Directional movement of steinernematid nematodes in response to electrical current

Steinernematid nematodes are parasites that are important natural regulating agents of insect populations. The infective juvenile nematodes respond to a variety of stimuli that aid in survival and host finding. Host finding strategies among steinernematids differ along a continuum from ambush (sit & wait) to cruiser (search & destroy). In this paper we describe directional movement in response to an electrical current, which was generated on agar plates. Specifically, Steinernema glaseri (a cruiser) moved to a higher electric potential, whereas Steinernema carpocapsae, an ambusher, moved to a lower electric potential. Thus, we hypothesize that steinernematids may detect electrical currents or electromagnetic fields in nature, and these stimuli may be used differentially among species for host finding or enhancing other fitness characters.     

Absence of Prey Chemical Discrimination by Tongue-flicking in an Ambush-foraging Lizard Having Actively Foraging Ancestors

Lingually mediated detection of prey chemicals is widespread in one major clade of lizards, Scleroglossa, but rare in the other, Iguania. It is absent in all ambush-foraging families tested and present in all actively foraging families. In Iguania, prey chemical discrimination is known only in the herbivorous Iguanidae; in Scleroglossa, it was heretofore known to be absent only in ambush-foraging gekkonids. Because ambush foraging precludes lingual sampling of a wide area and tongue-flicking would disrupt the crypticity ambushers maintain by immobility, we predicted that prey chemical discrimination would be absent in scleroglossans that have secondarily adopted ambush foraging. The Cape girdled lizard, Cordylus cordylus, is member of Cordylidae, a family of ambush foragers considered derived from active foragers in the Autarchoglossa, a group of scleroglossan families having highly developed lingual chemosensory behaviours. As predicted, this species did not discriminate surface chemicals of three prey species from control substances in a series of standardized experiments in which prey chemicals were presented on cotton-tipped applicators. Thus, even in taxa having highly developed prey chemical discrimination, adoption of ambush foraging may induce loss of prey chemical discrimination, providing further and stronger evidence that prey chemical discrimination is adaptively adjusted to foraging mode.     

Perspectives on the Behavior of Entomopathogenic Nematodes from Dispersal to Reproduction: Traits Contributing to Nematode Fitness and Biocontrol Efficacy

The entomopathogenic nematodes (EPN) Heterorhabditis and Steinernema are widely used for the biological control of insect pests and are gaining importance as model organisms for studying parasitism and symbiosis. In this paper recent advances in the understanding of EPN behavior are reviewed. The “foraging strategy” paradigm (distinction between species with ambush and cruise strategies) as applied to EPN is being challenged and alternative paradigms proposed. Infection decisions are based on condition of the potential host, and it is becoming clear that already-infected and even long-dead hosts may be invaded, as well as healthy live hosts. The state of the infective juvenile (IJ) also influences infection, and evidence for a phased increase in infectivity of EPN species is mounting. The possibility of social behavior - adaptive interactions between IJs outside the host - is discussed. EPNs’ symbiotic bacteria (Photorhabdus and Xenorhabdus) are important for killing the host and rendering it suitable for nematode reproduction, but may reduce survival of IJs, resulting in a trade-off between survival and reproduction. The symbiont also contributes to defence of the cadaver by affecting food-choice decisions of insect and avian scavengers. I review EPN reproductive behavior (including sperm competition, copulation and evidence for attractive and organizational effects of pheromones), and consider the role of endotokia matricida as parental behavior exploited by the symbiont for transmission.     

Foraging mode of a Madagascan iguanian lizard, Oplurus cuvieri cuvieri

The foraging mode of lizards has been extensively studied in members of most major families. One neglected taxon is Opluridae, a distinct Iguanian group endemic to Madagascar. We studied the foraging mode of Oplurus cuvieri cuvieri in a dry forest in north-western Madagascar. Quantitative data on the number of movements per minute and percentage of time spent in moving indicated that O. c. cuvieri is a typical ambusher, although the lizard occasionally fed on plant materials by active foraging. Possible seasonal changes in foraging activity were suggested. The results support the previous view that insectivorous iguanian lizards are ambush foragers.     

