.e. nauplius, copepodids and lar regions arranged a bilaterally symmetrical chalimus I and II) were examined in a cavity slide pattern (Fig. 1). These gland populations have been under a cover slip. Lactic acid (87.5%) was used named in accordance with their location within the to aid in the clearing of the body tissues of larval animal. stages, to enhance visualisation of In addition to these late Nil larvae gland structures glands, pos- without affecting the quality of staining, although sessed up to 10 small DAB-positive areas, arranged this method could not be used as a permanent in a symmetrical pattern, in the posterior half of mountant (Huys & Boxshall, 1991). the body between the lateral and posterior glands,

made BH- in 1. Observations were using an Olympus position ‘TIT Fig. These areas corresponded 2 compound microscope and photographs were to the position of glands present in the thoracic

taken limbs of the using an Olympus C-35AD-2 camera. following copepodid stage. The pos-

terior exited via small glands two pores, one lying

close to the ventral side of each balancer. Results During the staining process some individual, un-

hatched of salmonis eggs L. were inadvertently

stained Lepeophtheirus salmonis along with the hatched nauplius stages.

Examination of these showed that eggs pre-hatch DAB was successful in highlighting gland struc- nauplii had the same pattern of DAB-positive gland

tures in all the life-stages ofL. salmonis. In chalimus, distribution as N1 stage larvae. preadult and adult stages there was considerable

variability between specimens of the same sample with regard to their staining characteristics. Pic- Copepodid

tures of gland distribution in these stages were therefore constructed from multiple specimens. The Copepodid larvae displayed a similar pattern of DAB stain also to that of the picked up some other, non-glan- DAB-positive glands preceding dular features. In both L. salmonis and C. elongatus naupliar stages (Fig. 2). The posterior, lateral and

retinula cells of of the cerebrum anterior and the dorsal and ventral median the eye and parts groups often all showed little stained moderately with DAB, as did some glands apparent difference. In ad- of the body musculature, particularly the attach- dition, there were 10 distinct glands in the tho-

ment points of the muscles to the cuticle. In speci- racic limbs, four in the first pair of thoracic legs

mens with it and six in the second damaged was common (thj) (th ), the of integuments n precursors to which could see strong DAB-positive staining around the tears be seen under the ventral cuticle of nd a in the late Nil larvae. Some less distinct cuts cuticle. It is not possible to say smaller, regions

whether 5 such staining is due to DAB-positive com- of staining were also seen i.e. spots around the

ponents in this and 3 the the exposed cuticle or whether rep- eyes spots on free thoracic segment.

msents an active mechanism where DAB-positive The median glands overlay each other, both di-

products are the mouth-tube aggregating at sites of injury. rectly overlying and were seen to

have several ducts associated with them. Although

it was difficult to distinguish the precise relation-

and ship between these glands their ducts, some

Downloaded from Brill.com10/07/2021 11:30:31AM via free access 12 S. Bell et al. - Exocrine glands in Lepeophtheirus and Caligus

Downloaded from Brill.com10/07/2021 11:30:31AM via free access Contributions to Zoology, 69 (1/2) - 2000 13

in relationships could be visualised, as shown Fig. of staining, not apparent in the copepodid stage 3. The ventral median gland was located at the base had developed in the chalimus larvae. A promi- of the labrum and had duct which into of stained a passed nent feature specimens was a complex the labrum exit to on its anterior face approximately of approximately six separate elements lying close one thirdof the down from the end. the cuticle of the dorsal surface way proximal to immediately A second duct to lead from anterior to the This termed the ‘frontal DAB-positive appeared eyes. was one of the median glands and passed in an gland complex’ (FGC). This complex appeared to anterodorsal direction to enter the developing fila- consist of some of the elements of the filament-

The ment. duct stained clearly and extended to the producing glands described by Bron, Sommerville, distal tip of the undeveloped filament. The dorsal Jones & Rae (1991). In particular, the A-glands median gland had at least one duct arising from it described by Bron et al. (1991) stain strongly with which led is the dorsal cuticle immedi- did the three to a pore DAB, as mucous glands which lie ately the in front of the overlying gland. immediately eyes and extend ante-

As the have copepodid larvae a functional gut, riorly towards the A-glands. Other, smaller, asso- m contrast to the naupliar stage, it seems reason- ciated structures also stained but their origin and able to the could re-name the posterior glands of naup- significance not be determined, although it lius the ‘perianal glands’ at the copepodid stage. is possible that they are part of the filament-pro- This fits the terminology of (Boxshall, 1982). ducing complex.

