Fundulus Grandis Research Articles

Reproductive Output of Gulf Killifish at Different Stocking Densities in Static Outdoor Tanks

AbstractBroodstock of Gulf killifish Fundulus grandis (initial mean [±SD] weight = 8.2 ± 1.6 g) were held in static outdoor tanks at a 2:1 (female:male) sex ratio and at densities of 15, 30, or 45 fish/m3. Eggs were collected twice per week from spawning mats suspended 8 cm below the water surface. Collected eggs were measured volumetrically and assessed for total number, egg size, and embryo viability. Significantly more eggs were collected from tanks with manufactured (Spawntex) spawning mats than from tanks with cured Spanish moss spawning mats. Significant differences in the mean number of eggs collected were present among density treatments in April, June, and July. Stocking density was also associated with mortality; the greatest mortality was observed in tanks at the highest density (45 fish/m3). There was a clear pattern in egg production in relation to lunar phase, with egg production peaks occurring between full and new moon phases for all three stocking densities. Mean gonadosomatic index values differed significantly among sampling months for both females and males. Analysis of ovaries via histological sections indicated trends in oocyte stages that correlated to continued egg production for this fractionally spawning species. Peak egg production occurred between April and mid‐May, when mean (±SD) weekly water temperatures ranged from 23.8 ± 1.7°C to 28.2 ± 1.2°C. During April, the mean daily number of eggs collected translated to a reproductive output of approximately 0.90, 0.72, and 0.76 eggs/g of female for stocking densities of 15, 30, and 45 fish/m3, respectively. Based on egg output and mortality over the 5‐month spawning period, the intermediate density (30 fish/m3) appeared to provide the optimum number of eggs per individual over time for Gulf killifish broodstock held in static outdoor tanks at a 2:1 female:male sex ratio.

Orientation of larval and juvenile horseshoe crabs Limulus polyphemus to visual cues: Effects of chemical odors

Abstract Adult horseshoe crabs Limulus polyphemus have long served as models for the study of vision in marine arthropods. Yet, little is known about the ability of early life history stages to detect and respond to visual cues. We examined the visually directed movements of larvae and first stage juveniles to horizons containing dark visual targets of different sizes. The study tested the hypotheses that (1) larval and juvenile crabs can detect and respond to visual targets and (2) the direction of orientation varies with the presence of chemical cues associated with settlement habitats. Orientation of larval and juvenile crabs to rectangles subtending angles from 30–330° was tested in a circular arena containing water that either lacked estuarine chemical cues (offshore water) or contained odors from aquatic vegetation or known predators. In the absence of chemical odors, larvae oriented toward and juveniles moved away from dark horizons subtending angles > 60°. When placed in water containing chemical odors from potential nursery habitats, including the seagrasses Halodule wrightii and Syringodium filiforme, crabs reversed their direction of orientation relative to their responses in offshore water. Odors from two known predators, the mummichug Fundulus grandis and blue crab Callinectes sapidus, had no affect on the orientation of larvae. Yet, juveniles responded to both odors by moving toward the visual target. Results support the hypothesis that the visual orientation of larval and juvenile horseshoe crabs changes upon exposure to habitat and predator cues and that the direction of the response undergoes an ontogenetic shift following metamorphosis.

Production of Gulf Killifish in the Black Belt Region of Alabama Using Saline Groundwater

