Ultrastructural biomarkers as tools to characterize the health status of fish in contaminated streams

Ultrastructural biomarkers in gill, liver, andkidney of brown trout (Salmo truttaf. fario) and stone loach (Barbatulabarbatula) were investigated over afive year period. The cellular damage of theorgans was assessed semi-quantitatively basedon a three-step classification ofultrastructural responses. Data obtained forfish exposed under semi-field conditions in twodifferently polluted test streams and in thelaboratory demonstrated that the ultrastructureof the organs can be correlated with differentpollutant exposure conditions. Cellular damagewas generally more severe in fish exposed to acomplexly polluted stream than in those exposedto a moderately polluted stream or to tap waterin the laboratory. Histopathological effects inliver and gill of trout were more pronounced inferal fish than in transplanted fish, whereasresponses in the kidney in both species, and inliver and gill of loach, were similar forintroduced and feral fish. In a laboratoryexperiment where trout were exposed todifferent mixtures of pollutants includingpesticides, PAH, and ammonia, only theultrastructure of kidney and liver showedsignificant differences between the threeexperimental set-ups. In a recovery experiment,where trout were transferred from thesemi-field condition back to the laboratory,ultrastructural investigations showed adifferential capacity of the respective organsto recover from stress under field conditions.Kidney and liver fully recovered after threemonths under control condition whereas gillsdemonstrated only partial recovery.     

Histopathological alterations and parasite infection in fish: indicators of multiple stress factors

Within the scope of the multidisciplinaryresearch project Valimar (1995–1999), thepresent study emphasizes the use ofhistopathological investigations to evaluatethe effects of the different chemical impactof two small streams on the general health andcondition of the two sentinel fish species,brown trout (Salmo trutta f.fario) and loach (Barbatula barbatula).Parasitological investigations were alsoincluded to assess a possible relationshipbetween toxicant exposure, parasitic load, andthe occurence of histopathological organchanges. According to the multitiered approachof the Valimar project, fish from the field,individuals exposed to stream water undersemi-field conditions, and fish fromlaboratory experiments were investigated.Under semi-field conditions, thehistopathological responses in brown troutproved to reflect the different levels ofcontamination of the two small streams moreclearly than those detected in loach. Whencaptured from the field, both fish speciesrevealed, in most cases, a higher degree ofhistopathological alterations than thoseindividuals exposed at the respective siteunder semi-field conditions. Simulation of thetoxic load of the more polluted stream in alaboratory experiment did not result in thesame kind of tissue lesions in brown trout asin the field and semi-field investigations.Whereas results from the laboratoryexperiments could be specified astoxicant-induced, the histopathologicalalterations in fish from both semi-field andfield studies represent stress responses whichintegrate the effects of various abiotic andbiotic stress factors including both toxicantsand parasitic diseases.     

Mast cell responses to Ergasilus (Copepoda), a gill ectoparasite of sea bream

Immunocytochemical, light microscopy and ultrastructural studies were conducted on gill of sea bream, Sparus aurata L., naturally parasitized with the important parasitic copepod Ergasilus sp. to assess pathology and cellular responses. Thirty-seven S. aurata were examined from a fish farm; 26 (70%) were parasitized, with infection intensity ranging from 3 to 55 parasites per fish. Hosts were divided into two groups, lightly infected fish (<15 parasites per fish) and heavily infected fish (>15 parasites per fish). In histological sections, the copepod encircled gill lamellae with its second antennae, compressed the epithelium, provoked hyperplasia and hemorrhage, occluded arteries and often caused lamellar disruption. Fusion of the secondary lamellae due to epithelial hyperplasia was common in all infected fish; heavily infected fish showed more intense branchial inflammation. In both healthy and infected fish, mast cells (MCs) were free within the connective tissue inside and outside the blood vessels of the primary lamellae and made close contact with vascular endothelial cells, mucous cells and rodlet cells (RCs). MCs were irregular in shape with a cytoplasm filled by numerous electron-dense, membrane-bound granules. Immunostaining of primary and secondary gill filaments with an antibody against the antimicrobial peptide (AMP) piscidin 3 (anti-piscidin 3 antibody, anti-HAGR) revealed a subpopulation of MCs that were positive. These MCs were more abundant in gills of heavily infected fish than in either lightly infected or uninfected fish (ANOVA, P<0.05). Our report documents the response of gill to ectoparasite infection and provides further evidence that mast cells and their AMPs may play a role in responding to branchial ectoparasite infections.     