Two pairs of symmetrical glands (or one pair

ofbi-lobed glands) surround the base of the mouth- Chalimus tube on the ventral surface and were termed

‘circum-oral glands’. These glands are evident in All of 4a 4b. It stages chalimus larvae appeared to possess Figs. and is possible that these are the all the ventral DAB-positive glands located in the preced- reaches of the mucous glands of the FGC.

A similar ing larval stages, but during development from pattern was seen in all three later chalimus chalimus I the small to chalimus IV distribution of glands larvae, but with a few, differences. By changes. Chalimus larvae often exhibited variable chalimus 111 a pair of large glands close to the

staining which the bases of under the hindered interpretation of a stage- the maxillae was apparent ven-

specific dis- pattern. The simplest pattern of gland tral cuticle. From each of these glands a fine duct tribution II in was found in chalimus I and larvae: ran a medioventral direction and was presumed these still possessed the perianal, lateral and ante- to exit close to the bases of the maxillules and rior of maxillae. glands the previous stages, although the tho- Although no exit pore could be detected racic limb The in this there evidence that glands had become very indistinct. region, was a pore did anterior lie in and that it glands appeared to be composed of one/ this area was probably obscured two components in chalimus I and chalimus II, as by overlying appendages. This evidence was the

m Fig. 4a whereas it to consist of two/ of considerable of , appeared presence quantities DAB-posi- three tive components in the nauplius stage (see Fig. material of a granular nature adhering to the

!)• This cuticle close the bases of the change may represent either a regression, to maxillules where or stained amalgamation, of the gland units. Other regions the duct was seen to lead. These glands

1- DAB-stained second of L. salmonis and associated ducts the stage nauplius showing glands e.g. posterior gland ducts

(arrowed). Note the connected m median anterior anterior glands are composed of three regions, glands; ag glands; I lateral glands; th thoracic limb Scale bar 250 glands precursor; p posterior glands. pm.

Cg. 2. Dorsal view of DAB-stained _ larva ofL. salmonis. Note granular DAB secretion adhering to cuticle of _. antennules copepodidr x (x). m median anterior 1 lateral th thoracic limb Scale bar 250 glands; ag glands; glands; glands; p perianal glands. pm.

3. Lateral view of anterior cephalothorax of DAB-stained copepodid larva of L. salmonis. Large regions of dispersed stained granules can be within the anterior lateral seen cephalothorax (x). ag glands ; 1 glands; mxp maxillipeds; mt mouth-tube; vm ventral

median Scale bar 100 gland; a antenna; dm dorsal median gland; fd filament duct. pm.

Downloaded from Brill.com10/07/2021 11:30:31AM via free access 14 S. Bell et al. - Exocrine glands in Lepeophtheirus and Caligus

Fig. 4a. Ventral view of DAB-stained second stage chalimus larva of L. salmonis. Ducts are visible leading anteriorly from the

f df 1 anterior fcr first cuticular anterior glands (arrowed), existing filament; developing filament; antennule; ag glands; ridge; eg circum-oral bar 200 glands. Scale pm.

are here termed the ‘post-oral glands’ until their glands per se as no secreted material was seen on homology with previously described glands has the surface of the cuticle, and they shall therefore been determined. The possibilty that these glands be referred to only as ‘dorsal surface regions’ (DSR). correspond to the widely described maxillary glands

found in other species of copepod and attributed with an excretory function, should be considered. Preadults

In some specimens examined there was a large

numberof discrete DAB-positive granules/vesicles The gland distribution pattern of the preadult stages

that of the IV present within the body cavity. These regions are develops from chalimus and showed

shown in Fig. 4b. These regions correspond to the an increase in complexity. There was also an in-

of in the cells the mid-gut epithelium. crease number ofDAB-positive glands from

What were presumed to be the vestiges of the preadult I to preadult II in both sexes.

lateral glands could be discerned in the lateral re- All of the main gland structures located in the

gions of the cephalothorax, roughly level with the chalimus larvae were also found in the preadult

These con- i.e. frontal circum- maxillipeds. glands were very faint, stages perianal, gland complex,

sisting of loosley-packed grains. An anteriorly oral, postoral and dorsal surface regions, although

with leading stained duct was associated each of no evidence of the lateral glands could be found.

