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Chagas disease is a zoonosis caused by the protozoan Trypanosoma cruzi and transmitted through the feces of triatomines, mainly in Latin America. Since the 1950s, chemical insecticides have been the primary method for controlling these triatomines, yet resistance has emerged, prompting the exploration of alternative approaches. The objective of this research was to test the capacity of the entomopathogenic nematodes Heterorhabditis indica and its symbiotic bacteria Photorhabdus luminescens, to produce mortality of Triatoma dimidiata a key vector of T. cruzi in Mexico under laboratory conditions. Two bioassays were conducted. In the first bioassay, the experimental unit was a 250 ml plastic jar with 100 g of sterile soil and three adult T. dimidiata. Three nematode quantities were tested: 2250, 4500, and 9000 nematodes per 100 g of sterile soil (n/100 g) per jar, with 3 replicates for each concentration and 1 control per concentration (1 jar with 100 g of sterile soil and 3 T. dimidiata without nematodes). The experimental unit of the second bioassay was a 500 ml plastic jar with 100 g of sterile soil and 4 adult T. dimidiata. This bioassay included 5, 50, 500, and 5000 n/100 g of sterile soil per jar, with 3 replicates of each quantity and 1 control per quantity. Data were analyzed using Kaplan-Meyer survival analysis. Electron microscopy was used to assess the presence of nematodes and tissue damage in T. dimidiata. The results of the first bioassay demonstrated that the nematode induced an accumulated average mortality ranging from 55.5 % (2250 n/100 g) to 100 % (4500 and 9000 n/100 g) within 144 h. In the second bioassay, the 5000 n/100 g concentration yielded 87.5 % mortality at 86 h, but a concentration as small as 500 n/100 g caused 75 % mortality from 84 h onwards. Survival analysis indicated higher T. dimidiata mortality with increased nematode quantities, with significant differences between the 4500, 5000, and 9000 n/100 g and controls. Electron microscopy revealed the presence of nematodes and its presumably symbiotic bacteria in the digestive system of T. dimidiata. Based on these analyses, we assert that the H. indica and P. luminescens complex causes mortality in adult T. dimidiata under laboratory conditions., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Research on the use of entomopathogenic nematodes (EPNs) as a potential tool for the biological control of invertebrates has been growing in recent years, including studies involving snails with One Health importance. In this study, the effect of exposure time (24 or 48 h) of Heterorhabditis bacteriophora HP88 on the activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), as well as the concentration of total proteins, uric acid, and urea in the hemolymph of Biomphalaria glabrata, were investigated. The concentrations of these metabolic markers were measured weekly until the end of the third week after exposure. Along with a significant reduction in total protein levels, a significant increase (p < 0.01) in uric acid and urea contents in the hemolymph of B. glabrata exposed to H. bacteriophora was observed. The accumulation of urea in these mollusks could lead to deleterious effects due to its high toxicity, inducing significant cell damage. Variations in transaminase activities were also observed, with snails exposed to EPNs showing significantly higher values (p < 0.01) than individuals in the control group, both for ALT and AST. These results indicate that experimental exposure to infective juveniles of H. bacteriophora causes significant alterations in the metabolic pattern of B. glabrata, compromising the maintenance of its homeostasis. Finally, exposure for 48 h caused more damage to the planorbid in question compared to snails exposed for 24 h, suggesting that the exposure time may influence the intensity of the host's response., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Centrioles are the core constituent of centrosomes, microtubule-organizing centers involved in directing mitotic spindle assembly and chromosome segregation in animal cells. In sexually reproducing species, centrioles degenerate during oogenesis and female meiosis is usually acentrosomal. Centrioles are retained during male meiosis and, in most species, are reintroduced with the sperm during fertilization, restoring centriole numbers in embryos. In contrast, the presence, origin, and function of centrioles in parthenogenetic species is unknown. We found that centrioles are maternally inherited in two species of asexual parthenogenetic nematodes and identified two different strategies for maternal inheritance evolved in the two species. In Rhabditophanes diutinus, centrioles organize the poles of the meiotic spindle and are inherited by both the polar body and embryo. In Disploscapter pachys, the two pairs of centrioles remain close together and are inherited by the embryo only. Our results suggest that maternally-inherited centrioles organize the embryonic spindle poles and act as a symmetry-breaking cue to induce embryo polarization. Thus, in these parthenogenetic nematodes, centrioles are maternally-inherited and functionally replace their sperm-inherited counterparts in sexually reproducing species., (© 2024. The Author(s).)

Entomopathogenic nematodes (EPNs) are closely associated with Popillia japonica and potentially used as their biological control agents, although field results proved inconsistent and evoked a continual pursuit of native EPNs more adapted to the environment. Therefore, we surveyed the Azorean Archipelago to isolate new strains of Heterorhabditis bacteriophora and to evaluate their virulence against the model organism Galleria mellonella under laboratory conditions. Six strains were obtained from pasture and coastal environments and both nematode and symbiont bacteria were molecularly identified. The bioassays revealed that Az172, Az186, and Az171 presented high virulence across the determination of a lethal dose (LD50) and short exposure time experiments with a comparable performance to Az29. After 72 hours, these virulent strains presented a mean determination of a lethal dose of 11 infective juveniles cm -2 , a lethal time (LT50) of 34 hours, and achieved 40% mortality after an initial exposure time of only 60 minutes. Az170 exhibited an intermediate performance, whereas Az179 and Az180 were classified as low virulent strains. However, both strains presented the highest reproductive potential with means of 1700 infective juveniles/mg of larvae. The bioassays of the native EPNs obtained revealed that these strains hold the potential to be used in biological control initiatives targeting P. japonica because of their high virulence and locally adapted to environmental conditions.

