Brain‐encysting trematodes and altered monoamine activity in naturally infected killifish Fundulus parvipinnis

This paper presents novel evidence to address mechanisms by which trematode parasites effect behavioural changes in naturally infected fish hosts. California killifish Fundulus parvipinnis infected with the brain‐encysting trematode Euhaplorchis californiensis display conspicuous swimming behaviours that render them 30 times more likely to be eaten by birds, the parasite's final host. Prevalence of E. californiensis reaches nearly 100% in most F. parvipinnis populations, with parasite biomass constituting almost 2% of F. parvipinnis biomass in some locations. Despite having thousands of cysts on their brains, infected fish grow and mature at rates comparable to those of uninfected populations. The lack of general pathology combined with the specificity of the altered behaviours suggests that the behavioural changes are due to parasite manipulation. The monoamine neurotransmitters serotonin and dopamine, which control locomotion and social behaviour in fishes and other vertebrates, were examined to explore the underlying mechanisms of this behaviour modification. Whereas previous studies were similarly conducted with experimentally infected fish, in this study, brain dopaminergic and serotonergic activity were analysed in naturally infected fish to assess how E. californiensis may alter F. parvipinnis monoamines in a naturally occurring system. A parasite density‐associated decrease in serotonergic activity occurred in the hippocampus of naturally infected fish, as well as a decrease in dopaminergic activity in the raphe nuclei, suggesting that E. californiensis inhibits serotonin and dopamine signaling in naturally infected F. parvipinnis. The neurochemical profile of infected fish is consistent with the hypothesis that E. californiensis affects brain monoaminergic systems in order to induce impulse‐driven, active, and aggressive behaviour in its hosts.     

Measuring water‐borne cortisol in Poecilia latipinna:is the process stressful,can stress be minimized and is cortisol correlated with sex steroid release rates?

The stress of water-borne hormone collection process was examined in sailfin mollies Poecilia latipinna. Baseline release rates of the stress hormone cortisol were measured and minimum confinement time for water sampling was evaluated for a standard 60 min v. a 30 min protocol. A 30 min hormone collection period reflects release rates over 60 min. Potential stress response to confinement in the beaker for the water-borne collection process was tested over 4 days. There was no evidence of stress due to the collection methods, as cortisol release rates did not differ significantly across four sequential days of handling for P. latipinna. Males and females did not differ significantly in baseline cortisol release rates. Baseline cortisol release rates from fish immediately after being collected in the field were also not significantly different than those in the 4 day confinement experiment. After exposure to a novel environment, however, P. latipinna mounted a stress response. Stress may also affect sex steroids and behaviour but cortisol release rates were not significantly correlated with sex steroids [11-ketotestosterone (KT), testosterone, or oestradiol], or mating attempts. The correlation between water-borne release rates and plasma steroid levels was validated for both cortisol and KT. Finally, normalizing cortisol release rates using standard length in lieu of mass is viable and accurate. Water-borne hormone assays are a valuable tool for investigating questions concerning the role of hormones in mediating stress responses and reproductive behaviours in P. latipinna and other livebearing fishes.     

Behavioural Interactions and Hormones in Naturally and Hatchery‐Spawned Chinook Salmon

Artificial breeding programmes commonly lead to domestication, which is associated with many behavioural differences that can reduce the success of animals released into natural environments. To better understand the factors contributing to domestication, we used a captive population of Chinook salmon (Oncorhynchus tshawytscha) to partition hormonal and behavioural differences to effects of the breeding method and rearing environment. We compared 9‐mo‐old juveniles from three lines that shared a common genetic background: (1) the Channel line produced by natural spawning and reared in a low‐density environment with a natural substrate for approx. 6 mo before being transferred to the hatchery; (2) the Hatchery line produced by artificial spawning; and (3) the Transfer line produced by natural spawning but reared in the hatchery from the eyed‐egg stage. Plasma concentrations of 11‐ketotestosterone (11‐KT) and cortisol were measured in groups of 150 fish and again after 4 d of social interactions in groups of six fish. There was no difference in 11‐KT among lines in large groups, but in small groups, Transfer fish had lower 11‐KT concentrations and were significantly less aggressive than both Channel and Hatchery fish. Regardless of group size, concentrations of the stress hormone cortisol were nearly twofold higher in Channel fish than in Hatchery and Transfer fish. Furthermore, the elevated cortisol concentrations in Channel fish were associated with 35% lower feeding rates than in the other two lines. Our study details complex behavioural and hormonal responses to breeding method and rearing environment in juvenile salmon.     