these the there glands. In preadult stage were two large groups In chalimus II some small, discrete regions of of glands (or lobes of the same gland) present in

staining first appeared, lying close to the dorsal the anterolateral regions of the cephalothorax. These

cuticle of the both the dorsal cephalothorax. There was an increase groups, which were visible from in their number during each successive chalimus and ventral surfaces, showed an increase in num-

but no definite stage, pattern characteristic of any ber of regions from preadult I to preadult II in both

one could be determined. stage There was no evi- males and females. These gland complexes were

dence to that these of suggest regions staining were given the name ‘anterior gland complexes’ (AGC)

Downloaded from Brill.com10/07/2021 11:30:31AM via free access Contributions to Zoology, 69 (1/2) - 2000 15

brum with two further glands lying immediately

anterolateral to it, within the body cavity. These

glands appeared to connect to the gland at the base

of the labrum.

In common with the chalimus stages, the tho-

racic limbs showed little very staining. However,

in the contrast to chalimus stages, the DSR were

prolific and accounted for the bulk of DAB-posi-

tive staining found in preadults: =85 in preadult I

=140 in II in females, preadult females, =56 pre-

adult I males and =63 in preadult II males. They

distributed in were a bilaterally symmetrical pat- the tern across cephalothorax, and were especially

around the There concentrated margins. were simi-

lar regions of staining on the dorsal and ventral

surfaces of the genital segment. No secreted ma-

terial in association was ever seen with any of these

regions although quantities of DAB-positive ma-

terial were commonly found on the setae of the

caudal rami (particularly in preadult males) and

was presumed to have originated in the perianal

glands.

Adults

The greatest number of DAB-positive areas were

in found adult specimens. The pattern of distribu- Cg. 4b. Ventral view of DAB-stained second chalimus stage larva tion was similar to that of the but with of L. preadults salmonis. ag anterior glands; df developingfilament; c some differences. The bulk of the g circum-oral slight staining glands; fcr first cuticular ridge; v vesicles lining gut. Scale bar 350 could be accounted for the DSR pm. by (Fig. 5).

Adult specimens retained all of the main glan-

011 acc the dular in the ount of the similarity of their location to complexes described preceding stages. glands of the same name in earlier stages. They had well developed perianal glands which The postoral and consisted of one with glands were again prominent large region up to six pairs were situated smaller associated The close to the AGC in a region termed of regions. secretions from the lateral sinus’. When viewed from the ventral these glands were often seen to coat the setae of surface this lateral sinus was located immediately the caudal rami. The circum-oral glands take the lateral to the in maxillules and posterior to the post same form as those seen the preadult stages, as antennary the frontal processes. The posterior margin of this did gland complex. The anterior gland sinus was bounded the ‘first cuticular - were and of by ridge’ complexes large composed many el- a ndge of thickened cuticle extending posterola- ements, and together with the postoral glands fdled teral ly from close to the mouth-tube to the edge of the lateral sinus. The post-oral glands in adult stages the cephalothorax to have the same structural approximately half way along appeared morphology its length. seen in previous stages.

Circum-oral adults glands were present but their ap- in DAB-positive glands become promi- pearance differed from those in the first four ofthoracic in the chalimus stages; nent pairs legs, the setae a laige gland was evident of which were often seen be covered at the base of the la- to in granu-

Downloaded from Brill.com10/07/2021 11:30:31AM via free access - 16 S. Bell et al. Exocrine glands in Lepeophtheirus and Caligus

Fig. 5. Dorsal view of DAB-stained adult female L. salmonis. Fig. 6. Dorsal view of DAB-stained adult female C. elongatus.

Note the DAB-positive material adhering to the marginal Note stained muscle attachment points on dorsal cuticle (x). membrane of the and the of dsr dorsal anterior ccphalothorax (arrowed) regions surface regions; age glandcomplex; fgc frontal

associated attachments bar 500 staining with muscle (x). p perianal gland complex. Scale pm.

anterior glands; dsr dorsal surface regions; age glandcomplex;

fgc frontal glandcomplex. Scale bar 1 mm.

lar stained material. The staining of these glands clear whether this was secreted material, stained

was heavy and consistent between specimens. epibionts adhering to it, or regions of damaged The DSR at this stage, numberedat least 140 in integument.

and females 120 in males. The DSR were the most

prominent feature of the stained animals. There

was an obvious bilateral symmetry to the distribu- Caligus elongatus tion of these areas, the greatest number of which were located around the margins of the ccphal- The nauplius and copepodid larvae of C. elongatus othorax. There appeared to be a sexual dimorphism showed the same pattern of DAB-positive gland with to DSR which distribution found in the of regard distribution, was espe- corresponding stages

in the L. cially pronounced genital segment. salmonis.