Phasmarhabditis (syn. Pellioditis) californica is a facultative parasite that has been marketed as a popular biocontrol agent against pestiferous slugs in England, Scotland, and Wales. The necromenic nematode Pristionchus entomophagus has also been recovered from slugs infected with Ph. californica. In this study, we experimentally investigated the outcome of single and mixed applications of Pr. entomophagus and Ph. californica on the slug Deroceras reticulatum (Müller). Host mortality was comparable for single and mixed applications of Ph. californica, with time to death significantly shorter in both treatment groups compared with controls. However, trials with Pr. entomophagus alone did not cause any significant host mortality relative to controls. Compared with the single Ph. californica applications, mixed applications resulted in 67% fewer infective juveniles establishing in the host, and subsequently far fewer infective juveniles were recovered in the next generation. In contrast, the establishment rate and progeny production in Pr. entomophagus were not impacted by the presence of Ph. californica (i.e., mixed applications). Hence, the presence of Pr. entomophagus had a deleterious effect on the establishment success and progeny production of Ph. californica. Our findings reveal an asymmetrical, antagonistic interaction between Ph. californica and Pr. entomophagus and highlight the importance of understanding the ecological relationships between co-occurring species. A decrease in parasite establishment success and progeny production has the potential to directly impact the persistence, sustainability, and efficacy of Ph. californica as a biological control agent., (© American Society of Parasitologists 2024.)

Evolutionary arms races can alter both parasite infectivity and host resistance, and it is difficult to separate the effects of these twin determinants of infection outcomes. We used a co-introduced, invasive host-parasite system (the lungworm Rhabdias pseudosphaerocephala and cane toads Rhinella marina ), where rapid adaptation and dispersal have led to population differences in infection resistance. We quantified behavioural responses of parasite larvae to skin-chemical cues of toads from different invasive populations, and rates at which juvenile hosts became infected following standardized exposure to lungworms. Chemical cues from toad skin altered host-seeking behaviour by parasites, similarly among populations. The number of infection attempts (parasite larvae entering the host's body) also did not differ between populations, but rates of successful infection (establishment of adult worm in host lungs) were higher for range-edge toads than for range-core conspecifics. Thus, lower resistance to parasite infection in range-edge juvenile toads appears to be due to less effective immune defences of the host rather than differential behavioural responses of the parasite. In this ongoing host-parasite arms race, changing outcomes appear to be driven by shifts in host immunocompetence.

The gastropod parasitic nematode Phasmarhabditis hermaphrodita has been formulated into a successful biological control agent (Nemaslug®, strain DMG0001) used to kill slugs on farms and gardens. When applied to soil, P. hermaphrodita uses slug mucus and faeces to find potential hosts. However, there is little information on what cues other species of Phasmarhabditis (P. neopapillosa and P. californica) use to find hosts and whether there is natural variation in their ability to chemotax to host cues. Therefore, using chemotaxis assays, we exposed nine wild isolates of P. hermaphrodita, five isolates of P. neopapillosa and three isolates of P. californica to mucus from the pestiferous slug host Deroceras invadens, as well as 1% and 5% hyaluronic acid - a component of slug mucus that is highly attractive to these nematodes. We found P. hermaphrodita (DMG0010) and P. californica (DMG0018) responded significantly more to D. invadens mucus and 1% hyaluronic acid than other strains. Also, P. hermaphrodita (DMG0007), P. neopapillosa (DMG0015) and P. californica (DMG0017) were superior at locating 5% hyaluronic acid compared to other isolates of the same genera. Ultimately, there is natural variation in chemoattraction in Phasmarhabditis nematodes, with some strains responding significantly better to host cues than others.

A new nematode species of the genus Phasmarhabditis was isolated from the body surface of a slug (Philomycus bilineatus Benson, PB). Morphological and molecular analyses confirmed this nematode as a new species. The nematode was named Phasmarhabditis zhejiangensis sp. nov. (Nematoda: Rhabditidae) and is dioecious. In males, the open bursa with genital papillae is characterized by the formula 1-1-1-2-1-3, and the spicule length is 58μm. In female, the vulva is located approximately in the middle of the body. The nematode belongs to papillosa group because of its tail shape pointed with filiform tip. The phasmids are rod-shaped. The posterior anus is slightly swollen. P. zhejiangensis was further characterized by internal transcribed spacer (ITS), 18S rDNA and 28S rDNA sequences. After the sequencing results were compared with sequences available from the National Center for Biotechnology Information (NCBI), the maximum similarities of ITS, 18S and 28S sequences were 89.81%, 96.22% and 95.28%, respectively. Phylogenetic analyses placed Phasmarhabditis zhejiangensis sp. nov. in the genus Phasmarhabditis., Competing Interests: No authors have competing interests The authors have declared that no competing interests exist.