AbstractThe Gulf killifish Fundulus grandis, commonly called the bull minnow, mudminnow, or cacahoe, is a popular live bait along the Gulf of Mexico coast in the United States. Although culture techniques for Gulf killifish have been established for more than 30 years, this species is not commonly produced. Suitable coastal areas are thought to be one of the limitations. Low‐salinity groundwater occurs in much of the United States, including portions of the Alabama Black Belt region; however, the relative abundance of minerals in the water varies. Spawning success was compared among Gulf killifish held in saline groundwater (salinity = 4.2–11.1‰) from four inland sites and in 5‰ synthetic seawater. Gulf killifish production (fry to subadult) was compared between two sites of similar salinity but with different mineral profiles (site A had 4.0‰ salinity and received water from a fertilized source pond; site B had 3.2‰ salinity and received water with no fertilizer additions); initial stocking densities in 1.15‐m2 outdoor tanks were 25, 50, and 100 fry/m2. Gulf killifish spawned successfully under all conditions tested. Mean (±SD) fry production of 1.7 ± 0.88 fry/g of female was obtained. The yields and mean weights obtained differed by site and stocking density. After 104 d of production, the highest yield was 133.3 ± 40.4 g/m2 (1,333 ± 404 kg/ha) at site A for fish stocked at 100 fry/m2. The yield at site B for fish at the same stocking density was 62.6 ± 18.8 g/m2. Market‐sized fish (total length ≥ 6.35 cm; weight = 3.3 g) were obtained at site A for the stocking density of 25 fry/m2, but market‐sized fish were not obtained for any stocking density at site B. Although the mineral profiles differ between saline groundwater and natural seawater, saline groundwater is suitable for the reproduction and growth of this species. Egg production and fish growth were similar to levels expected for Gulf killifish culture in brackish‐water ponds along the Gulf of Mexico coast.

Adaptation versus Allometry: Population and Body Mass Effects on Hypoxic Metabolism inFundulus grandis

Hypoxia has significant effects on organisms, from metabolic reduction to death, and could be an important evolutionary force affecting the variation among populations within a species. To determine intraspecific variation in hypoxic metabolism and the effect of body mass, we examine rates of oxygen consumption (M(O2)) at seven oxygen concentrations among seven populations of Fundulus grandis that inhabit a mosaic of habitats with different frequencies and intensities of hypoxia. For M(O2), there is a significant interaction (P< 0.05) between body mass and oxygen concentrations: log(10) body mass: log(10) M(O2) slopes were steeper at intermediate oxygen partial pressures (Po(2)) than either normoxic or lowest Po(2) (ANCOVA, P<0.001). Additionally, the PO(2crit) (Po(2) where M(O2) can no longer be maintained) was a negative function of body mass (P < 0.04). At the lowest Po(2) (1.8 kPa), there was a significant difference in M(O2) among populations: one of the populations from environments more frequently stressed by hypoxia has greater M(O2) at the lowest oxygen concentrations. With few differences among populations, the most important effects were how body mass affected M(O2) at intermediate Po(2) and the negative relationship between body mass and PO(2crit). These findings suggest that an increase in body size is a useful strategy to minimize the effect of hypoxia.

Diel hypoxia in marsh creeks impairs the reproductive capacity of estuarine fish populations

Seasonal hypoxia in estuaries and continuous hypoxia in the laboratory significantly reduce the reproductive capacity of estuarine fish, but the impacts of diel hypoxia are unknown. This field study quantifies reproductive capacity in populations of gulf killifish (Fundulus grandis) sam- pled from sites with different dissolved oxygen cycles. Condition factor, gonadosomatic index (GSI), steroid sex hormones, and vitellogenin were measured in F. grandis captured in Weeks Bay (WB), Alabama (July 2004 and 2005) and Pensacola Bay (PB), Florida (July 2005). Compared to a control site with little or no diel hypoxia, testes and ovaries were significantly smaller under both moderate (WB: 2.61 mg l -1 , 0.6 h d -1 ; PB: 2.41 mg l -1 , 1.5 h d -1 ) and severe (0.93 mg l -1 , 3.4 h d -1 ) diel hypoxia. Male 11-ketotestosterone (11KT) concentrations were significantly lower under moderate hypoxia, while both testosterone (T) and 11KT concentrations were significantly reduced under severe diel hypoxia. In females, T concentrations were similar regardless of the occurrence or severity of hypoxia, but estradiol-17β (E2) concentrations were lower under severe diel hypoxia. Since T is unchanged, but its products 11KT and E2 are significantly reduced, hypoxia may affect reproduction by inhibiting spe- cific steroidogenic enzymes in the gonad. The association between diel hypoxia and lower reproduc- tive capacity was consistent across estuaries. Hypoxia-related changes in gulf killifish populations could affect trophic structure in marshes and bays since F. grandis move carbon from the upper marsh surface to the lower marsh, as increased biomass and as prey for juvenile piscivores.