Ambient salinity modulates the expression of sodium pumps in branchial mitochondria-rich cells of Mozambique tilapia,Oreochromis mossambicus

Na,K-ATPase (sodium pumps) provide the primitive driving force for ion transport in branchial epithelial cells. Immunoblots of epithelial homogenates of both seawater (SW)- and freshwater (FW)-adapted tilapia gills as well as rat brain homogenate, a positive control, revealed one major band with a molecular weight of about 100 kDa. SW-adapted tilapia gills possessed larger (about 2-fold) amounts of sodium pumps compared with FW-adapted tilapia gills. (3)H-ouabain binding representing functional binding sites of Na,K-ATPase was also higher (about 3.5-fold) in gills of SW-adapted tilapia compared to that of FW-adapted fish. Moreover, specific activities of SW fish were higher (about 2-fold) than those of FW fish. Double labeling of Na,K-ATPase and Con-A, a fluorescent marker of MR cells, in tilapia gills followed by analysis with confocal microscopy showed that sodium pumps were localized mainly in MR cells, including the SW type and different FW types. Although more-active expression of Na,K-ATPase was demonstrated in gills of SW-adapted tilapia, no significant differences in densities of apical openings of MR cells were found between SW- and FW-adapted fish. These results indicate that, during salinity challenge, tilapia develop more "functional" Na,K-ATPase in SW-type MR cells to meet physiological demands.     

Changes to chloride and rodlet cells in gills, kidney and intestine of Dicentrarchus labrax (L.) exposed to reduced salinities

The effect of osmotic shock was investigated mainly in the chloride cells (CCs) and rodlet cells (RCs) of gills, and RCs of intestine and kidney of the European sea bass Dicentrarchus labrax obtained from a farm in the northern Adriatic Sea. During the experiment, fish were abruptly transferred from sea water to a salinity of 15 (15 SW) or to fresh water (FW). Numeric variation and ultrastructural changes of both cell types were evaluated at 24, 48 and 96 h after the transfer to lower salinity levels, using light and transmission electron microscopy (TEM). Exposure to FW produced a significant increase ( P < 0·05) in the number of branchial CCs and RCs within 96 and 24 h, respectively. Following osmotic challenge (either transfer to 15 SW or FW), kidney and intestine showed an evident increase in RC numbers. The cellular damage detected by TEM was the same for each sampling time (24, 48 and 96 h), but appeared more severe in fish exposed to FW (higher osmotic shock) than in those exposed to 15 SW. In RCs cytoplasmic vacuolizations, autophagosomes and autophagolysosomes with myelinoid bodies, dissolution and shrinkage of the typical inclusions were documented. Nevertheless, CCs showed vacuolization of endoplasmic reticulum and cytoplasmic dissolution and maintained the apical crypt typical of seawater acclimated fish. Renal tubular cells and intestinal epithelial cells showed similar changes to those reported for CCs and RCs.     

Haliotrema abaddon n. sp. (Monogenoidea: Dactylogyridae) from the gills of wild and maricultured West Australian dhufish Glaucosoma hebraicum (teleostei: Glaucosomatidae), in Australia

Haliotrema abaddon n. sp. (Dactylogyridae) is described from the gills of West Australian dhufish Glaucosoma hebraicum (Teleostei: Glaucosomatidae), from the Indian Ocean near Fremantle, Western Australia. The parasite is recorded from cultured dhufish and from a major portion of the known geographic distribution of its host. In cultured dhufish, H. abaddon causes branchitis and hyperplasia and metaplasia of the lamellar epithelial cells and may be a limiting factor in the development of an economically feasible mariculture industry. Comparable prevalences and densities of the parasite on wild and cultured dhufish were observed. In heavily infested captive fish, clusters of eggs (egg strands) appear to restrict blood flow to localized regions of the gills as a result of entanglement of egg filaments with secondary gill lamellae. Hatching of oncomiracidia occurs in 7-10 days at 20-25 C.     