Background: Pseudogamy is a reproductive system in which females rely on the sperm of males to activate their oocytes, generally parasitizing males of other species, but do not use the sperm DNA. The nematode Mesorhabditis belari uses a specific form of pseudogamy, where females produce their own males as a source of sperm. Males develop from rare eggs with true fertilization, while females arise by gynogenesis. Males thus do not contribute their genome to the female offspring. Here, we explored the diversity of reproductive mode within the Mesorhabditis genus and addressed species barriers in pseudogamous species., Results: To this end, we established a collection of over 60 Mesorhabditis strains from soil and rotting vegetal matter. We found that males from pseudogamous species displayed a reduced size of their body, male tail and sperm cells compared to males of sexual Mesorhabditis species, as expected for males that face little competition. Using rDNA sequences and crosses, we could define 11 auto-pseudogamous biological species, with closely related species pairs and a possible single origin of pseudogamy in the Mesorhabditis genus. Most crosses between males and females of different species did not even produce female progeny. This surprising species barrier in pseudogamous egg activation was pre or postcopulatory depending on the species pair. In the latter case, when hybrid embryos were produced, most arrested before the first embryonic cell division. Hybrid incompatibility between auto-pseudogamous species was due to defective interaction between sperm and oocyte as well as defective reconstitution of zygotic centrosomes., Conclusions: We established a collection of sexual and pseudo-sexual species which offer an ideal framework to explore the origin and consequences of transition to asexuality. Our results demonstrate that speciation occurs in the pseudogamous state. Whereas genomic conflicts are responsible for hybrid incompatibility in sexual species, we here reveal that centrosomes constitute key organelles in the establishment of species barrier.

Different model systems have, over the years, contributed to our current understanding of the molecular mechanisms underpinning the various types of interaction between bacteria and their animal hosts. The genus Photorhabdus comprises Gram-negative insect pathogenic bacteria that are normally found as symbionts that colonize the gut of the infective juvenile stage of soil-dwelling nematodes from the family Heterorhabditis . The nematodes infect susceptible insects and release the bacteria into the insect haemolymph where the bacteria grow, resulting in the death of the insect. At this stage the nematodes feed on the bacterial biomass and, following several rounds of reproduction, the nematodes develop into infective juveniles that leave the insect cadaver in search of new hosts. Therefore Photorhabdus has three distinct and obligate roles to play during this life-cycle: (1) Photorhabdus must kill the insect host; (2) Photorhabdus must be capable of supporting nematode growth and development; and (3) Photorhabdus must be able to colonize the gut of the next generation of infective juveniles before they leave the insect cadaver. In this review I will discuss how genetic analysis has identified key genes involved in mediating, and regulating, the interaction between Photorhabdus and each of its invertebrate hosts. These studies have resulted in the characterization of several new families of toxins and a novel inter-kingdom signalling molecule and have also uncovered an important role for phase variation in the regulation of these different roles.

In insects, diet plays an important role in growth and development. Insects can vary their diet composition based on their physiological needs. In this study we tested the influence of diet composition involving varying concentrations of macronutrients and zinc on the immune-tolerance following parasite and pathogen exposure in Spodoptera litura larvae. We also tested the insecticidal potential of Mesorhabditis belari, Enterobacter hormaechei and its secondary metabolites on Spodoptera litura larvae. The results shows macronutrient composition does not directly affect the larval tolerance to nematode infection. However, Zinc supplemented diet improved the immune tolerance. While larvae exposed to bacterial infection performed better on carbohydrate rich diet. Secondary metabolites from bacteria produced an immune response in dose dependent mortality. The study shows that the larvae maintained on different diet composition show varied immune tolerance which is based on the type of infection., (Copyright © 2019 Elsevier Inc. All rights reserved.)

We report the reproductive strategy of the nematode Mesorhabditis belari This species produces only 9% males, whose sperm is necessary to fertilize and activate the eggs. However, most of the fertilized eggs develop without using the sperm DNA and produce female individuals. Only in 9% of eggs is the male DNA utilized, producing sons. We found that mixing of parental genomes only gives rise to males because the Y-bearing sperm of males are much more competent than the X-bearing sperm for penetrating the eggs. In this previously unrecognized strategy, asexual females produce few sexual males whose genes never reenter the female pool. Here, production of males is of interest only if sons are more likely to mate with their sisters. Using game theory, we show that in this context, the production of 9% males by M. belari females is an evolutionary stable strategy., (Copyright © 2019 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

The Spanish Slug (Arion vulgaris, formerly known as Arion lusitanicus) is an invasive agricultural and horticultural pest species that causes great damages all over Europe. Numerous options to control this slug are on the market; among the most commonly used are slug pellets containing the active ingredients metaldehyde or iron-III-phosphate and the application of parasitic nematodes (Phasmarhabditis hermaphrodita). These control measures potentially also affect non-target organisms like earthworms (Lumbricidae), which themselves can directly and/or indirectly alter a plant's susceptibility against slug herbivory. Also, the efficacy of slug control treatments is expected to be influenced by watering. In a greenhouse experiment we investigated the influence of daily watering vs. every third day watering on slug control efficacy and potential interactions with earthworms. We found significant interactions between watering and slug control efficacy. Slug herbivory and biomass decreased after application of slug pellets; metaldehyde was more effective under less frequent watering while iron-III-phosphate was unaffected by watering. Parasitic nematodes had no effect on slug herbivory and biomass production. Earthworm activity was reduced with less frequent watering but did not interact with slug control. We conclude that watering patterns should be considered when choosing slug control measures.