Genetic Assessment of Species Ranges inFundulus heteroclitusandF. grandisin Northeastern Florida Salt Marshes

Abstract The limits of species ranges can be determined by a number of biotic and abiotic interactions, and in areas where closely related species overlap, some degree of reproductive isolation must exist in order for them to remain distinct. Understanding these interactions is essential for understanding what limits species distributions or causes hybridization. Fundulus heteroclitus (Mummichog) and Fundulus grandis (Gulf Killifish) are two closely related species with similar morphologies and ecological niches. Both species have widespread distributions that overlap in northeastern Florida. In the present study, two highly divergent loci (one nuclear and one mitochondrial) were utilized to distinguish these fundulid species in order to identify their ranges and to detect hybrids. Analysis of specimens collected along a north to south gradient in northeastern Florida salt marshes established that a relatively sharp transition (≈38 km) from relatively pure Mummichog populations to relatively pure F. grand...

Parasitic crustaceans as vectors of viruses, with an emphasis on three penaeid viruses

Parasitic crustaceans serve as both hosts and vectors of viruses as well as of parasites and other microbial pathogenic agents. Few of the presumably numerous associations are known, but many can be anticipated. Recently, branchiurans and gnathiid isopods have been documented to host helminths and blood parasites. Because the agents can be observed readily with a microscope, these are better recognized than are the smaller viral, bacterial, and fungal agents. Some agents are harmful to the host of the crustacean parasite and others are not. Viruses probably fit both these categories, since viruses that do not appear pathogenic are often seen in ultrastructural images from a range of invertebrate hosts, including crustaceans. Some viruses have been implicated in causing disease in the host, at least under appropriate conditions. For example, lymphocystis virus may possibly be transmitted to the dermis of its fish hosts by copepods and to the visceral organs by a cymothoid isopod. Similarly, argulid branchiurans seem to transmit the viral agent of spring viremia of carp as well as carp pox, and copepods have been implicated in transmitting infectious hematopoietic necrosis, infectious salmon anemia, and infectious pancreatic necrosis to salmon. Other viruses can be vectored to their hosts through an additional animal. We exposed three viruses, Taura syndrome virus (TSV), white spot syndrome virus (WSSV), and yellowhead virus (YHV), all of which cause mortalities in wild and cultured penaeid shrimps, to crustacean parasites on fish and crabs. Using real-time polymerase chain reaction analysis, we show that TSV in the cyclopoid copepod Ergasilus manicatus on the gill filaments of the Gulf killifish, Fundulus grandis, the acorn barnacle Chelonibia patula on the carapace of the blue crab, Callinectes sapidus, and gooseneck barnacle Octolasmis muelleri on the gills of C. sapidus, can replicate for at least 2 weeks and establish what should be an infective dose. This result was additionally supported by positive in situ hybridization reactions. All three parasites are the first known non-penaeid hosts in which replication occurs. The mean log copy number of WSSV also suggested that replication occurred in E. manicatus. The mean log copy number of YHV gradually decreased in all three parasites and both hosts over the 2-week period. The vector relationships indicate an additional potential means of transmitting and disseminating the disease-causing agents to the highly susceptible and economically valuable penaeid shrimp hosts.