Further refinement of the non‐invasive procedure for measuring steroid production in the male three‐spined stickleback Gasterosteus aculeatus

Measurement of steroids that are released into the water via the gills has previously been shown to be an effective way of studying the reproductive endocrinology of the male three‐spined stickleback Gasterosteus aculeatus without having to kill the fish. In the present paper, a previous observation on the existence of a compound other than 11‐ketotestosterone (11‐KT) in water, which cross‐reacted in the 11‐KT radioimmunoassay was repeated. The amounts of this compound, however, were not sufficient to warrant a separation step prior to carrying out assay. The lack of association between androstenedione levels in water and those in plasma was also confirmed. For the first time, the amounts of testosterone released into the water were shown to be positively correlated with the amounts in plasma, the sampling procedure (placing the fish for 30 min in 50 ml water) had no effect on the rate of release of cortisol but caused a rapid drop in the rate of release of 11‐KT (which means that the fish should not be sampled twice in short succession), physical interaction between two nesting males (which was accompanied by aggression) significantly increased the rate of release of 11‐KT, androstenedione and testosterone (but not of cortisol) and the rate of release of 11‐KT was at its maximum between 2 and 4 h after exposure.     

Responses to handling and confinement stressors in juvenile great sturgeon Huso huso

The effects of acute stressors on physiological responses of juvenile great sturgeon or beluga Huso huso L. were investigated in two experiments. In the first experiment, fish were handled by placing them in containers at either low density (LD, one fish l^?1) or high density (HD, four fish l^?1) for 60 s. Concentrations of plasma cortisol, glucose and lactate were determined from blood collected at 0, 1, 3, 6 and 12 h after application of the stressor. Plasma cortisol concentrations increased after the disturbance in H. huso from both handling treatments, but changes were not significant. Plasma glucose rose significantly by 22·9 and 31·6% in LD and HD handling treatments, respectively, after 3 h. Significant increases in plasma lactate occurred within 1 h in both treatment groups, but that of the HD group was much higher. In the second experiment, fish were held at two different densities, LD (2 kg m^?2 tank bottom surface area) and HD (7 kg m^?2), for 8 weeks and then subjected to an aerial emersion handling stressor in a net for 60 s; blood samples were taken before handling (resting, 0 h) and at 1, 3, 6 and 9 h after handling. Plasma cortisol increased significantly in fish from the HD treatment from 8·8 ± 0·3 to 19·2 ± 2·4 ng ml^?1 (mean ±s.e. ) by 1 h after stress, but post‐handling changes in the LD group were not significant. Significant increases in both plasma glucose and lactate were observed by 1 h in both treatment groups, with peak levels of plasma glucose evident at 3 h [69·4 ± 2·9 and 60·9 ± 1·7 mg dl^?1 (mean ±s.e .) in LD and HD groups, respectively]. Plasma glucose levels were significantly higher in the LD group than in the HD group at 3 and 6 h. Post‐handling haemoglobin content increased by 1 h and white blood cell numbers were reduced by 3 and 6 h in the HD treatment group compared with resting values, but changes in these blood features in the LD group were not significant. Acute handling did not affect haematocrit in either treatment. The results suggest that H. huso is relatively resistant to handling and confinement, and could tolerate normal hatchery practices associated with aquaculture. Because changes in cortisol concentrations were relatively low compared with those in most teleosts, glucose and lactate concentrations may be more useful as stress indicators in juvenile H. huso. This study also demonstrated that prior exposure to a chronic stressor, specifically high stocking density, could alter the physiological response to subsequent acute handling in H. huso.     

Milt characteristics and plasma levels of gonadal steroids in greenback flounder Rhombosolea tapirina following treatment with exogenous hormones