Characterization of New Entomopathogenic Nematodes from Thailand: Foraging Behavior and Virulence to the Greater Wax Moth, Galleria mellonella L. (Lepidoptera: Pyralidae)

Entomopathogenic nematodes (EPNs) in the genera Steinernema and Heterorhabditis and their associated bacteria (Xenorhabdus spp. and Photorhabdus spp., respectively) are lethal parasites of soil dwelling insects. We collected 168 soil samples from five provinces, all located in southern Thailand. Eight strains of EPNs were isolated and identified to species using restriction profiles and sequence analysis. Five of the isolates were identified as Heterorhabditis indica, and one as Heterorhabditis baujardi. Two undescribed Steinernema spp. were also discovered which matched no published sequences and grouped separately from the other DNA restriction profiles. Behavioral tests showed that all Heterorhabditis spp. were cruise foragers, based on their attraction to volatile cues and lack of body-waving and standing behaviors, while the Steinernema isolates were more intermediate in foraging behavior. The infectivity of Thai EPN strains against Galleria mellonella larvae was investigated using sand column bioassays and the LC(50) was calculated based on exposures to nematodes in 24-well plates. The LC(50) results ranged from 1.99-6.95 IJs/insect. Nine centimeter columns of either sandy loam or sandy clay loam were used to determine the nematodes' ability to locate and infect subterranean insects in different soil types. The undescribed Steinernema sp. had the greatest infection rate in both soil types compared to the other Thai isolates and three commercial EPNs (Heterorhabditis bacteriophora, Steinernema glaseri and Steinernema riobrave).     

Directional movement of entomopathogenic nematodes in response to electrical field: effects of species, magnitude of voltage, and infective juvenile age

Entomopathogenic nematodes respond to a variety of stimuli when foraging. Previously, we reported a directional response to electrical fields for two entomopathogenic nematode species; specifically, when electrical fields were generated on agar plates Steinernema glaseri (a nematode that utilizes a cruiser-type foraging strategy) moved to a higher electric potential, whereas Steinernema carpocapsae, an ambush-type forager, moved to a lower potential. Thus, we hypothesized that entomopathogenic nematode directional response to electrical fields varies among species, and may be related to foraging strategy. In this study, we tested the hypothesis by comparing directional response among seven additional nematode species: Heterorhabditis bacteriophora, Heterorhabditis georgiana, Heterorhabditis indica, Heterorhabditis megidis, Steinernema feltiae, Steinernema riobrave, and Steinernema siamkayai. S. carpocapsae and S. glaseri were also included as positive controls. Heterorhabditids tend toward cruiser foraging approaches whereas S. siamkayai is an ambusher and S. feltiae and S. riobrave are intermediate. Additionally, we determined the lowest voltage that would elicit a directional response (tested in S. feltiae and S. carpocapsae), and we investigated the impact of nematode age on response to electrical field in S. carpocapsae. In the experiment measuring diversity of response among species, we did not detect any response to electrical fields among the heterorhabditids except for H. georgiana, which moved to a higher electrical potential; S. glaseri and S. riobrave also moved to a higher potential, whereas S. carpocapsae, S. feltiae, and S. siamkayai moved to a lower potential. Overall our hypothesis that foraging strategy can predict directional response was supported (in the nematodes that exhibited a response). The lowest electric potential that elicited a response was 0.1 V, which is comparable to electrical potential associated with some insects and plant roots. The level of response to electrical potential diminished with nematode age. These results expand our knowledge of electrical fields as cues that may be used by entomopathogenic nematodes for host-finding or other aspects of navigation in the soil.     