Phasmarhabditis hermaphrodita is a parasitic nematode of terrestrial gastropods that has been formulated into a biological control agent for farmers and gardeners to kill slugs and snails. In order to locate slugs it is attracted to mucus, faeces and volatile cues; however, there is no information about whether these nematodes are attracted to snail cues. It is also unknown how wild isolates of P. hermaphrodita or different Phasmarhabditis species behave when exposed to gastropod cues. Therefore, we investigated whether P. hermaphrodita (commercial and wild isolated strains), P. neopapillosa and P. californica were attracted to mucus from several common snail species (Cepaea nemoralis, Cepaea hortensis, Arianta arbustorum and Cornu aspersum). We also examined whether snails (C. aspersum) collected from different locations around the UK differed in their attractiveness to wild isolates of P. hermaphrodita. Furthermore, we also investigated what properties of snail mucus the nematodes were attracted to, including hyaluronic acid and metal salts (FeSO4, ZnSO4, CuSO4 and MgSO4). We found that the commercial strain of P. hermaphrodita responded poorly to snail mucus compared to wild isolated strains, and C. aspersum collected from different parts of the UK differed in their attractiveness to the nematodes. We found that Phasmarhabditis nematodes were weakly attracted to all metals tested but were strongly attracted to hyaluronic acid. In a final experiment we also showed that pharmacological manipulation of cyclic guanosine monophosphate (cGMP) increased chemoattraction to snail mucus, suggesting that the protein kinase EGL-4 may be responsible for Phasmarhabditis sp. chemoattraction.

Coexistence of highly similar species is at odds with ecological theory of competition; coexistence, then, requires stabilizing mechanisms such as differences in ecological niche. In the bacterivore nematode Litoditis marina species complex, which occurs associated with macro-algae, four cryptic lineages (Pm I-IV) co-occur in the field along the south-western coast and estuaries of The Netherlands. Here we investigate the temporal and/or spatial niche differentiation in their natural environment using a qPCR-based detection and relative quantification method. We collected different algal species (i.e. two Fucus species and Ulva sp.) and separated algal structures (i.e. receptacula, thalli, non-fertile tips and bladders) at different sampling months and times (i.e. twice per sampling month), to examine differences in microhabitat use between coexisting L. marina species. Results demonstrate that the cryptic species composition varied among different algal species and algal structures, which was also subject to temporal shifts. Pm I dominated on Fucus spp., Pm II showed dominance on Ulva sp., while Pm III overall had the lowest frequencies. Microhabitat partitioning was most pronounced between the two cryptic species which had similar microbiomes (Pm I and Pm II), and less so between the two species which had significantly different microbiomes (Pm I and Pm III), suggesting that species which share the same microhabitats may avoid competition through resource partitioning. The interplay of microhabitat differentiation and temporal dynamics among the cryptic species of L. marina implies that there is a complex interaction between biotic components and abiotic factors which contributes to their coexistence in the field., Competing Interests: The authors have declared that no competing interests exist.

Interest has recently grown in developing the entomopathogenic nematode Heterorhabditis bacteriophora as a model to genetically dissect the process of parasitic infection. Despite the availability of a full genome assembly, there is substantial variation in gene model accuracy. Here, a methodology is presented for leveraging RNA-seq evidence to generate improved annotations using ab initio gene prediction software. After alignment of reads and subsequent generation of a RNA-seq supported annotation, the new gene prediction models were verified on a selection of genes by comparison with sequenced 5' and 3' rapid amplification of cDNA ends products. By utilising a whole transcriptome for genome annotation, the current reference annotation was enriched, demonstrating the importance of coupling transcriptional data with genome assemblies., (Copyright © 2018 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.)

Three key steps in meiosis allow diploid organisms to produce haploid gametes: (1) homologous chromosomes (homologs) pair and undergo crossovers; (2) homologs segregate to opposite poles; and (3) sister chromatids segregate to opposite poles. The XX/XO sex determination system found in many nematodes [1] facilitates the study of meiosis because variation is easily recognized [2-4]. Here we show that meiotic segregation of X chromosomes in the trioecious nematode Auanema rhodensis [5] varies according to sex (hermaphrodite, female, or male) and type of gametogenesis (oogenesis or spermatogenesis). In this species, XO males exclusively produce X-bearing sperm [6, 7]. The unpaired X precociously separates into sister chromatids, which co-segregate with the autosome set to generate a functional haplo-X sperm. The other set of autosomes is discarded into a residual body. Here we explore the X chromosome behavior in female and hermaphrodite meioses. Whereas X chromosomes segregate following the canonical pattern during XX female oogenesis to yield haplo-X oocytes, during XX hermaphrodite oogenesis they segregate to the first polar body to yield nullo-X oocytes. Thus, crosses between XX hermaphrodites and males yield exclusively male progeny. During hermaphrodite spermatogenesis, the sister chromatids of the X chromosomes separate during meiosis I, and homologous X chromatids segregate to the functional sperm to create diplo-X sperm. Given these intra-species, intra-individual, and intra-gametogenesis variations in the meiotic program, A. rhodensis is an ideal model for studying the plasticity of meiosis and how it can be modulated., (Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The nervous system controls most rhythmic behaviors, with a remarkable exception. In Caenorhabditis elegans periodic defecation rhythm does not appear to involve the nervous system. Such oscillations are studied in detail with genetic and molecular biology tools. The small size of C. elegans cells impairs the use of standard electrophysiological methods. We studied a similar rhythmic pacemaker in the noticeably larger gut cells of Heterorhabditis megidis nematode. H. megidis defecation cycle is driven by a central pattern generator (CPG) associated with unusual all-or-none hyper-polarization "action potential". The CPG cycle period depends on the membrane potential and CPG cycling also persisted in experiments where the membrane potential of gut cells was continuously clamped at steady voltage levels. The usual excitable tissue description does not include the endoderm or imply the generation of hyper-polarization spikes. The nematode gut cells activity calls for a reevaluation of the excitable cells definition.