Mucous cell responses in gill and skin of brown trout Salmo trutta fario in acidic, aluminium-containing stream water

Morphometric examination was carried out on the gills and skin of wild and caged hatchery brown trout Salmo trutta fario in an acidic (pH 4.9 to 5.4; Al 203 to 250 microg l(-1)) and in a non-acidic (pH 6.7 to 7.0; Al 27 to 67 microg l(-1)) stream in the Vosges Mountains (NE France) to assess the sublethal effects of acidic water on the mucous cell response. The caged fish were randomly collected after 2, 4, 7 and 11 d and the wild fish were obtained by electrofishing. After 2 d, a reduction of both mucous cell (MC) number and size was observed in the gills of fish held in the acidic stream, suggesting a massive mucus discharge. Hyperplasia and hypertrophy of cells immediately followed this mucus secretion. In the same fish population, skin examination showed a slight and delayed decrease of MC number but a significant increase of cell size. The number of mucous cells of gills and skin was similar in both wild trout populations, whereas a significant MC hypertrophy was observed in the wild fish of the acidic stream. The present field experiment indicates that caged fish could be useful as early indicators of acidification. In addition, the examination of wild populations suggested the occurrence of adaptive mechanisms, information that might be of importance in the context of river recovery programs.     

Macrozoobenthos communities and biocoenotic assessment of ecological status in relation to degree of human impact in small streams in southwest Germany

One of the major objectives of the VALIMAR project is to determine the ecological significance of various fish biomarker studies as indicators of chronic pollution in small streams in southwest Germany. Results of these fish biomarker investigations were compared to information from complementary studies on the meiobenthos, macrobenthos, and fish community studies to assess the ecological significance of these biomarker investigations. The main objective of this study was to provide biological assessments of the biomarker sites on the basis of the macrozoobenthos communities. Since no validated framework for the assessment of the biological integrity existed in Germany, two multimetric approaches were adapted to the whole stream system by investigating benthos and fish communities of 46 sites of varying degrees of human disturbance. Assessment of the communities was conducted in accordance with the European Community Water Framework Directive. Species distribution of benthos depended upon stream type and pollution status of streams. Biological attributes and bioindices of benthos communities, however, did not correlate with typological parameters like stream size or dominant substrate but correlated better with pollution parameters like conductivity or chloride concentration. Using a set of 18 measures, such as portion of sessile individuals, Rheoindex, oxygen availability index, and portion of pool dwellers, the benthos communities were characterized and evaluated. The composition of the fish communities was mainly determined by stream type, pollution and migration barriers. The influence of chemical parameters could be assessed by developing a“fish chemistry index”, which calculatesthe similarity of the present fish community with the potential natural community, but excludes those species strongly effected by deficits in stream channel morphology. Both fish chemistry index and benthic indices strongly correlated with pollution index parameters, clearly distinguishing between the more polluted Körsch sites and the less pollutedKrähenbach and Aich sites. Most of the single bioindices as well as overall assessment by multimetric indices indicated a gradient of decreasing quality from the reference stream Krähenbach to theslightly polluted Aich and Körsch upstream site (KE, upstream of all sewage treatment plants) and finally to the most polluted Körsch site directlybelow the most upstream sewage treatment plant (KD). According to the Water Framework Directive, the classification of ecological status of the benthos communities ranges from “high” (best of 5 classes) forthe reference stream to “bad” (5th class) for KD. Assessment of the fish community tends to score somewhat worse than the benthos due to deficits in morphological quality of the stream reaches. The benthos assessment and the newlydeveloped “fish chemistry index” correlated well with chemical water quality and hence with biomarkers, whereas ecological status of fish and overall ecological status was also influenced by river morphology. In conclusion all tested assessment methods on biocoenotic level are reliable indicators for the degree of human disturbance on small streams, whereas biomarkers are more suited for risk assessment and the investigation ofcause-effect-relationships.     