Fish were treated with exogenous hormones, and milt and blood samples were collected for up to 96 h post‐treatment. Blood plasma samples were assayed for the gonadal steroids testosterone (T), 11‐ketotestosterone (11KT) and 17,20ß‐dihydroxy‐4‐pregnen‐3‐one (17.20ßP). Milt volume, spermatocrit and sperm motility were measured from milt samples. Non‐spermiated fish showed increased plasma T and 11KT in response to human chorionic gonadotropin (hCG) but not luteinising hormone releasing hormone analogue (LHRHa). Fish did not become spermiated in response to treatment with hCG, LHRHa, 11KT, 17‐hydroxyprogesterone (17P) or 17,20ßP. Spermiated fish showed an increase in milt volume in response to hCG and LHRHa but not exogenous steroids. Sperm motility declined to zero over 120 s and was not affected by hormone treatment or sampling time. Increased milt volume was accompanied by increased plasma T and 11KT, but not 17.20ßP levels. In a separate experiment, LHRHa delivered by injection or pellet was equally effective at increasing milt volume but had no effect on plasma steroid levels. Spermatocrit declined with stripping but was not affected by hormone treatment, nor was sperm motility. Co‐treatment of fish with 17P plus LHRHa had no additive effect on plasma steroid concentrations or milt volume. The results suggest that as in other teleosts, gonadotropin mediates milt production in greenback flounder.     

The rate of uptake of sex steroids from water by Tinca tinca is influenced by their affinity for sex steroid binding protein in plasma

Two experiments were carried out in which male and female tench Tinca tinca were placed in individual containers and tritiated steroids then added to the water. Water samples were collected over the next 6 or 7 h and the fish then sacrificed, bled and the gall bladder removed. Radioactivity was counted in all the samples. Over the course of the exposure period in the first experiment (7 h), radioactivity of 11‐ketotestosterone (11‐KT) in the water was depleted by 11%, 17,20β‐dihydroxypregn‐4‐en‐3‐one (17,20ß‐P) and 17,20α‐dihydroxypregn‐4‐en‐3‐one (17,20α‐P) by 28%, testosterone (T) by 56% and androstenedione (AD) by 68%. HPLC analysis of water samples at 3 h indicated that none of the steroids was extensively metabolized during the experiment. Females had a faster rate of uptake of AD than males. In the second experiment (6 h), radioactivity of cortisol in the water was depleted by 5%, 11‐KT by 7%, 17‐hydroxypregnen‐4‐ene (17‐P) by 17%, 17β‐oestradiol (E₂) by 35%, T by 37% and AD by 44%. In both experiments, the amounts of radioactivity that were recovered from the gall bladder and plasma were positively correlated with the rate of disappearance of radioactivity from the water. The ability of the steroids to bind to sex steroid binding protein (SBP) of tench plasma was tested by incubating plasma with radioactive steroids and then separating bound and free with ice cold dextran‐coated charcoal. When plasma at a final dilution of 1 : 60 (v/v) was incubated with 5 nM of each steroid, the percentage of radiolabel bound to SBP was: T 48% AD 44%, E₂ 30%, 17‐P 17%, 11‐KT 13·2%, 17,20α‐P 10·3%, 17,20β‐P 4·5% and cortisol 0%. Saturation analysis established dissociation constants (K_d; mean ± s .e .) of 3·4 ± 0·4, 2·2 ± 0·2, 4·0 ± 0·3. 9·0 ± 2·8 and 51·8 nM and binding capacities (B_max) of 201 ± 29, 201 ± 33, 165 ± 3, 187 ± 15 and 13·4 nM for T, AD, E₂,17‐P and 17,20β‐P respectively. The ability of steroids to displace tritiated T and AD from SBP was in the rank order AD > T > E₂ > 17,20αP = 17,20β‐P = 11‐KT = 17‐P > cortisol. Thus, the ability of tench plasma to bind certain steroids showed a relatively strong correlation with the ability of the fish to take up these steroids from water. Modelling of data for AD and 17,20β‐P helped to show why and how plasma binding had a strong influence on the rate of uptake (and hence release) of the steroids.     

Biometric and Allozymic Characterization of Three Coastal and Inland Killifish Populations (Pisces: Fundulidae) from the Peninsula of Baja California,Mexico

Type localities and/or central populations of coastal and inland killifishes along the peninsula of Baja California were sampled to survey the biometric and allozymic relationships among them. The standard stepwise discriminatory function analysis of biometric characters classified organisms into three well distinctive groups (p < 0.001). The number of scales in the lateral series successfully split the species, while the caudal length/standard length ratio did so between subspecies. Two diagnostic loci (PROT–2* and AAT–2*) segregated private alleles in each taxon. Fundulus lima showed lower genetic variability levels than F. parvipinnis spp., in both polymorphism (18.8% in F. lima vs. 25.0% in F. parvipinnisspp.) and expected heterozygosity (0.060 in F. lima vs 0.099 in F. p. brevis and 0.089 in F. p. parvipinnis). The reduced genetic variability in the threatened species F. lima may be related to its evolutionary history. Nei's genetic distance between F. lima and F. p. parvipinnis was 0.247, while 0.075 between F. parvipinnis spp.     