Contemporary and historical influences on the genetic structure of the estuarine-dependent Gulf killifish Fundulus grandis

In comparison to species living in open marine environments, estuarine-dependent species are expected to exhibit stronger genetic population structure due to dispersal limitations. Estuarine habitats are relatively transitory on geological time scales; thus, populations may not be at migration-drift equilibrium, which could confound estimates of current day gene flow or selection. We used 8 nuclear microsatellite loci to investigate the genetic structure of the estuarine Gulf killifish Fundulus grandis across 10 populations along the northwestern and northeastern Gulf of Mexico. Patterns of isolation by distance, spatial autocorrelation, and assignment tests indicate that dispersal is limited and occurs primarily between neighboring sites. Principal component analysis and Bayesian clustering revealed evidence for genetic discontinuities located in Mobile Bay and western Florida which are near hypothesized biogeographical boundaries. There was also a significant nega- tive relationship between genetic diversity and latitude, a pattern consistent with the presence of hypothesized refugia in the southern Gulf regions during the Pleistocene that later recolonized the northern Gulf. Results suggest that populations may be at or near migration-drift equilibrium at a regional scale (e.g the western Gulf), but that dispersal barriers and potential historical signatures on population structure will need to be taken into consideration at larger spatial scales.

Simultaneous Extraction of High-Quality RNA and DNA from Small Tissue Samples

Purification of high-quality DNA and RNA from a single sample is becoming increasingly important for studies seeking both genomic and transcriptomic data. We compare different methods for isolating DNA and RNA from fish embryos (Gulf killifish; Fundulus grandis) and describe an optimal technique to extract high-quality DNA and RNA from a single embryo. The optimal method utilizes a chaotropic buffer and spin column technology. From embryos weighing approximately 4 mg, we were able to isolate an average of 6.1 microg of DNA and 1.1 microg of RNA per sample. Relative amounts of DNA and RNA can be adjusted as needed per study. Although these extraction trials were conducted on fish embryos, they can be potentially applied to small samples that typically do not yield high concentrations of nucleic acids.

Vertebrate osmoregulation: a student laboratory exercise using teleost fish

Here, we describe a laboratory experiment as part of an upper-level vertebrate physiology course for biology majors to investigate the physiological response of vertebrates to osmoregulatory challenges. The experiment involves measuring plasma osmolality and Na+-K+-ATPase activity in gill tissue of teleost fish acclimated to water of differing salinity. We describe results obtained using the widely available goldfish (Carassius auratus) and a common baitfish, the Gulf killifish (Fundulus grandis). The procedures described are generally applicable to other fish species, and they provide an alternative to the experimental use of humans or other mammalian species to investigate osmoregulation mechanisms. In addition to reenforcing the conceptual material covered in lecture, this laboratory exercise trains students in a wide range of laboratory and analytical skills, such as calculating and performing dilutions, pipetting, tissue sampling and homogenizing, preparing standard curves, conducting enzymatic assays, and analyzing and interpreting results. Typical student results are presented and discussed, as are common experimental and conceptual mistakes made by students.

Production sources and food web structure of a temperate tidal estuary: integration of dietary and stable isotope data

Food web structure and major sources of primary production consumed by metafauna of Mad Island Marsh, a coastal saltmarsh on the NW coast of the Gulf of Mexico, were compared using stable isotopes and dietary analysis. Carbon and nitrogen isotope data were entered into a mixing model containing 5 potential production sources. Results were inconclusive due to overlapping iso- topic signatures of certain sources, but nonetheless indicated that most fishes and macroinvertebrates assimilated material derived mostly from variable mixtures of macrophytes and filamentous algae. Highest estimates of percentage of material assimilated directly or indirectly from C4 marsh grasses (ranging from 30 to 82%) were for spot Leiostomus xanthurus and Gulf killifish Fundulus grandis. Isotopic analysis could not reveal the detailed structure of predator-prey interactions at the species level; greater detail of trophic pathways was revealed by the dietary analysis. Estimates of vertical web structure (species trophic levels) by the 2 methods were largely concordant. The exceptions were 2 zooplanktivorous and detritivorous fish species and grass shrimp Palaemonetes pugio that had higher trophic levels according to nitrogen isotope ratios. For these taxa, the isotopic method more accurately indexed the number of trophic transfers than the dietary method, which depends on accurate dietary estimation for all food chain components leading to a consumer, and which assumes equal assimilation efficiencies for items found in stomach contents. The isotopic method underesti- mated trophic levels of several invertebrates, possibly due to inaccurate estimation of mean δ 15 N for production sources supporting these taxa and/or differential trophic fractionation. Together, stable isotope and dietary analyses provide a more accurate assessment of food web structure and dynam- ics of coastal marsh ecosystems than either method alone.