Entomopathogenic nematodes in the Heterorhabditis genus and their symbiotic Photorhabdus bacteria are important biocontrol agents of insect pests and models for the study of microbe-host interactions. In this work, we used larvae of the tobacco budworm (Heliothis virescens) as a model to study its defensive mechanisms against Heterorhabditis bacteriophora nematodes carrying symbiotic Photorhabdus temperata. We first determined time points of initial nematode entry and release of bacteria into the haemolymph to perform transcriptional analysis of insect gene expression during these steps in the infective process. RNA-Sequencing analyses were then performed to profile differential gene expression in the insect during nematode invasion, bacterial release and final steps of infection, relative to the untreated controls. Our results support the theory that insect immune response genes are induced upon nematode invasion, but the majority of these genes are suppressed upon the release of bacteria by the nematodes into the haemolymph. Overall, these findings provide information on the dynamics of the insect's response to a progressing infection by this entomopathogenic nematode-bacteria complex and facilitate development of Hel. virescens as a pest model for future functional studies of the key insect defence factors., (© 2017 The Royal Entomological Society.)

Background: Aedes aegypti is a potential vector of West Nile, Japanese encephalitis, chikungunya, dengue and Zika viruses. Alternative control measurements of the vector are needed to overcome the problems of environmental contamination and chemical resistance. Xenorhabdus and Photorhabdus are symbionts in the intestine of entomopathogenic nematodes (EPNs) Steinernema spp. and Heterorhabditis spp. These bacteria are able to produce a broad range of bioactive compounds including antimicrobial, antiparasitic, cytotoxic and insecticidal compounds. The objectives of this study were to identify Xenorhabdus and Photorhabdus isolated from EPNs in upper northern Thailand and to study their larvicidal activity against Ae. aegypti larvae., Results: A total of 60 isolates of symbiotic bacteria isolated from EPNs consisted of Xenorhabdus (32 isolates) and Photorhabdus (28 isolates). Based on recA gene sequencing, BLASTN and phylogenetic analysis, 27 isolates of Xenorhabdus were identical and closely related to X. stockiae, 4 isolates were identical to X. miraniensis, and one isolate was identical to X. ehlersii. Twenty-seven isolates of Photorhabdus were closely related to P. luminescens akhurstii and P. luminescens hainanensis, and only one isolate was identical and closely related to P. luminescens laumondii. Xenorhabdus and Photorhabdus were lethal to Ae aegypti larvae. Xenorhabdus ehlersii bMH9.2&#95;TH showed 100% efficiency for killing larvae of both fed and unfed conditions, the highest for control of Ae. aegypti larvae and X. stockiae (bLPA18.4&#95;TH) was likely to be effective in killing Ae. aegypti larvae given the mortality rates above 60% at 72 h and 96 h., Conclusions: The common species in the study area are X. stockiae, P. luminescens akhurstii, and P. luminescens hainanensis. Three symbiotic associations identified included P. luminescens akhurstii-H. gerrardi, P. luminescens hainanensis-H. gerrardi and X. ehlersii-S. Scarabaei which are new observations of importance to our knowledge of the biodiversity of, and relationships between, EPNs and their symbiotic bacteria. Based on the biological assay, X. ehlersii bMH9.2&#95;TH begins to kill Ae. aegypti larvae within 48 h and has the most potential as a pathogen to the larvae. These data indicate that X. ehlersii may be an alternative biological control agent for Ae. aegypti and other mosquitoes.

Terrestrial molluscs (Mollusca: Gastropoda) are important economic pests worldwide, causing extensive damage to a variety of crop types, and posing a health risk to both humans and wildlife. Current knowledge indicates that there are eight nematode families that associate with molluscs as definitive hosts, including Agfidae, Alaninematidae, Alloionematidae, Angiostomatidae, Cosmocercidae, Diplogastridae, Mermithidae and Rhabditidae. To date, Phasmarhabditis hermaphrodita (Schneider, 1859) Andrássy, 1983 (Rhabditida: Rhabditidae) is the only nematode that has been developed as a biological molluscicide. The nematode, which was commercially released in 1994 by MicroBio Ltd, Littlehampton, UK (formally Becker Underwood, now BASF) under the tradename Nemaslug®, is now sold in 15 different European countries. This paper reviews nematodes isolated from molluscs, with specially detailed information on the life cycle, host range, commercialization, natural distribution, mass production and field application of P. hermaphrodita.