Forager abundance and its relationship with colony activity level in three species of South American Pogonomyrmex harvester ants

The proportion of foragers in ant colonies is a fairly constant species-specific characteristic that could be determined by intrinsic or extrinsic factors. If intrinsic factors are relevant, species with similar life history characteristics (e.g., colony size and foraging strategies) would be expected to have a similar proportion of foragers in their colonies. Within the genus Pogonomyrmex, North American species can vary largely in their colony size, whereas only species with small colonies are known in South America. We studied the characteristics of the foraging subcaste in three sympatric South American species of Pogonomyrmex harvester ants, and compared it with the available information on other species of the same genus. We used two mark-recapture methods and colony excavations to estimate the number and proportion of foragers in the colonies of P. mendozanus, P. inermis, and P. rastratus, and to test the relationship between forager external activity levels and abundance per colony. Forager abundance in the three studied species was lower than in most North American species. The percentage of foragers in their colonies ranged 7–15 %, more similar to North American species with large colonies than to those with small colony size. Foraging activity was positively correlated with forager abundance in all three species, implying that colony allocation to number of foragers allows for higher food acquisition. Further comparative studies involving a wider range of traits in South and North American species would allow to unveil the role of environmental factors in shaping each species’ particular traits.     

An Adaptive Difference in the Relationship between Foraging Mode and Responses to Prey Chemicals in two Congeneric Scincid Lizards

Sensory abilities must allow efficient detection of prey, but the senses used and their relative importance may vary with hunting methods. In lizards, ambush foragers locate prey visually and active foragers use a combination of vision and vomerolfaction, the chemical sense associated with the vomeronasal system. Active foragers, but not ambush foragers, discriminate between prey chemicals and other chemical stimuli sampled by tongue-flicking. In active foragers, features of the tongue that may improve chemical sampling, such as elongation and forking are more pronounced and density of vomeronasal chemoreceptors is greater, than in ambush foragers. Foraging mode is fixed in most lizard families, and correlated evolution has been demonstrated among foraging mode, discrimination of prey chemicals, and lingual-vomeronasal morphology by interfamilial comparisons. Here I present information on a rare case of an intrageneric difference in foraging mode in the genus Mabuya . Laboratory experiments on the discrimination of prey chemicals showed that the active forager M . striata sparsa exhibits prey chemical discrimination, but the ambush forager M . acutilabris does not. The active forager also has a slightly more elongated tongue with deeper notching at the tip than the ambush forager, which might be a response to a change in foraging behavior or a reflection of unrelated differences in head shape. These findings confirm predictions based on correlated evolution between the hunting method and use of the chemical sense to locate food. They further show that chemosensory behavior is adjusted to change in foraging mode more rapidly than was previously known and suggest that behavioral changes may occur more rapidly than associated modifications of chemosensory morphology.     

A multigene approach for assessing evolutionary relationships of Xenorhabdus spp. (γ-Proteobacteria), the bacterial symbionts of entomopathogenic Steinernema nematodes

Xenorhabdus spp., are gram-negative bacterial symbionts of entomopathogenic nematodes in the genus Steinernema. A specialized and intimate relationship exists between nematode and bacteria, affecting many of their life history traits, such as nutrition, dispersal, host-finding, foraging and defense from biotic and abiotic factors. Xenorhabdus currently comprises more than 20 species isolated from Steinernema spp. with diverse host range, host foraging behavior, reproductive modes and environmental tolerance. Xenorhabdus phylogenies have historically been based on 16s rDNA sequence analyses, and only recently has data from housekeeping genes been employed. The prevalence of lateral gene transfer among bacteria calls for a wider perspective when considering their phylogeny. With the increasing number of Xenorhabdus species and strains, various perspectives need to be considered for investigating the evolutionary history of these nematode bacterial symbionts, In this study, we reconstruct the evolutionary histories of 30 species of Xenorhabdus considering the traditional 16s rDNA gene region as well as the housekeeping genes recA and serC. Datasets were analyzed individually and then combined, using a variety of phylogenetic criteria.     

Social traditions and social learning in capuchin monkeys (Cebus)

Capuchin monkeys (genus Cebus) have evolutionarily converged with humans and chimpanzees in a number of ways, including large brain size, omnivory and extractive foraging, extensive cooperation and coalitionary behaviour and a reliance on social learning. Recent research has documented a richer repertoire of group-specific social conventions in the coalition-prone Cebus capucinus than in any other non-human primate species; these social rituals appear designed to test the strength of social bonds. Such diverse social conventions have not yet been noted in Cebus apella, despite extensive observation at multiple sites. The more robust and widely distributed C. apella is notable for the diversity of its tool-use repertoire, particularly in marginal habitats. Although C. capucinus does not often use tools, white-faced capuchins do specialize in foods requiring multi-step processing, and there are often multiple techniques used by different individuals within the same social group. Immatures preferentially observe foragers who are eating rare foods and hard-to-process foods. Young foragers, especially females, tend to adopt the same foraging techniques as their close associates.     