Long term hypoxia suppresses reproductive capacity in the estuarine fish, Fundulus grandis

Human nutrient input has significantly altered dissolved oxygen (DO) cycles in coastal waters such that summertime hypoxia (DO < 2 mg/L) and anoxia of bottom water are common worldwide. Prolonged hypoxia usually reduces metabolic rate in fish and potentially reduces reproduction, particularly in a spring and summer spawning species such as the Gulf killifish, Fundulus grandis. To evaluate the effects of long term hypoxia on reproduction, Gulf killifish were subjected to either normoxia (6.68 ± 2.1 mg/L DO) or hypoxia (1.34 ± 0.45 mg/L DO) for one month. Fecundity, growth, gonadosomatic index (GSI), circulating sex steroids (testosterone, T; 11-ketotestosterone, 11KT; and estradiol-17β, E2), and egg yolk protein (vitellogenin, VTG) were measured. Hypoxia significantly reduced growth and reproduction. E2 was 50% lower in females and 11KT was 50% lower in males, although the precursor hormone T was unchanged in either sex after hypoxic exposure. Hypoxia-exposed females produced significantly fewer eggs and initiated spawning later than control fish. Plasma VTG concentration was unchanged, suggesting that hypoxia may delay VTG uptake by oocytes. Long term laboratory exposure clearly suppressed reproductive capacity in Gulf killifish. Wild populations experience cyclic hypoxia which could have equivalent effects if daily hypoxic periods are long and frequent — a potential consequence of anthropogenic nutrient enrichment in marsh systems.

Effects of long-term hypoxia on enzymes of carbohydrate metabolism in the Gulf killifish,Fundulus grandis

The goal of the current study was to generate a comprehensive, multi-tissue perspective of the effects of chronic hypoxic exposure on carbohydrate metabolism in the Gulf killifish Fundulus grandis. Fish were held at approximately 1.3 mg l(-1) dissolved oxygen (approximately 3.6 kPa) for 4 weeks, after which maximal activities were measured for all glycolytic enzymes in four tissues (white skeletal muscle, liver, heart and brain), as well as for enzymes of glycogen metabolism (in muscle and liver) and gluconeogenesis (in liver). The specific activities of enzymes of glycolysis and glycogen metabolism were strongly suppressed by hypoxia in white skeletal muscle, which may reflect decreased energy demand in this tissue during chronic hypoxia. In contrast, several enzyme specific activities were higher in liver tissue after hypoxic exposure, suggesting increased capacity for carbohydrate metabolism. Hypoxic exposure affected fewer enzymes in heart and brain than in skeletal muscle and liver, and the changes were smaller in magnitude, perhaps due to preferential perfusion of heart and brain during hypoxia. The specific activities of some gluconeogenic enzymes increased in liver during long-term hypoxic exposure, which may be coupled to increased protein catabolism in skeletal muscle. These results demonstrate that when intact fish are subjected to prolonged hypoxia, enzyme activities respond in a tissue-specific fashion reflecting the balance of energetic demands, metabolic role and oxygen supply of particular tissues. Furthermore, within glycolysis, the effects of hypoxia varied among enzymes, rather than being uniformly distributed among pathway enzymes.

Field investigation on the toxicity of Alaska North Slope crude oil (ANSC) and dispersed ANSC crude to Gulf killifish, Eastern oyster and white shrimp

A field investigation was conducted on a Louisiana Spartina alterniflora shoreline to evaluate the toxic effects of crude oil (Alaska North Slope crude oil, ANSC) and dispersed oil (ANSC + dispersant Corexit 9500) on three aquatic species indigenous to the Gulf of Mexico: Fundulus grandis (Gulf killifish), Crassostrea virginica (Eastern oyster), and Litopenaeus setiferus (white shrimp). Results indicated that total hydrocarbons concentration value in oiled treatments decreased rapidly in 3 h and were below 1 ppm at 24 h after initial treatment. Corexit 9500 facilitated more ANSC fractions to dissolve and disperse into the water column. L. setiferus showed short-term sensitivity to the ANSC and ANSC + 9500 at 30 ppm. However, most test organisms (>83%) of each species survived well after 24 h exposure to the treatments. Laboratory tests conducted concurrent with the field investigation indicated that concentrations of crude oil higher than 30 ppm were required for any significant toxic effect on the juvenile organisms tested.