瓦氏雅罗鱼生殖洄游过程中离子调节相关生理变化研究

为了解达里湖瓦氏雅罗鱼(Leuciscus waleckii)生殖洄游过程中血清离子调节相关生理变化, 对比了达里湖和贡格尔河瓦氏雅罗鱼血清离子(Na+、K+、Cl?、Ca2+和Mg2+)水平, 鳃、肠和肾组织Na+/K+-ATPase和鳃Ca2+/Mg2+-ATPase活性、血清催乳素(PRL)、生长激素(GH)和类胰岛素生长因子-1(IGF-1)水平及鳃组织结构差异; 并利用实验生态学方法, 研究达里湖中瓦氏雅罗鱼转入贡格尔河水24h后上述生理参数的响应。研究结果显示, 与达里湖未洄游的瓦氏雅罗鱼相比, 洄游到贡格尔河后其血清Na+含量显著降低(P<0.05), Cl?含量显著升高(P<0.05), 肾脏和肠组织中Na+/K+-ATPase活性显著升高(P<0.05), 而鳃组织中Na+/K+-ATPase活性无显著变化; 血清K+、Ca2+、Mg2+水平和GH、IGF-1、PRL含量无显著变化。将达里湖瓦氏雅罗鱼转入河水中24h后, 其血清Cl?含量显著升高(P<0.05)、K+含量显著降低(P<0.05), 且在鳃、肠和肾组织中Na+/K+-ATPase及鳃Ca2+/Mg2+-ATPase活性均显著升高(P<0.05), 血清PRL和IGF-1水平显著升高(P<0.05); 比较湖中和河中瓦氏雅罗鱼鳃组织形态结构, 显示湖中瓦氏雅罗鱼鳃基底膜分布着大量黏液细胞, 洄游到河水中后黏液细胞数量明显减少, 鳃基底膜上氯细胞体积增大而数量未见明显变化。本研究结果表明: 瓦氏雅罗鱼从达里湖洄游到贡格尔河后通过提高血清PRL和IGF-1水平, 进而介导鳃、肠和肾组织中Na+/K+-ATPase活性增加, 从而维持鱼体较高或稳定的血清离子水平。     

Transfection efficiency and cytotoxicity of cationic liposomes in primary cultures of rainbow trout (Oncorhynchus mykiss) gill cells

Immunisation of fish by immersion has been applied for inactivated, whole cell bacterins, where the gill epithelial cells are considered as one of the prime uptake sites. Antigen entry is a critical factor for delivery of vaccine antigens through the immersion route, also for DNA vaccines, and delivery systems like cationic liposomes may enhance uptake. In this study, the aim was to examine the efficiency of cationic liposomes as a means to transfect primary cultures of rainbow trout gill cells with plasmids encoding viral or reporter proteins. Furthermore, the effects of the concentration and composition of liposomes/lipoplex on the viability of the cells were evaluated. Transfection of the gill cells was possible with both plasmids following transfection with lipoplexes of a neutral charge. Low concentrations and neutral/negatively charged formulations were favourable with respect to the toxicity of the formulations. Given that the mucous barrier covering the gills is overcome, this system might be useful for the priming of the local immunity in the fish gills.     

Goussia girellae n. sp. (Apicomplexa: Eimeriorina) in the opaleye, Girella nigricans

Goussia girellae n. sp. is described from the opaleye fish, Girella nigricans. Merogonic stages were observed in the apices of intestinal epithelial cells, in the lamina propria, and in extra-intestinal sites including liver, gills, and spleen. Gamonts were observed in the intestinal epithelial cells. Only unsporulated oocysts were detected in the intestine, and sporulation occurred when feces containing oocysts were incubated for 48 h in seawater at 21 degrees C. Oocysts are elongated (24.8 x 14.7 micron) with a wall about 200 nm thick and have no residuum, micropyle, or polar granule. Sporocysts are ellipsoid (8.5 x 4.5 micron), have a thin two-layered wall approximately 30 nm thick, and consist of two valves joined by a suture. Although moribund opaleye were also infected with Gyrodactylus sp., Cryptobia sp., Cardicola sp., and epitheliocystis organisms (chlamydia), all fish were heavily infected with G. girellae and morbidity was thus attributed to the coccidium.     

Heavy metal pollution in Antarctica: a molecular ecotoxicological approach to exposure assessment