The Both adult marginal membrane often had DAB-posi- male and female C. elongatus pos- tivc material sessed all of the main adhering to it, although it was not DAB-positive regions seen

Downloaded from Brill.com10/07/2021 11:30:31AM via free access Contributions to Zoology, 69 (1/2) - 2000 17

in L. salmonis adults. Some differences were noted been reported previously. Bron (1993) first de- i.e. the ‘swellings’ (Boxshall, 1974) on the ven- scribed DAB-positive glands in the thoracic limbs tral cuticle between th, and ,th always stained of L. salmonis copepodids and also located glands

DAB structural differ- which to to the circum-oral heavily with and a slight appear correspond ence of the frontal gland complex was noted. glands seen in chalimus larvae (Bron, Soinmerville,

The Specimens of C. elongatus had considerably more & Rae, 1993). structure of the filament-produc- dorsal L. also surface regions than their L. salmonis coun- ing glands in salmonis chalimus larvae has terparts with adult males having -280 distinct re- been previously investigated (Bron et al. 1991; Bron, gions while females had =160 (Fig. 6). A sexual 1993) and superficially, components of the fron- dimorphism of these regions was evident, at least tal gland complex found in the present study re- in showed of the genital segment. These regions again semble some those glands. Further analysis using a bilateral symmetry in their distribution, with the histological/histochemical techniques will be re- greatest proportion found around the extreme quired to determine the homologies of these glands. margins of the cephalothorax, close to the point The fact that some of the filament-producing glands of marginal membrane attachment. found by (Bron et ah, 1991) stained positively with

stains other than DAB, suggests a multi-component

composition to the gland product and it is likely

Other that of the stained have species many glands may multiple

functional components. Many exocrine secretions

Noneof the other species of aquatic arthropod stain- of copepods have a mucous component (Flicks and ed using the technique employed for L. salmonis Grahame, 1979) and, whilst it has been demonstrat- and C.i ed stain elongatus showed comparable positive stain- that sulphated mucopoly-saccharides may ing. A. and all had with Alcian Blue 1.0 and tonsa, E. affinis D. magna a positively DAB, (pH 2.5) pair of their in the granular DAB-positive regions present in staining failed to demonstrate presence the buccal cavity region. Both A. tonsa and E. affinis exocrine glands of L. salmonis. often be showed a region of staining within the gut, The glands in the thoracic limbs may impor- but whether this represented secretions from the tant with regard to the functioning of the limbs buccal food could in the cavity glands or material not be since they stain strongly only copepodid

determined. in where swimming ability is important and adults The of female E. where be and egg strings affinis were gen- swimming may similarly important the moults the erally only other regions of the body to stain where there are no further to renew with DAB. Specimens ofA. salina, and M. rosen- limbs and their setae. They are conspicuous by

bergii in the chalimus showed no definite staining, although a slight their greatly reduced appearance

of darkening the general body tissues was often stages where limb function is relatively less im-

seen. when filament-attached, and in portant i.e. preadult

stages where there is a relatively short period be-

tween moults. As it was common to find large Discussion quantities of DAB-positive material adhering to

the the secretion limb setae it may be that acts in The DAB stain has useful tool for some to either maintain, or the func- proven as a way improve

highlighting a population of exocrine glands in L. tion of those setae. This supports the suggestions salmonis and C. of researchers who cuticle tan- elongatus.. Whilst the present study previous proposed

has not attempted to determinethe identity of prod- ning (Stevenson, 1961), anti-fouling (Boxshall, ucts giving a positive reaction to DAB, this will 1982; Bannister, 1993) and hydrophobic surfac- be addressed & in future. This work has shown that tants (Brunet, Cuoc, Arnaud, Mazza, 1991) as

DAB-positive both actions of It has been shown reactants are present in spe- possible gland products.

cies of sea lice at all life-stages, including pre- (Gresty & Warren, 1993; Dumontet et al., 1996) hatch nauplii. that copepods frequently have epibiotic organisms Several of the glands found in this study have attached to their cuticle and it is possible, there-