Guidelines for reporting statistics in journals published by the American Physiological Society

Guidelines for reporting statistics in journals published by the American Physiological Society

The Effects of Season and Proximity to Fringing Mangroves on Seagrass-Associated Fish Communities in Charlotte Harbor, Florida

Little has been published on habitat use by fishes in Charlotte Harbor, one of Florida's largest-and still relatively pristine-estuaries. Multivariate analyses of data from 21.3-m-seine samples (1996-2000) were used to examine spatiotemporal patterns of seagrass habitat use by Charlotte Harbor fishes. Two habitats (mangrove-associated seagrass shorelines and offshore seagrass flats) were examined. Throughout the year, the mangrove-seagrass habitat was distinguished by Menidia spp., Mugil gyrans, Eucinostomus harengulus, and Floridichthys carpio, and the offshore seagrass flats habitat was distinguished by Bairdiella chrysoura, Orthopristis chrysoptera, and Cynoscion nebulosus. The dry season (Dec.-May) was distinguished by Lagodon rhomboides, Leiostomus xanthurus, and O. chrysoptera in both habitats and by Mugil cephalus in the mangrove-seagrass habitat. The wet season (June-Nov.) was distinguished by Lucania parva, F. carpio, and Fundulus grandis in the mangrove-seagrass habitat and by Eucinostomus gula, Microgobius gulosus, C. nebulosus, and E. harengulus in the offshore seagrass flats habitat. Eucinostomus spp., Anchoa mitchilli, and Strongylura notata were abundant in both habitats during the wet season. In general, many species were collected in both habitats and were widely distributed in the estuary. Various combinations of four environmental variables (i.e., water temperature, salinity, dissolved oxygen, water depth) and their coefficients of variation were well correlated with the biotic community patterns. These results show that different fish communities use different seagrass habitats in Charlotte Harbor, depending on the season and the proximity of the seagrasses to fringing mangroves.

Histopathological Evidence of Regeneration following Hepatotoxic Effects of the Cyanotoxin Microcystin-LR in the Hardhead Catfish and Gulf Killifish.

The cyanobacterial toxin microcystin-LR (MC-LR) causes liver damage in several freshwater fish species. In the present study, two estuarine species, hardhead catfish Arius felis and gulf killifish Fundulus grandis, were injected intraperitoneally with MC-LR at 45-300 μg/kg and their livers examined histopathologically for up to 23 d postinjection (PI) in the catfish and 5 d PI in the killifish. The livers from both species exhibited extensive, diffuse hepatocellular necrosis by 6 h PI. The necrosis persisted, and by day 2 large numbers of basophilic cells had emerged throughout the liver parenchyma. These cells occurred individually or in small clusters. By 72 h, the basophilic cells appeared to be highly proliferative with numerous mitotic figures and were arranged in cords and tubules similar to mature hepatic parenchyma. Regeneration of the liver parenchyma was noted in gulf killifish at 5 d PI, as tracts of basophilic cells were still evident. In hardhead catfish, there were no signs of necrosis or proliferating basophilic cells by 9 d PI, and the hepatic parenchyma appeared normal except for the vacuolation of many hepatocytes and some areas of hepatic megalocytosis. The degenerative changes in the two species studied were similar to but more pronounced than those observed in freshwater fish species exposed to MC-LR. Evidence of regeneration from the hepatotoxic effects of MC-LR suggests that the toxin can be used to study regenerative processes in fish livers.