Effects of release procedures on the primary stress response and post‐release survival and growth of hatchery‐reared spotted seatrout Cynoscion nebulosus

To help explain the apparent poor post‐release success of hatchery‐reared (HR) spotted seatrout Cynoscion nebulosus, this study examined the effects of handling, transport and release procedures on the stress response of two age classes [48 and 80 day post‐hatch (dph)] of HR C. nebulosus, as measured by cortisol concentrations and the post‐release survival and growth of 48 and 80 dph HR C. nebulosus. As a proxy for stress, tissue cortisol was measured at various times during the handling, tagging (80 dph), transport, acclimation and release process. To consider the implications of the pre‐release stressors, growth and survival were monitored in separate field experiments for each age class of acclimated post‐transport C. nebulosus using control C. nebulosus that only experienced anaesthesia, transport, acclimation and a net release v. experimental C. nebulosus that underwent the entire routine procedure, including anaesthesia, tagging, transport, acclimation and gravity release through a pipe. For 48 dph C. nebulosus, mean cortisol varied significantly throughout handling and transport, increasing more than six‐fold from controls before decreasing in mean concentration just prior to release. For 80 dph C. nebulosus, cortisol varied throughout handling, tagging and transport, first increasing more than three‐fold compared with control C. nebulosus, before decreasing and rising slightly just prior to release. For 48 dph C. nebulosus within field enclosures, survival was high and similar for control and experimental groups; experimental C. nebulosus, however, were shorter, lighter and lower in condition than control C. nebulosus. For 80 dph C. nebulosus within field enclosures, fewer experimental C. nebulosus survived and those that did survive were of lower condition than C. nebulosus from the control group. Small untagged C. nebulosus may survive the release procedure better than larger C. nebulosus carrying a coded‐wire tag. These findings document some ways in which pre‐release practices may translate into detrimental effects on post‐release success of HR C. nebulosus.     

Ichthyofaunal utilization of newly-created versus natural salt marsh creeks in Mission Bay,CA

Loss of wetland habitat has proceeded at an alarmingrate in southern California, and increasingly marshrestoration and creation are being used to mitigatethese losses. As part of an effort to evaluatefunctional equivalence of created systems, theichthyofaunal assemblages in a created and adjacentnatural marsh in Mission Bay, San Diego, Californiawere compared. Fishes trapped in both marshes includedFundulus parvipinnis, Gillichthysmirabilis, Acanthogobius flavimanus, Ctenogobius sagittula, Atherinops affinis, andMugil cephalus. Fundulus parvipinniswasnumerically dominant in both systems, representing onaverage 69% of all fishes trapped in the createdmarsh and 65% of all fishes trapped in the naturalmarsh. Gillichthys mirabiliswas the second-mostabundant species, representing on average 31% of allfishes trapped in the created marsh and 28% of allfishes trapped in the natural marsh. Species richnessand dominance measures were similar between the twosystems, while abundances were higher in the naturalrelative to the created marsh. The size-structure ofF. parvipinnisand G. mirabilisdifferedbetween the created and natural marsh creeks, with thecreated marsh populations being skewed towards largersize classes. These size differences are believed toarise from differences in creek morphology between thecreated and natural systems, and potentially affectboth predators and prey of these species in the marsh.Mark-release-recapture revealed considerable marshfidelity, with as many as 35% of the F.parvipinnistagged in a marsh being recovered one daylater in the same marsh. Stable isotope analyses ofF. parvipinnisrevealed similar ¹⁵Nand ³⁴S values between marshes; howeverthere was a consistent enrichment in ¹³C (>3per mil) in tissues of F. parvipinnisfrom thecreated marsh, supporting the high marsh fidelitysuggested by tagging results. This first publisheddocumentation of the Mission Bay marsh resident fishessuggests that the created marsh ichthyofaunalassemblage was distinct in density and size structurefrom the adjacent natural marsh, and provides lessonsfor future restoration efforts.     