Downloaded from Brill.com10/07/2021 11:30:31AM via free access 18 S. Bell et al. - Exocrine glands in Lepeophtheirus and Caligus

in- includes chemicals released fore, that secretions from the glands may be at sites of tissue in- volved in killing or preventing the establishment jury. The peroxidase compounds found in sea lice of such organisms on the cuticle surface, a phe- may be involved in protecting the parasite from

nomenon found in corixid beetles where gland the deleterious effects of such compounds. Pros- secretions bacterial the hairs of which in in- prevent growth on taglandins, are synthesized a process the & This have been shown to physical gill (Kovac Maschwitz, 1991). volving peroxidase enzymes, hypothesis does not account for why theseregions be involved in host inflammatory response modu-

be unless the envi- lation. Peroxidases and have been appear to unique to caligids, prostaglandins

in in tick ronment which they live is more likely to ex- isolated species of with unusually long host- pose them to such fouling organisms. attachment periods (Bowman et al., 1996). It seems

Peroxidases have been shown to be present in reasonable to suppose that sea lice species may red blood central have methods maintain benevolent cells, hepatocytes, nervous sys- developed to a

environment tem neurons and oestrogen-sensitive tissues (Rain- for themselves and this should be bow 1996) and are also known to be fundamental considered as a possible function of these glands.

As few to the process of prostaglandin production from very DAB-positive regions were present arachidonic acid (Bowman et al., 1996). Peroxi- in free-living species, as compared to the two dase and catalase have also been shown examined stain enzymes parasitic species here, the DAB

be neutralisers of be features which to important potentially damag- may picking out represent spe- ing free radicals, which result following tissue cific developments associated with the parasitic damage and are formed as by-products of metabo- mode of life.

lism and also from the interaction of U.V with It is probable that the glands highlighted by this natural organic matter. Peroxidase activity has also stain do not all have the same function. In most been show in tunicates where it is believed to be cases the stain highlighted secreted material and involved in iodination fundamental to that it is ultrastructural processes we can presume not gland protothyroid activity (Fredriksson et al., 1988). elements which arc being stained, but rather the

The only DAB-positive glandular regions seen secretion itself which shows the positive reaction. in free-living species are in the buccal cavity area. This is not however, true for the DSR where no

Whilst the function of these glands is unknown it associated ducts or secreted material have been may be suggested that peroxidases are being pro- observed. The possibility that the stain here is duced here to assist in the elimination of free radi- highlighting ultrastructural or neurological features cals produced as a side-effect of feeding activity, will have to be addressed using other staining tech- a phenomenon witnessed in caterpillars (Felton & niques and TEM. The results of a preliminary SEM

If DSR associ- Duffey, 1991). such products are not quickly examination suggest that the may be neutralised they might otherwise damage the gut ated with cuticular sensillae, adding further doubt epithelium. Glands in the buccal cavity region of to the idea that they are exocrine glands.

marine been well docu- the free-living copepods have In summary, DAB-positive gland distribu-

tion mented (Boxshall, 1982; Arnaud et al., I988ab; of L. salmonis had a consistent pattern which

Vaupel Klein & Koomen, 1994) and L. salmonis could be specifically identified at each life-stage, and C. elongatus also have DAB-positive glands with the exception ofchalimus larvae where staining associated with their hindered mouth-parts which may func- difficulties interpretation. The develop- tion in a similar it that of the fundamental could be manner, although appears ment gland groups at least one of them exits via the outside followed several i.e. a pore on through life-stages perianal, of the mouth-tube and it is therefore unclear how lateral and anterior from nauplius to chalimus, while the secretion would interact with the food mate- the perianal glands (and possibly the anterior gland rial. complexes) were retained from nauplius to the adult.

It is that sea lice will be The median could be followed from possible exposed to glands naup- free-radicals, both as a result of host tissue dam- lius and be to copepodid may possibly represented and host immune age, following a response which by the circum-oral and frontal gland complexes in

Downloaded from Brill.com10/07/2021 11:30:31AM via free access 19 Contributions to Zoology, 69 (1/2) - 2000

later that these have and Fanned Fish: Sea Lice. Chichester: Ellis Horwood, stages. We can presume glands 125-142 a fundamental role to play in the biology of these Brunet M, Cuoc C, Arnaud J, Mazza J. 1991. Tegumental species. Histological evidence is being sought to glands in a copepod Hemidiaptomus ingens: structural, determine the characteristics of the glands physical ultrastructural and cytochemical aspects. Tissue and Cell

and their relationship with other organs and tis- 23: 733-743.