Trematomus bernacchii sampled from two sites at Ross Island, Antarctica, were assessed for condition (gonadal and hepatic somatic indices and condition factor), pathological state (liver and gill histology), hepatic metal (Zn, Cu, Cd and Ni) concentrations, and metallothionein induction by quantitative competitive (qc) RT-PCR. Fish from a polluted site (Winter Quarters Bay) had pathological anomalies including necrosis and periductal inflammation in their livers, and X-cell disease, epithelial hyperplasia, lamellar fusion and aneurysms in their gills. Such anomalies were less common in fish from a relatively pristine site (Backdoor Bay, Cape Royds). Fish from both sites had similar liver concentrations of Zn, Cu and Cd and qcRT-PCR revealed similar levels of hepatic metallothionein mRNA. Ni in the livers of fish from Winter Quarters Bay were higher than those in fish from Backdoor Bay, but the differences were not great enough to affect hepatic metallothionein mRNA significantly. Despite the polluted state of Winter Quarters Bay waters, it seems that the heavy metals present may have only limited impact on the health status of fish collected from this locality. This may reflect a low bioavailability of the heavy metals in Ross Island marine sediments and suggests that other factors, such as relatively high levels of organic pollutants (PAHs, PCBs) or pathogens from the nearby sewage outlet, may play a more significant role in the aetiology of pathological conditions in fish from Winter Quarters Bay.     

The application of chemical and biological multi-metric models to a small urban stream for ecological health assessments

Nutrient or/and organic pollutions have been used as one of the key factors influencing the stream health, so research approaches were frequently focused on chemical analysis rather and biological analysis. Our objective of the present study was to diagnose the chemical and biological stream health using chemical multi-metric model of nutrient pollution index (NPI) and biological multi-metric fish model of the index of biotic integrity (IBI), respectively in an urban stream. Seven years dataset from 2008 to 2014 were used for the health assessments. The nutrients regime (TN, TP), sestonic chlorophyll, total suspended solids (TSS), electrical conductivity, BOD, and COD showed a typical polluted stream with large temporal and spatial fluctuations due to variations of Asian monsoon rain. The analysis of fish trophic guilds and fish tolerance guilds showed that omnivore fish species and tolerant species were dominated the community and the proportions were directly determined by water chemistry (nutrient and organic matters). Chemical model of NPI showed “poor-very poor health condition” by the criteria of the model and showed same results with the biological model of fish IBI. The degradation of the stream health was mainly due to massive effluents from the wastewater treatment plants. Overall, our data suggest that the multi-metric chemical and biological models may be used as a key tool diagnosing the stream health condition in an urban stream.     

Does sulphide detoxication occur in the gills of the hydrothermal vent shrimp, Rimicaris exoculata?Y a-t-il détoxication des sulfures dans les branchies de la crevette hydrothermale, Rimicaris exoculata ?

Ultrastructural observations of the gills of the hydrothermal vent shrimp Rimicaris exoculata reveal that the epithelial cells contain numerous mitochondria clustered around unusual organelles (diameter of 0.7 to 2.5 microns) containing membrane stacks. These organelles were termed sulphide-oxidising bodies (SOBs) by structural analogy with organelles observed in the tissues of species adapted to sulphide-rich environments. Moreover, in the gills of R. exoculata, mitochondria display numerous electron-dense granules in their stroma. Such ultrastructural features suggest that sulphide detoxication may probably occur in the gills of R. exoculata. Comparable structures were also described in the gills of other hydrothermal vent species, as the alvinellid Pompeii worms that, as R. exoculata, are housing ectosymbiotic bacteria.     

Brown trout Salmo trutta f. Fario liver ultrastructure as a biomarker for assessment of small stream pollution

T he effects of environmental pollutants from two small streams in south-west Germany on the liver of brown trout (S almo trutta f. Fario ) were studied as biomarkers by means of quantitative and semi-quantitative electron microscopy, and quantitatively by morphometrical measurements. Cellular damage was assessed semi-quantitatively based on a classification of ultrastructural responses. Both methods revealed more severe cellular effects in the liver of trout which had been exposed to the highly polluted stream than in those exposed to the lightly polluted river. Morphometrical studies showed a significant reduction of glycogen storage and a significant increase in number of mitochondria, peroxisomes and cisternae of the endoplasmic reticulum. The biomarker responses of this study were correlated with the results obtained by limnological and analytical investigations, and reflect the levels of pollution in each stream.     