Ambush or active life: foraging mode influences haematocrit levels in snakes

The rate at which organisms acquire resources is a critical trait and foraging mode can vary from sit‐and‐wait tactics to being highly mobile and active. Snakes provide a robust opportunity to examine the physiological correlates of contrasted foraging strategies. In this context, haematocrit (Hct), a proxy of blood oxygen carrying capacity, should be a reliable indicator of aerobic activity levels. We used phylogenetically informed models to examine the relationship between foraging mode and Hct in 80 snake species. After accounting for clade and habitat effects, we found a significant relationship of Hct with foraging mode; Hct is lower in snakes that ambush prey compared to active foragers across habitats. Species using both foraging tactics had marginally lower Hct than active foragers. Ambush foraging tactics are widespread in snakes, notably among low‐energy specialists that usually display low feeding frequency, as well as limited activity and daily movements. Because Hct influences blood viscosity, low levels may thus be advantageous by reducing maintenance and locomotory costs. Further studies are required to better understand the implication of foraging mode on blood characteristics and other aspects of snake physiology. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 111 , 636–645.     

How phylogeny and foraging ecology drive the level of chemosensory exploration in lizards and snakes

The chemical senses are crucial for squamates (lizards and snakes). The extent to which squamates utilize their chemosensory system, however, varies greatly among taxa and species’ foraging strategies, and played an influential role in squamate evolution. In lizards, ‘Scleroglossa’ evolved a state where species use chemical cues to search for food (active foragers), whereas ‘Iguania’ retained the use of vision to hunt prey (ambush foragers). However, such strict dichotomy is flawed as shifts in foraging modes have occurred in all clades. Here, we attempted to disentangle effects of foraging ecology from phylogenetic trait conservatism as leading cause of the disparity in chemosensory investment among squamates. To do so, we used species’ tongue‐flick rate (TFR) in the absence of ecological relevant chemical stimuli as a proxy for its fundamental level of chemosensory investigation, that is baseline TFR. Based on literature data of nearly 100 species and using phylogenetic comparative methods, we tested whether and how foraging mode and diet affect baseline TFR. Our results show that baseline TFR is higher in active than ambush foragers. Although baseline TFRs appear phylogenetically stable in some lizard taxa, that is a consequence of concordant stability of foraging mode: when foraging mode shifts within taxa, so does baseline TFR. Also, baseline TFR is a good predictor of prey chemical discriminatory ability, as we established a strong positive relationship between baseline TFR and TFR in response to prey. Baseline TFR is unrelated to diet. Essentially, foraging mode, not phylogenetic relatedness, drives convergent evolution of similar levels of squamate chemosensory investigation.     

Routine Cryopreservation of Isolates of Steinernema and Heterorhabditis spp.

Infective-stage juveniles of Steinernema and Heterorhabditis spp. were cryopreserved using two-stage incubation in glycerol and 70% methanol before storage in cryotubes in liquid nitrogen. Optimal glycerol concentrations and incubation times for survival were determined for different species, but acceptable survival of all species and isolates of entomopathogenic nematodes can be obtained using 15% (w/w) glycerol and incubation for 48 hours. Mean survival was 69% for isolates of Steinernema and 68% for isolates of Heterorhabditis (n = 84). The maximum survival recorded was 97% for S. feltiae K254 stored in liquid nitrogen for 12 months.     