demonstration of sues in the body. Bussolati G. 1971. Histochemical sulfate of diaminobenzidine. Hislochem. J. 3: Bron have groups by means (1993) and Bron et al. (1993) pro- 445-449. vided evidence of the existence of further glands Evidence for dissolved Chapman PM. 1981. glucose up- in L. salmonis which are not DAB-positive sug- take from seawater by Neocalanus plumchrus (Arthro- DAB does of the gesting the stain not highlight all poda, Copepoda). Can. J. Zool. 59: 1618-1621.

glands Clarke Conover David Nicol JAC. 1962. present in this species. GL, RJ, CN, Comparative studies of luminescence in copepods and

other pelagic marine animals. J. Mar. Biol. Assoc. U.K.

42: 541-564. References Den noil R. 1947. The occurrence and significance of phe-

nolic hardening in the newly formed cuticle of Crusta-

Andrade-Salas and the ecto- O. 1997. Feeding digestion in cea. Proc. Roy. Soc. B 134: 485-503.

parasilic copepod Lepeophtheirus salmonis (Kroyer, 1837). Dumont HJ, Silva-Briano M. 1997. Sensory and glandular

Unpublished doctoral dissertation. University of Stirling. equipmentof the trunk limbs ofthe Chydoridae and Macro- Arnaud J, Brunet M, Mazza J. 1988a. Labral glands in thricidae (Crustacea: Anomopoda). Hydrobiologia 360:

Centropages typicus (Copepoda, Calanoida). I. Sites of 33-46.

synthesis. J. Morphol. 197: 21-32. Dumontct S, Krovacck K, Baloda SB, Grottoli R, Pasqualc

Arnaud labral J, Brunet M, Maz/.a J. 19881). The glands V, Vanucei S. 1996. Ecological relationship between

in Calanoida). II. Sites Aeromonas and Vibrio and in Centropages typicus (Copepoda, spp planktonic copepods

of secretory release. J. Morphol. 197: 209-219. the coastal marine environment in southern Italy. Comp.

Bannister NJ. 1993. Distribution and structure of sub-cu- Immunol. Microbiol. Infect. Dis. 19: 245-254.

ticular J. Mar. glands in the copepod Temora longicornis. Fahrenbach WH. 1962. The biology of a harpacticoid cope- Biol. Assoc. U.K. 73: 97-107. pod. La Cellule 62: 303-376. Bannister NJ, Herring PJ. 1989. Distribution and structure Felton GW, Duffey SS. 1991. Protective action of midgut

ol luminous cells in four marine copepods. J. Mar. Biol. catalase in lepidopteran larvae against oxidative plant

Assoc. U.K. 69: 523-533. defenses. J. Chem. Ecol. 17: 1715-1732. Bowman AS, Dillwith .IW, Sauer JR. 1996. Tick salivary Frcdriksson G, Ofverholm T, Ericson LE. 1988. Iodine

and Para- in the of prostaglandins: presence, origin significance. binding and peroxidase activity endostyle Salpa

sitology 12: 388-396. Dolioletta -<*, Today, fusiformis, Thalia democratica, gegenbauriand Boxshall GA. 1974. Lepeopththeirus pectoralis (O.F. Doliolum nationalis (Tunicata, Thaliacea). Cell and Tis- M iiller, and 1776): a description, review some compari- sue Res. 253: 403-411.

sons with Muller. J. Nat. Hist. 8: 445- Ginneken ID. 1979. Etude the genus Caligus Gharagozlou-van ultrastructurale

468. des et cytochimique de 1’activite temporaire glandes tegu-

Boxshall GA. 1982. the Ann. Sci. Nat. Zool. On the anatomy of misophrioid raentaires d’un Crustace Copepode.

copepods, with special reference to Bentomisophriapalliata Paris 13: 205-212.

Sars. Phil. Trans. Roy. Soc. London. B 297: 125-181. Gresty KA, Warren A. 1993. Incidence of ciliate epibionts KrigJ-s Rl>. 1978. Paranthessius from salmon in Structure ofthe integument in on Lepeophtheirus salmonis Japan and

anemoniae Claus, a of the Snakelocks copcpod associate Scotland; a scanning electron microscopic study. Boxshall anemone Ammonia sulcata (Pennant). J. Morphol. 156: GA, Defaye D, eds. Pathogens of Wild and Fanned Fish: 293-316. Sea Lice. Chichester: Ellis Horwood, 356-363.