Seasonal Differences in Hypoxia Tolerance in Gulf Killifish, Fundulus Grandis (Fundulidae)

Many estuarine habitats are characterized by episodes of hypoxia, the frequency and severity of which may vary seasonally. Accordingly, resident fish species may show seasonal differences in their capacity to tolerate hypoxia. We have tested this hypothesis in the gulf killifish, Fundulus grandis, sampled from the Lake Pontchartrain estuary (Louisiana) at different times of the year. We measured 2 indicators of hypoxia tolerance, the frequency of aquatic surface respiration (ASR) during gradual reduction in dissolved oxygen (D.O.) and survival time during severe hypoxic stress, and found both to be significantly affected by season. Fish collected during the summer did not engage in ASR until the D.O. concentration dropped to values lower than that required to elicit ASR by fish collected during other seasons. Laboratory acclimation of fish to low oxygen did not change the relationship between ASR behavior and D.O., suggesting that the observed seasonal effect on ASR was not simply due to previous exposure of summer fish to environmental hypoxia. Furthermore, fish collected during the summer and winter had significantly longer survival times during exposure to severe hypoxia than fish collected during the fall. Survival analysis indicated that the condition of fish was positively associated with survival time, and seasonal variation in condition accounted for about half of the observed difference between survival times of fish collected during the summer and fall. Seasonal variation in ASR and survival, when taken together, demonstrate that hypoxia tolerance in F. grandis may be subject to acclimatization. An increase in hypoxia tolerance during the summer could increase survivorship of fish when exposed to elevated temperatures and low oxygen concentrations which prevail during the summer months.

Immune function, hepatic CYP1A, and reproductive biomarker responses in the gulf killifish, Fundulus grandis, during dietary exposures to endocrine disrupters

The gulf killifish, Fundulus grandis, was used to determine the influence of biological rhythms on three biomarker responses. We first developed monoclonal antibodies against the model's immunoglobulins and vitellogenin in order to measure antibody responses and vitellogenesis, respectively. We then treated adults with 10, 1, 1, and 10 ppm of Aroclor 1254, tribuyltin, 3-methylcholanthrene, and nonyl-phenol, respectively, in mixtures over a 16-week period. The study followed Vibrio anguillarum-specific antibody responses, hepatic CYP1A, and plasma vitellogenin levels in the morning and again in the evening at 2-week intervals. The contaminated diet suppressed secondary antibody responses, but only in the morning. The contaminated diet also altered CYP1A, but not vitellogenesis. In addition, fish in the control group exhibited daily and seasonal differences in specific antibody levels and CYP1A induction. Moreover, circulating vitellogenin levels in control males sampled in the morning increased throughout the exposure, but remained below those of females. This study underscores the need to consider normal physiological rhythms when employing biomarkers in toxicology.

Oxygen consumption, blood lactate and inter-individual variation in the gulf killifish, Fundulus grandis, during hypoxia and recovery

Rates of oxygen consumption ( M ̇ O 2 ) for Fundulus grandis, the gulf killifish, were measured in air-saturated water, at four progressively lower levels of oxygen and upon normoxic recovery. The pattern of M ̇ O 2 versus oxygen partial pressure ( P wO 2) was that of an oxygen regulator, with a critical oxygen pressure ( P c) of 34 torr (1 torr=133.3 Pa). Below this value, M ̇ O 2 decreased and the concentration of blood lactate increased, indicating anaerobic metabolism during hypoxia. Recovery was characterized by elevated M ̇ O 2 compared to the initial normoxic exposure, coupled with the rapid clearance of blood lactate. Variation in M ̇ O 2 among the individual fish was appreciable and, in general, it was greater at higher levels of P wO 2. This inter-individual variation was significantly larger than the variation between replicate measures of M ̇ O 2 for a given individual, i.e. it cannot be attributed solely to random error. Furthermore, values for M ̇ O 2 during normoxia were found to be repeatable when the same fish were used in multiple experimental trials. The observation of significant, repeatable inter-individual variation in M ̇ O 2 suggests that such variation is a real and potentially important feature of fish metabolism.