Evolution of host resistance and trade‐offs between virulence and transmission potential in an obligately killing parasite

Standard epidemiological theory predicts that parasites, which continuously release propagules during infection, face a trade‐off between virulence and transmission. However, little is known how host resistance and parasite virulence change during coevolution with obligate killers. To address this question we have set up a coevolution experiment evolving Nosema whitei on eight distinct lines of Tribolium castaneum. After 11 generations we conducted a time‐shift experiment infecting both the coevolved and the replicate control host lines with the original parasite source, and coevolved parasites from generation 8 and 11. We found higher survival in the coevolved host lines than in the matching control lines. In the parasite populations, virulence measured as host mortality decreased during coevolution, while sporeload stayed constant. Both patterns are compatible with adaptive evolution by selection for resistance in the host and by trade‐offs between virulence and transmission potential in the parasite.     

Density‐dependent parasitism in biological control of soil‐borne insects,nematodes, fungi and bacteria

Temporal density‐dependent parasitism appears to explain the development of some pest‐ and pathogen‐suppressive soils. In many examples, an enrichment of hosts (soil‐borne pests or pathogens) precedes and supports a build up of beneficial parasites in perennial crops or in monocultures, and the population dynamics of hosts and parasites may be described in epidemiological terms. Although the examples support the concept of a balance of nature based on density‐dependent regulation, we lack detailed understanding of the processes that drive these beneficial epidemics. In addition, the epidemics develop slowly and often do not provide the control expected by farmers. Current research, therefore, emphasizes inundative release of beneficial agents. Nevertheless, we should study epidemics in soil in order to learn how they might be enhanced; to learn how better to utilize inoculum that is inundatively added to soil; and to contribute to the general discussion of population biology and regulation.     

Quantitative trait loci for magnitude of the plasma cortisol response to confinement in rainbow trout

Better understanding of the mechanisms underlying interindividual variation in stress responses and their links with production traits is a key issue for sustainable animal breeding. In this study, we searched for quantitative trait loci (QTL) controlling the magnitude of the plasma cortisol stress response and compared them to body size traits in five F2 full‐sib families issued from two rainbow trout lines divergently selected for high or low post‐confinement plasma cortisol level. Approximately 1000 F2 individuals were individually tagged and exposed to two successive acute confinement challenges (1 month interval). Post‐stress plasma cortisol concentrations were determined for each fish. A medium density genome scan was carried out (268 markers, overall marker spacing less than 10 cM). QTL detection was performed using qtlmap software, based on an interval mapping method ( http://www.inra.fr/qtlmap ). Overall, QTL of medium individual effects on cortisol responsiveness (<10% of phenotypic variance) were detected on 18 chromosomes, strongly supporting the hypothesis that control of the trait is polygenic. Although a core array of QTL controlled cortisol concentrations at both challenges, several QTL seemed challenge specific, suggesting that responses to the first and to a subsequent exposure to the confinement stressor are distinct traits sharing only part of their genetic control. Chromosomal location of the steroidogenic acute regulatory protein (STAR) makes it a good potential candidate gene for one of the QTL. Finally, comparison of body size traits QTL (weight, length and body conformation) with cortisol‐associated QTL did not support evidence for negative genetic relationships between the two types of traits.     

Effectively vertical transmission of a Drosophila‐parasitic nematode: mechanism and consequences

1. Long‐term control of insects by parasites is possible only if the parasite populations persist. Because parasite transmission rate depends on host density, parasite populations may go extinct during periods of low host density. Vertical transmission of parasites, however, is independent of host density and may therefore provide a demographic bridge through times when their insect hosts are rare. 2. The nematode Howardula aoronymphium, which parasitises mycophagous species of Drosophila, can experience both horizontal and effectively vertical transmission, relative rates of which depend, in theory at least, on the density of hosts at breeding sites. 3. A nine‐generation experiment was carried out in which nematodes were transmitted either exclusively vertically or primarily horizontally. This experiment revealed that these parasites can persist and exhibit positive population growth even when there is only vertical transmission. 4. Assays at the end of the experiment revealed that the vertically transmitted nematodes had suffered no inbreeding depression and that they were similar to the horizontally transmitted nematodes in terms of virulence, infectivity, within‐host growth rate, and fecundity. Thus, vertical transmission of H. aoronymphium did not appear to compromise the ability of these parasites to control Drosophila populations.     