Bron JE. 1993. A study of the biology .and behaviour of the Herring PJ. 1988. Copepod luminescence. In; Boxshall GA,

copepodid larvae of the salmon louse Lepeophtheirus Schminke HK, eds. Biology of Copepods. Hydrobiologia salmonis (Kroyer, 1837) (Copepoda: Caligidae). Unpub- 167/168: 183-195 lished doctoral dissertation. University of Stirling. Hicks GRF, Grahame J. 1979. Mucus production and its Bron JE, Sommerville C, Jones M, Rae GH. 1991. The role in the feedingbehaviour of Diarthrodes nobilis (Cope-

settlement and attachment of early stages of the salmon poda: Harpacticoida). J. Mar. Biol. Assoc. U.K. 50: 321-

louse, Lepeophtheirus salmonis (Copepoda: Caligidae) on 330. the salmon host, Salmo I 224: 201-212. R. 1987. Ultrastructure and salar.„ 7nnl, presumed , ... Hipcau-Jacquotte Bron JE, Som merville C, Rae GH. 1993. Aspects of the function of the pleural dermal glands in the atypical male

behaviour of copepodid larvae ofthe salmon louse (Kroyer, of the parasitic copepod Pachypygus gibber (Crustacea: '837). Boxshall GA, Defaye D, eds. Pathogens of Wild Notodelphyidae). J. Crust. Biol. 7: 60-70.

Downloaded from Brill.com10/07/2021 11:30:31AM via free access 20 S. Bell et al. - Exocrine glands in Lepeophtheirus and Caligus

lluys R, Boxshall GA. 1991. Copepod Evolution. London parasitic copepod Cucumaricola curvatus (Copepoda,

Ray Society. Cucumaricolidae). Zoologicheskiy Zhurnal 70: 44-54.

Kovac D, Maschwitz U. 1991. The function ofthe metatho- Salin ML, Brown-Peterson NJ. 1993. Dealing with active

racic scent in corixid intermediates: A gland bugs (Hemiptera, Corixidae): oxygen halophilic perspective. Experientia

Secretion grooming on the water surface. J. Nat. Hist. 49: 523-529.

25: 331-340. Stevenson JR. 1961. Polyphenol oxidase in the tegumental

Maiiclilinc J. 1977. The integumental sensilla ami glands of glands in relation to the molting cycle of the isopod crus-

pelagic Crustacea. J. Mar. Biol. Assoc. U.K. 57: 973-994. tacean Armadillidium vulgare. Biol. Bull. (Woods Hole)

Park TS. 1966. The biology of a calanoid copepod, Epilabido- 121: 554-560.

cera amphitrites McMurrish. La Cellule 66: 129-251. Vaupel Klein JC van, Koomcn P. 1994. The possible ori-

Pochon-Masson Kenaud-IVIornant Cals P. 1975. Con- of used for in J, J, gins mucus jets immobilizing prey species

tribution a la connaissance des glandes tegumentaires ofEuchirella (Copepoda, Calanoida, Aetideidae), 1. Theo-

metameriques d’un crustace meiobenthique interstitiel retical considerations in relation to swimming and feed-

(Crustacea: Mystacocarida). Cytologic structurale et infra- ing behaviour. Crustaceana 66: 184-203.

structurale. Arch. Zool. Exp. Gen. 116; 123-146. Yonge CM. 1932. On the nature and permeability of chitin.

Rainbow RD. 1996. Immunohistochemistry. Woods AE, Ellis 1. The chitin lining of the foregut of decapod Crustacea

RC, eds. Laboratory Histopathology. A Complete Refer- and the function of the tegumental glands. Proc. Roy.

ence (Second ed.). Churchill Livingstone. Soc. B 111: 298-329.

et Richards J. 1891. Recherches sur le systeme glandulaire Zeni C, Zal'fagnini F. 1992. Labral glands of Leptestheira

sur le systeme nerveux des Copepodeslibres d’eau douce dahalacensis (Branchiopoda: Spinicaudata): An ultrastruc-

des de vivent tural J. Crust. Biol. 12: 661-676. suivies d’une revision especes ce groupe qui study.

en France. Ann. Sci. Nat. B 12: 113-260.

Rybakov AV, Dolmatov IY. 1991. Internal structure of the Received: 4 December 1998

Downloaded from Brill.com10/07/2021 11:30:31AM via free access