Various responses to copper and manganese exposure of Carassius auratus gibelio from two populations

The aim of this study was to compare the effect of pro-oxidants copper (Cu(2+), 0.005 and 0.050mg L(-1)) or manganese (Mn(2+), 0.17 and 1.7mg L(-1)) on Carassius auratus gibelio from polluted (B) and unpolluted (Z) sites after exposure for fourteen days. Fish from site B showed high levels of lipid peroxidation (TBARS concentration), lower levels of metallothionein (MT)-related metal, total glutathione (GSH), its redox index, superoxide dismutase (Cu,Zn- and Mn-SOD), glutathione-S-transferase (GST) and cholinesterase (ChE) activities and also higher MT-related thiol concentration in the liver and gills. A common effect of exposure was related to genotoxicity, a decrease in GSH and an increase in microsomal ethoxyresorufin O-deethylase activity in the liver. However, the systems of oxidative stress and biotransformation were more efficient in fish from the polluted site, while the responsivity of MTs in this fish was impaired. Principle Component Analysis separated the subgroups from the unpolluted site and fish loaded by lesser concentrations of metals on the one side, and fish from the polluted site jointly with fish exposed to higher concentrations of metals on the other side. The main distinguishing indices of sites and exposures selected by classification and regression tree (CART) analysis were MT characteristics and genotoxicity.     

Atlantic salmon paramyxovirus (ASPV) infection contributes to proliferative gill inflammation (PGI) in seawater-reared Salmo salar

Proliferative gill inflammation (PGI) causes significant losses in farmed Atlantic salmon Salmo salar L. in Norway, especially during the first months following seawater transfer. The aetiology is apparently multifactorial, including infection with chlamydia-like bacteria and Atlantic salmon paramyxovirus (ASPV). In the present study, gills from diseased fish from 3 farms on the western coast of Norway were sampled. The pathological changes were briefly described and the aetiological significance of ASPV studied by immunofluorescent staining of cryosections and by immunohistochemistry on sections of formalin-fixed and paraffin-embedded tissue. The pathological changes were macroscopically characterized by palour of the gills, and histologically by inflammation, circulatory disturbances, cell death and epithelial cell proliferation. ASPV was demonstrated in fish from all farms studied, as immunostaining consistent with ASPV was obtained in lamellar epithelial and endothelial cells of pathologically altered tissues. It is concluded that ASPV is at least a contributing cause of PGI. As far as we know, this is the first demonstration of fish disease related to infection with a paramyxovirus.     

THE STRUCTURE OF FISH GILLS IN RELATION TO THEIR RESPIRATORY FUNCTION

1. The general structure of the gills of different fishes is compared and it is concluded that, though essentially the same, there are certain differences by which they can be recognized. Possible ways in which they may have evolved from one another are considered. 2. A detailed account is given of the structure of the secondary lamellae, where gaseous exchange takes place, and it is shown that two epithelial sheets are separated by a vascular axis mainly composed of pillar cells overlain by a basement membrane on each side. Blood pathways through the gills are discussed in relation to their respiratory function. 3. The embryonic development of gills is described and evidence regarding homo-logies of different structures, particularly the pillar cells, is reviewed. 4. The gills of fish having different modes of life show variations in (a) the number of arches, (b) the number and length of the gill filaments, and (c) the size and frequency of the secondary lamellae. Ways in which measurements of gill area may be carried out and some of the complications involved are reviewed and a summary given of measurements made for a wide variety of species. Measurements of the thickness of the water-blood barrier are also discussed; the more active fish generally have thinner water-blood barriers and larger gill areas. 5. The different mechanisms of gill ventilation are summarized and characteristics of gill resistance in elasmobranchs and teleosts are compared. Gas exchange is discussed in relation to available techniques and the current terminology and symbols, and to indicate the value of analogies between gill exchangers and systems studied by engineers. 6. It is outlined how studies of the functioning of gills during coughing, parasitic infection, and in polluted waters add to knowledge of their role in respiration.     

Trichodina sp. (Ciliophora: Peritrichida) in eel Anguilla anguilla in recirculation systems in denmark: host-parasite relations

Farmed eel cultured in recirculation systems in Denmark were found infected by Trichodina jadranica Raabe, 1958. Associations between parasite abundance and fish size was examined in 2 different production systems. In one system, stocked with relatively well-nourished eels (3 to 31 g), most of the parasites (66%) were found on the dorsal part on the skin and relatively few were found on the gills (approx. 8%). The infection level was significantly positively correlated both with fish weight and length. In the other system, stocked with relatively malnourished small eels (0.5 to 4 g), significantly more parasites (0.06 +/- 0.02 [SD]) were present on fish with a low condition factor than on fish with a higher condition factor (0.13 +/- 0.01 [SD]). In this eel stock the vast majority of the trichodines were found on the gills.