Neosteinernema longicurvicauda n. gen., n. sp. (Rhabditida: Steinernematidae), a Parasite of the Termite Reticuldermes flavipes (Koller)

A nematode isolated from the termite Reticulitermes flavipes (Koller) was identified and described as a new genus and species, Neosteinernema longicurvicauda. Primary distinguishing characters, by contrast to members of the genus Steinernema, were females having prominent phasmids, a curved tail longer than the body width at the anus, a spiral shape in juvenile-bearing females, and juveniles becoming infective-stage juveniles before emerging from the female; males having prominent phasmids, a digitate tail tip, a characteristic shape of the spicules (foot-shaped with a hump on the dorsal side), and 13-14 pairs of genital papillae, with eight pairs preanal; and infective juveniles having prominent phasmids and a filiform curved tail as long as the esophagus. Adult nematodes are found outside the termite cadaver. Diagnosis of the family Steinernematidae was emended to accommodate the new species.     

Comparison of Three Methods for Estimating the Number of Entomopathogenic Nematodes Present in Soil Samples

Numbers of Steinernema sp. (CB2B) and S. carpocapsae (Agriotos) exponentially declined after application into a clay loam soil. Over a 35-day sampling period, Steinernema sp. (CB2B) was more persistent than S. carpocapsae (Agriotos). The presence or absence of the second-stage cuticle on the third-stage juveniles (J3) at the time of application did not alter the rate of population decline of Steinernema sp. (CB2B). Nearly all J3 of Steinernema sp. (CB2B) and S. carpocapsae (Agriotos) lost their cuticle within 24 hours of being in soil. Centrifugal flotation recovered the greatest number of nematodes, with a lower variance than either the live bait or Baermann funnel techniques. A strong positive linear relationship was evident between numbers of nematodes present in the soil and the numbers that established in a bait insect. Approximately 40% of Steinernema sp. (CB2B) and 30% of the S. carpocapsae (Agriotos) present in the soil established in Galleria mellonella larvae. The extraction techniques had different efficiencies and gave different relative estimates of persistence for the two species. Persistence and infectivity was best measured using a combination of live bait and flotation techniques.     

Assessing Nuclear and Mitochondrial DNA Sequence Variation Within Steinernema (Rhabditida: Steinernematidae)

DNA sequence analysis was used to characterize the nuclear ribosomal DNA ITS1 region and a portion of the COII and 16S rDNA genes of the mitochondrial genome from Steinernema entomopathogenic nematodes. Nuclear ITS1 nucleotide divergence among seven Steinernema spp. ranged from 6 to 22%, and mtDNA divergence among five species ranged from 12 to 20%. No intraspecific variation was observed among three S. feltiae strains. Phylogenetic analysis of both nuclear and mitochondrial DNA sequences confirms the existing morphological relationships of several Steinernema species. Both the rDNA ITS1 and mtDNA sequences were useful for resolving relationships among Steinernema taxa.     

Attraction of entomopathogenic nematodes Steinernema carpocapsae and Heterorhabditis bacteriophora to the red palm weevil (Rhynchophorus ferrugineus)

The red palm weevil (RPW), Rhynchophorus ferrugineus, is a serious pest of date palms. Its larvae bore deep into the trunk disrupt the vascular tissues and kill the infested trees. Behavioral features of entomopathogenic nematodes (EPNs), reflected by attraction and distribution patterns, are fundamental aspect in determining their parasitic ability and potential management of RPW. We studied the attraction behavior of the EPNs Steinernema carpocapsae and Heterorhabditis bacteriophora to the RPW under simulated natural conditions in tubes to evaluate their infective potential. In all experiments, a certain proportion of infective juveniles (IJs) (16–20%) stayed near the inoculated site and a major proportion (38–48%) was attracted to the host end. Both H. bacteriophora and S. carpocapsae were efficient crawlers, climbing up and descending when locating their insect host. They were efficiently attracted to the various larval sizes and stages of the RPW life cycle. Host localization by ascending movement was more prominent in S. carpocapsae than in H. bacteriophora. In general, H. bacteriophora is classified as a cruiser forager and S. carpocapsae as an ambusher. However, in this study, we discovered a higher percentage of cruiser foragers among S. carpocapsae IJs. They dispersed much faster and their cruising behavior was prominent characteristic in controlling the cryptic RPW concealed in organic habitats.     