This study investigated the possible biological, biochemical and histological changes in Bradybaena similaris(Gastropoda: Pulmonata) infected by Heterorhabditis indica (Rhabditida: Heterorhabditidae), strain LPP1. Two groups of 16 snails were formed: the control group (unexposed) and the treated group, which was exposed for three weeks to infective juveniles (J 3 ) of H. indica LPP1. The experiment was conducted in duplicate, using a total of 64 snails. After the exposure period, the snails were dissected to collect the hemolymph and tissues, for evaluation of the physiological changes caused by the infection. The number of eggs laid/snail and the viability of these eggs were also assessed as indicators of the reproductive activity of B. similaris. Intense glycogenolysis was accompanied by a significant reduction (p < 0.05) in the glucose content of the hemolymph of the exposed snails, indicating that infection by H. indica induces breakdown of the host's glycemic homeostasis. Significant variations (p < 0.05) in the lactate dehydrogenase activity occurred together with changes in the concentration of pyruvic and lactic acid in the hemolymph of the infected B. similaris snails, corroborating the transition from aerobic to anaerobic metabolism in the hosts. These metabolic alterations reflect the parasitic castration process in this interface. The results suggest that the use of H. indica LPP1 is a potential alternative for biological control of B. similaris., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Entomopathogenic nematodes (EPN) belonging to the Heterorhabditidae family are lethal parasites of soil-dwelling insects. Two species were reported in Argentina: Heterorhabditis argentinensis and Heterorhabditis bacteriophora characterized mainly by morphometric features. In this work a comparative and phylogenetic study between five Heterorhabditis populations from Argentina was conducted to analyze the variability between strains and to evaluate the taxonomic position of Heterorhabditis argentinensis. The PCA analyses of morphometric characters separated the larger juvenile, female and male H. argentinensis from H. bacteriophora populations. The juvenile (IJs) stage provided the clearest separation of Heterorhabditis populations presenting the least variability between strains. The variable L and MBW were highly related to H. argentinensis IJs. Three groups were separated by this stage considering PC1 and PC2: one formed by H. bacteriophora OLI, RIV and RN strains, (isolates from Córdoba and Río Negro province), one for H. bacteriophora VELI strain (Buenos Aires province) and one for H. argentinensis (Santa Fe province). Heterorhabditis bacteriophora VELI and H. argentinensis isolated from regions with more rainfalls and humidity presented larger values for morphometric features. Molecular analyses showed the Argentinian populations (H. bacteriophora VELI strain and H. argentinensis), forming a same clade, with six other H. bacteriophora populations (not from Argentina) with a genetic similarity between them of 99%. Heterorhabditis argentinensis presented one unique nucleotide that was not present in any of the other species of the clade. Considering the results of this study H. argentinensis would be conspecific to H. bacteriophora, constituting a strain with a great morphometric variation where the host and climatic conditions could have influenced on the measurements.

Entomopathogenic nematodes (EPNs) of the genera Steinernema and Heterorhabditis are lethal parasites of many insect species. To investigate defensive mechanisms towards EPNs in relation to antioxidative and detoxifying enzymes, we chose Tenebrio molitor (Coleoptera: Tenebrionidae) as experimental insect. We studied the activity changes of superoxide dismutases (SODs), peroxidases (PODs), and catalases (CATs), as well as tyrosinase (TYR), acetylcholinesterase (AChE), carboxylesterase (CarE), and glutathione S-transferase (GSTs) for 40 h in T. molitor larvae infected with Heterorhabditis beicherriana infective juveniles (IJs) at 5 rates (0, 20, 40, 80, and 160 IJs/larva). We found that when T. molitor larvae infected with H. beicherriana at higher rates (80 and 160 IJs/larva), SOD activity quickly increased to more than 70 % higher than that control levels. The activities of POD and CAT increased after 24 h. TYR activity increased slowly at lower rates of infection for 16 h, followed by a slight decrease, and then increasing from 32 to 40 h. The other detoxifying enzymes (GST, CarE, and AChE) were enhanced at lower infection rates, but were inhibited at higher rates. Our results suggested that host antioxidative response and detoxification reactions played a central role in the defensive reaction to EPNs, and that this stress which was reflected by the higher level enzymes activity contributed to the death of hosts. Further study should explore the exact function of these enzymes using different species of EPNs and investigate the links between enzyme activity and host susceptibility to EPNs.

Photorhabdus luminescens subsp. laumondii is closely associated with the entomopathogenic nematode Heterorhabditis bacteriophora and has, to date, not been isolated from other nematode species. This study is the first report of P. luminescens subsp. laumondii from two South African isolates of entomopathogenic nematodes, Heterorhabditis safricana SF281 and H. bacteriophora SF351. Both symbiotic bacterial strains are phenotypically closely related to P. luminescens subsp. laumondii previously isolated and described from H. bacteriophora. The genetic relatedness between P. luminescens subsp. laumondii strains SF281B and SF351B was confirmed by comparing 16S rDNA, recA, gyrB and gltX sequences with sequences of P. luminescens subsp. laumondii, including the type strain (TT01 T ) and strain E21.

Bradysia odoriphaga Yang & Zhang (Diptera: Sciaridae) is the most serious pest of Chinese chive, Allium tuberosum Chemical pesticide application has become a necessary step to control B. odoriphaga after each of the four to six harvests during the growing season. We compared the effects of application type (nematode-infected cadaver, aqueous nematode suspension, and mixture of cadaver and aqueous suspension) and nematode species (Heterorhabditis bacteriophora vs. Steinernema carpocapsae) on B. odoriphaga control. Nematode species combinations and the use of the cadaver method has only been tested in a relatively few studies, and has not been tested for this target pest, B. odoriphaga. Furthermore, this is the first report of combining application methods (aqueous and cadaver). Results indicated that the cadaver treatment produced higher mortality of B. odoriphaga than the aqueous treatment, and H. bacteriophora caused higher mortality of B. odoriphaga than S. carpocapsae. The mortality of B. odoriphaga was 96.7% in H. bacteriophora cadaver treatment and only 27.5% in control treatment without nematode addition. Cadaver treatments caused higher biomass of A. tuberosum than that of aqueous treatment. The plant biomass in H. bacteriophora cadaver treatment was 10 folds as much as that in the control treatment. The mixed aqueous suspension of H. bacteriophora and S. carpocapsae did not increase the mortality in pot trial. Our findings indicate that entomopathogenic nematodes applied as cadavers might be a potential alternative biological agent in the integrated management of B. odoriphaga for Chinese chive production., (© The Authors 2016. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