Effects of acute stress on aggression and the cortisol response in the African sharptooth catfish Clarias gariepinus: differences between day and night

African sharptooth catfish Clarias gariepinus were housed under continuous dim light (1 lx) or 12L:12D (350–0 lx) cycles. The number of skin lesions, as indicator of aggressive acts, and plasma cortisol levels, as indicator of stress‐axis activity, were measured at baseline as well as following a stressor (given in the light or dark phase). Results showed that (1) baseline plasma cortisol levels were not different between photoperiods, (2) the number of baseline skin lesions was highest for C. gariepinus housed under continuous dim light, (3) stressor‐induced peak levels of plasma cortisol were highest in the light phase and (4) the number of skin lesions following a stressor was highest in the dark phase. The higher number of stressor‐related skin lesions in the dark (active) phase suggests increased stressor‐induced aggression while in the active phase. In addition, the data suggest that housing under continuous dim light does not result in higher stress‐axis activity, as measured by baseline levels of cortisol, but does result in more stressor‐induced aggression, as measured by the higher number of skin lesions. The latter may be related to the fact that the continuous dim light photoperiod has twice the number of dark‐phase (active) hours in which stressor‐induced aggression is stronger compared to the 12L:12D photoperiod, which has a light phase in which stressor‐induced aggression is lower.     

How integrated are behavioral and endocrine stress response traits? A repeated measures approach to testing the stress-coping style model

It is widely expected that physiological and behavioral stress responses will be integrated within divergent stress‐coping styles (SCS) and that these may represent opposite ends of a continuously varying reactive–proactive axis. If such a model is valid, then stress response traits should be repeatable and physiological and behavioral responses should also change in an integrated manner along a major axis of among‐individual variation. While there is some evidence of association between endocrine and behavioral stress response traits, few studies incorporate repeated observations of both. To test this model, we use a multivariate, repeated measures approach in a captive‐bred population of Xiphophorus birchmanni. We quantify among‐individual variation in behavioral stress response to an open field trial (OFT) with simulated predator attack (SPA) and measure waterborne steroid hormone levels (cortisol, 11‐ketotestosterone) before and after exposure. Under the mild stress stimulus (OFT), (multivariate) behavioral variation among individuals was consistent with a strong axis of personality (shy–bold) or coping style (reactive–proactive) variation. However, behavioral responses to a moderate stressor (SPA) were less repeatable, and robust statistical support for repeatable endocrine state over the full sampling period was limited to 11‐ketotestosterone. Although post hoc analysis suggested cortisol expression was repeatable over short time periods, qualitative relationships between behavior and glucocorticoid levels were counter to our a priori expectations. Thus, while our results clearly show among‐individual differences in behavioral and endocrine traits associated with stress response, the correlation structure between these is not consistent with a simple proactive–reactive axis of integrated stress‐coping style. Additionally, the low repeatability of cortisol suggests caution is warranted if single observations (or indeed repeat measures over short sampling periods) of glucocorticoid traits are used in ecological or evolutionary studies focussed at the individual level.     

Robust geographical determinants of infection prevalence and a contrasting latitudinal diversity gradient for haemosporidian parasites in Western Palearctic birds

Identifying robust environmental predictors of infection probability is central to forecasting and mitigating the ongoing impacts of climate change on vector‐borne disease threats. We applied phylogenetic hierarchical models to a data set of 2,171 Western Palearctic individual birds from 47 species to determine how climate and landscape variation influence infection probability for three genera of haemosporidian blood parasites (Haemoproteus, Leucocytozoon, and Plasmodium). Our comparative models found compelling evidence that birds in areas with higher vegetation density (captured by the normalized difference vegetation index [NDVI]) had higher likelihoods of carrying parasite infection. Magnitudes of this relationship were remarkably similar across parasite genera considering that these parasites use different arthropod vectors and are widely presumed to be epidemiologically distinct. However, we also uncovered key differences among genera that highlighted complexities in their climate responses. In particular, prevalences of Haemoproteus and Plasmodium showed strong but contrasting relationships with winter temperatures, supporting mounting evidence that winter warming is a key environmental filter impacting the dynamics of host‐parasite interactions. Parasite phylogenetic community diversities demonstrated a clear but contrasting latitudinal gradient, with Haemoproteus diversity increasing towards the equator and Leucocytozoon diversity increasing towards the poles. Haemoproteus diversity also increased in regions with higher vegetation density, supporting our evidence that summer vegetation density is important for structuring the distributions of these parasites. Ongoing variation in winter temperatures and vegetation characteristics will probably have far‐reaching consequences for the transmission and spread of vector‐borne diseases.     