Morphometrics of Infective Juveniles of Steinernema spp. and Heterorhabditis bacteriophora (Nemata: Rhabditida)

Selected morphometrics of Heterorhabditis bacteriophora and seven species of Steinernema from in vivo culture were compared in relation to time of harvest. In addition, five Steinernema species were reared in vitro and their morphometrics were compared with those from in vivo culture. With in vivo culture, there was generally a negative linear relationship between body length of infective juveniles (IJ) and time of harvest. The distance from the anterior end to the excretory pore (EP) and the tail length (T) of IJ also varied with time of harvest. The E percentage (= EP/T x 100) was the least variable. Body lengths of IJ reared in vitro were much less than those of IJ reared in vivo. The study suggests that IJ harvested from in vivo culture within 1 week of emergence from cadavers are best for species identification. Infective juveniles from in vitro culture should not be used for species identification.     

Phylogeny,biogeography and ecological diversification of Sarcocornia (Salicornioideae,Amaranthaceae)

Background and Aims Sarcocornia comprises about 28 species of perennial succulent halophytes distributed worldwide, mainly in saline environments of warm-temperate and subtropical regions. The genus is characterized by strongly reduced leaves and flowers, which cause taxonomic difficulties; however, species in the genus show high diversity in growth form, with a mat-forming habit found in coastal salt marshes of all continents. Sarcocornia forms a monophyletic lineage with Salicornia whose species are all annual, yet the relationship between the two genera is poorly understood. This study is aimed at clarifying the phylogenetic relationship between Sarcocornia and Salicornia, interpreting biogeographical and ecological patterns in Sarcocornia, and gaining insights into putative parallel evolution of habit as an adaptation to environmental factors.Methods A comprehensively sampled and dated phylogeny of Sarcocornia is presented based on nuclear ribosomal DNA (external transcribed spacer) and chloroplast DNA (atpB-rbcL, rpl32-trnL) sequences; representative samples of Salicornia were also included in the analyses. To infer biogeographical patterns, an ancestral area reconstruction was conducted.Key Results The Sarcocornia/Salicornia lineage arose during the Mid-Miocene from Eurasian ancestors and diversified into four subclades: the Salicornia clade, the American Sarcocornia clade, the Eurasian Sarcocornia clade and the South African/Australian Sarcocornia clade. Sarcocornia is supported as paraphyletic, with Salicornia nested within Sarcocornia being sister to the American/Eurasian Sarcocornia clade. The American and the South African/Australian Sarcocornia clade as well as the Salicornia clade were reconstructed to be of Eurasian origin. The prostrate, mat-forming habit arose multiple times in Sarcocornia.Conclusions Sarcocornia diversified in salt-laden environments worldwide, repeatedly evolving superficially similar prostrate, mat-forming habits that seem advantageous in stressed environments with prolonged flooding, high tidal movement and frost. Some of these prostrate-habit types might be considered as ecotypes (e.g. S. pacifica or S. pillansii) while others represent good ecospecies (e.g. S. perennis, S. decumbens, S. capensis), hence representing different stages of speciation.     

Interaction between Varroa destructor and imidacloprid reduces flight capacity of honeybees

Current high losses of honeybees seriously threaten crop pollination. Whereas parasite exposure is acknowledged as an important cause of these losses, the role of insecticides is controversial. Parasites and neonicotinoid insecticides reduce homing success of foragers (e.g. by reduced orientation), but it is unknown whether they negatively affect flight capacity. We investigated how exposing colonies to the parasitic mite Varroa destructor and the neonicotinoid insecticide imidacloprid affect flight capacity of foragers. Flight distance, time and speed of foragers were measured in flight mills to assess the relative and interactive effects of high V. destructor load and a field-realistic, chronic sub-lethal dose of imidacloprid. Foragers from colonies exposed to high levels of V. destructor flew shorter distances, with a larger effect when also exposed to imidacloprid. Bee body mass partly explained our results as bees were heavier when exposed to these stressors, possibly due to an earlier onset of foraging. Our findings contribute to understanding of interacting stressors that can explain colony losses. Reduced flight capacity decreases the food-collecting ability of honeybees and may hamper the use of precocious foraging as a coping mechanism during colony (nutritional) stress. Ineffective coping mechanisms may lead to destructive cascading effects and subsequent colony collapse.