The crucifer flea beetle, Phyllotreta cruciferae (Goeze), is a key pest of canola (Brassica napus L.) in the northern Great Plains of North America. The efficacies of entomopathogenic nematodes (Steinernema spp. and Heterorhabditis spp.), a sprayable polymer gel, and a combination of both were assessed on canola for flea beetle management. Plots were treated soon after colonization by adult flea beetles, when canola was in the cotyledon to one-leaf stage. Ten plants along a 3.6-m section of row were selected and rated at pre-treatment and 7 and 14 d post treatment using the damage-rating scheme advanced by the European Plant Protection Organization, where 1 = 0%, 2 = 2%, 3 = 5%, 4 = 10%, and 5 = 25% leaf area injury. Under moderate flea beetle feeding pressure (1-3.3% leaf area damaged), seeds treated with Gaucho 600 (Bayer CropScience LP Raleigh, NC) (imidacloprid) produced the highest yield (843.2 kg/ha). Meanwhile, Barricade (Barricade International, Inc. Hobe Sound, FL) (polymer gel; 1%) + Scanmask (BioLogic Company Inc, Willow Hill, PA) (Steinernema feltiae) resulted in the highest yields: 1020.8 kg/ha under high (2.0-5.3% leaf area damaged), and 670.2 kg/ha at extremely high (4.3-8.6 % leaf area damaged) feeding pressure. Our results suggest that Barricade (1%) + Scanmask (S. feltiae) can serve as an alternative to the conventional chemical seed treatment. Moreover, Scanmask (S. feltiae) can be used to complement the effects of seed treatment after its protection has run out., (© The Authors 2016. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

The microhabitats in which hosts live can potentially influence the ability and success of parasites in finding and infecting these hosts. The infection dynamics of both digenetic trematode parasites and a nematode parasite (Daubaylia potomaca) infecting a pulmonate snail, Helisoma anceps , were observed in a small North Carolina lake using 3 different classifications of substratum type based on percent coverage by leaves and debris. There were no differences in snail site occupancy or density between substratum types, but small-scale differences in microhabitat impacted parasite prevalence and intensity in their gastropod hosts. Snails inhabiting substrata covered in more leaf litter and debris had a lower prevalence and intensity of infection by all of the trematode species and life stages when compared to H. anceps inhabiting other substrata types, while only the intensity of infection was impacted in D. potomaca infections. These findings emphasize the importance of microhabitat, specifically its physical components, in influencing parasite infection in intermediate hosts and that small-scale differences may significantly affect the patterns of infection.

Certain species of entomopathogenic nematodes, such as Heterorhabditis indica Poinar, Karunakar & David, have the potential to be effective controls for Aethina tumida (Murray), or small hive beetles, when applied to the soil surrounding honey bee (Apis mellifera L.) hives. Despite the efficacy of H. indica, beekeepers have struggled to use them successfully as a biocontrol. It is believed that the sensitivity of H. indica to certain environmental conditions is the primary reason for this lack of success. Although research has been conducted to explore the impact of specific environmental conditions--such as soil moisture or soil temperature-on entomopathogenic nematode infectivity, no study to date has taken a comprehensive approach that considers the impact of multiple environmental conditions simultaneously. In exploring this, a multivariate logistic regression model was used to determine what environmental conditions resulted in reductions of A. tumida populations in honey bee colonies. To obtain the sample sizes necessary to run a multivariate logistic regression, this study utilized citizen scientist beekeepers and their hives from across the mid-Atlantic region of the United States. Results suggest that soil moisture, soil temperatures, sunlight exposure, and groundcover contribute to the efficacy of H. indica in reducing A. tumida populations in A. mellifera colonies. The results of this study offer direction for future research on the environmental preferences of H. indica and can be used to educate beekeepers about methods for better utilizing H. indica as a biological control., (© The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Nematodes have diverse reproductive strategies, which make them ideal subjects for comparative studies to address how mating systems evolve. Here we present the sex ratios and mating dynamics of the free-living nematode Rhabditis sp. SB347, in which males, females and hermaphrodites co-exist. The three sexes are produced by both selfing and outcrossing, and females tend to appear early in a mother's progeny. Males prefer mating with females over hermaphrodites, which our results suggest is related to the female-specific production of the sex pheromones ascr#1 and ascr#9. We discuss the parallels between this system and that of parasitic nematodes that exhibit alternation between uniparental and biparental reproduction.

Root exudates can play an important role in plant-nematode interactions. Recent studies have shown that the root cap exudates obtained from several plant species trigger a state of dormancy or quiescence in various genera of nematodes. This phenomenon is not only of fundamental ecological interest, but also has application potential if the plant-produced compound(s) could be used to control harmful nematodes or help to prolong the shelf-life of beneficial entomopathogenic nematodes (EPNs). The identification of the compound(s) involved in quiescence induction has proven to be a major challenge and requires large amounts of active material. Here, we present a high-throughput method to obtain bioactive root extracts from flash-frozen root caps of green pea and maize. The root cap extract obtained via this method was considerably more potent in inducing quiescence than exudate obtained by a previously used method, and consistently induced quiescence in the EPN Heterorhabditis megidis, even after a 30-fold dilution. Extracts obtained from the rest of the root were equally effective in inducing quiescence. Infective juveniles (IJs) of H. megidis exposed to these extracts readily recovered from their quiescent state as soon as they were placed in moist soil, and they were at least as infectious as the IJs that had been stored in water. Excessive exposure of IJs to air interfered with the triggering of quiescence. The implications of these results and the next steps towards identification of the quiescence-inducing compound(s) are discussed from the perspective of applying EPN against soil-dwelling insect pests.