Concurrent levels of 11-ketotestosterone in fish surface mucus, muscle tissue and blood

A rapid, reproducible method is described for extracting and comparing levels of the ether‐soluble fish androgen 11‐ketotestosterone (11‐KT) in blood serum, muscle tissue and surface mucus. Because widely different volumes of extract were recovered after centrifugation from the three sources, it was important to express androgen levels as pg 11‐KT/mg of total soluble protein (TSP). For six male and four female sexually‐staged freshwater Koi (Cyprinus carpio), the method yielded similar pg 11‐KT/mg TSP ratios in blood serum and extracts of muscle tissue and surface mucus, with the strongest correlation between blood serum and surface mucus. While male Koi were distinguishable from females based on the magnitude of 11‐KT levels, reproductive stage and gonadosomatic index levels were not correlated with the 11‐KT levels of either sex. Similar pg 11‐KT/mg TSP ratios were also found for autologous muscle tissue and surface mucus extracts of 37 captured and sexed wild marine fishes representing seven genera. However, high 11‐KT levels were not restricted to mature males. Collectively, results suggest that surface mucus collection (followed by 11‐KT assay) is a useful alternative to more invasive methods of determining systemic hormone levels in fish. Without knowledge of seasonal variation in levels of this and other sex hormones, however, reliance on 11‐KT levels alone may lead to spurious identification of gender, let alone reproductive stage.     

Avian Plasmodium lineages found in spot surveys of mosquitoes from 2007 to 2010 at Sakata wetland,Japan: do dominant lineages persist for multiple years?

The ecology and geographical distribution of disease vectors are major determinants of spatial and temporal variations in the transmission dynamics of vector‐borne pathogens. However, there are limited studies on the ecology of vectors that contribute to the natural transmission of most vector‐borne pathogens. Avian Plasmodium parasites are multihost mosquito‐borne pathogens transmitted by multiple mosquito species, which might regulate the diversity and persistence of these parasites. From 2007 to 2010, we conducted entomological surveys at Sakata wetland in central Japan, to investigate temporal variation in mosquito occurrence and prevalence of avian Plasmodium lineages in the mosquito populations. A polymerase chain reaction (PCR)‐based method was used to detect Plasmodium parasites and identify the blood sources of mosquitoes. Culex inatomii and C. pipiens pallens represented 60.0% and 34.8% of 11 mosquito species collected, respectively. Our results showed that the two dominant mosquito species most likely serve as principal vectors of avian Plasmodium parasites during June, which coincides with the breeding season of bird species nesting in the wetland reed beds. Fourteen animal species were identified as blood sources of mosquitoes, with the oriental reed warbler (Acrocephalus orientalis) being the commonest blood source. Although there was significant temporal variation in the occurrence of mosquitoes and prevalence of Plasmodium lineages in the mosquitoes, the dominant Plasmodium lineages shared by the two dominant mosquito species were consistently found at the same time during transmission seasons. Because vector competence cannot be confirmed solely by PCR approaches, experimental demonstration is required to provide definitive evidence of transmission suggested in this study.     

Fine‐scale distribution modeling of avian malaria vectors in north‐central Kansas

Infectious diseases increasingly play a role in the decline of wildlife populations. Vector‐borne diseases, in particular, have been implicated in mass mortality events and localized population declines are threatening some species with extinction. Transmission patterns for vector‐borne diseases are influenced by the spatial distribution of vectors and are therefore not uniform across the landscape. Avian malaria is a globally distributed vector‐borne disease that has been shown to affect endemic bird populations of North America. We evaluated shared habitat use between avian malaria vectors, mosquitoes in the genus Culex and a native grassland bird, the Greater Prairie‐Chicken (Tympanuchus cupido), by (1) modeling the distribution of Culex spp. occurrence across the Smoky Hills of north‐central Kansas using detection data and habitat variables, (2) assessing the occurrence of these vectors at nests of female Greater Prairie‐Chickens, and (3) evaluating if shared habitat use between vectors and hosts is correlated with malarial infection status of the Greater Prairie‐Chicken. Our results indicate that Culex occurrence increased at nest locations compared to other available but unoccupied grassland habitats; however the shared habitat use between vectors and hosts did not result in an increased prevalence of malarial parasites in Greater Prairie‐Chickens that occupied habitats with high vector occurrence. We developed a predictive map to illustrate the associations between Culex occurrence and infection status with malarial parasites in an obligate grassland bird that may be used to guide management decisions to limit the spread of vector‐borne diseases.