In addition to an increase in mean temperature, climate change models predict decreasing amplitudes of daily temperature fluctuations. In temperate regions, where daily and seasonal fluctuations are prominent, such decreases in daily temperature fluctuations can have a pronounced effect on the fitness of species and on the outcome of species interactions. In this study, the effect of a temperature regime with daily fluctuations versus a constant temperature on the fitness and interspecific interactions of three cryptic species of the marine nematode species complex of Litoditis marina (Pm I, Pm III and Pm IV) were investigated. In a lab experiment, different combinations of species (monospecific treatment: Pm I and Pm IV and Pm III alone; two-species treatment: Pm I + Pm IV; three-species treatment: Pm I + Pm IV + Pm III) were subjected to two different temperature regimes: one constant and one fluctuating temperature. Our results showed that fluctuating temperature had minor or no effects on the population fitness of the three species in monocultures. In contrast, interspecific interactions clearly influenced the fitness of all three species, both positively and negatively. Temperature regime did have a substantial effect on the interactions between the species. In the two-species treatment, temperature regime altered the interaction from a sort of mutualism to commensalism. In addition, the strength of the interspecific interactions changed depending on the temperature regime in the three-species treatment. This experiment confirms that interactions between the species can change depending on the abiotic environment; these results show that it is important to incorporate the effect of fluctuations on interspecific interactions to predict the effect of climate change on biodiversity.

Entomopathogenic nematodes have been classified into cruisers (active searchers) and ambushers (sit and wait foragers). However, little is known about their dispersal and foraging behavior at population level in soil. We studied lateral dispersal of the ambush foraging Steinernema carpocapsae (ALL strain) and cruise foraging Heterorhabditis bacteriophora (GPS11 strain) from infected host cadavers in microcosms (0.05 m2) containing Wooster silt-loam soil (Oxyaquic fragiudalf) and vegetation in the presence or absence of non-mobile and mobile hosts. Results showed that the presence of a non-mobile host (Galleria mellonella larva in a wire mesh cage) enhanced H. bacteriophora dispersal for up to 24 hr compared with no-host treatment, but had no impact on S. carpocapsae dispersal. In contrast, presence of a mobile host (G. mellonella larvae) increased dispersal of S. carpocapsae compared with no host treatment, but had no effect on H. bacteriophora dispersal. Also H. bacteriophora was better at infecting non-mobile than mobile hosts released into the microcosms and S. carpocapsae was better at infecting mobile than non-mobile hosts, thus affirming the established cruiser-ambusher theory. However, results also revealed that a large proportion of infective juveniles (IJs) of both species stayed near (≤ 3.8 cm) the source cadaver (88-96% S. carpocapsae; 67-79% H. bacteriophora), and the proportion of IJs reaching the farthest distance (11.4 cm) was significantly higher for S. carpocapsae (1.4%) than H. bacteriophora (0.4%) in the presence of mobile hosts. S. carpocapsae also had higher average population displacement than H. bacteriophora in the presence of both the non-mobile (5.07 vs. 3.6 cm/day) and mobile (8.06 vs. 5.3 cm/day) hosts. We conclude that the two species differ in their dispersal and foraging behavior at the population level and this behavior is affected by both the presence and absence of hosts and by their mobility.

Biological invasions can bring both the invader and native taxa into contact with novel parasites. As cane toads ( Rhinella marina ) have spread through Australia, they have encountered lungworms (Rhabdias hylae) that occur in native frogs. Field surveys suggest that these lungworms have not host-switched to toads. In our laboratory studies, R. hylae infected cane toads as readily as it infected native frogs, but failed to reach the lungs of the novel host (i.e., were killed by the toads' immune response). Plausibly, then, R. hylae might reduce the viability both of their native hosts (frogs, that can exhibit high parasite burdens) and cane toads (that must deal with infective larvae traveling through the host body). Our laboratory trials suggest, however, that the impacts of the parasite on infected anuran hosts (both frogs and toads) were minimal, with no significant decrements to host survival, activity, growth, or locomotor performance. Ironically, the lack of impact of the parasite on its native hosts appears to be an outcome of co-evolution (frogs tolerate the lungworm), whereas the lack of impact on the novel host is due to a lack of co-evolution (toads can recognize and eliminate the lungworm).

Behavioural innovation and morphological adaptation are intrinsically linked but their relationship is often poorly understood. In nematodes, a huge diversity of feeding morphologies and behaviours can be observed to meet their distinctive dietary and environmental demands. Pristionchus and their relatives show varied feeding activities, both consuming bacteria and also predating other nematodes. In addition, Pristionchus nematodes display dimorphic mouth structures triggered by an irreversible developmental switch, which generates a narrower mouthed form with a single tooth and a wider mouthed form with an additional tooth. However, little is known about the specific predatory adaptations of these mouth forms or the associated mechanisms and behaviours. Through a mechanistic analysis of predation behaviours, in particular in the model organism Pristionchus pacificus, we reveal multifaceted feeding modes characterised by dynamic rhythmic switching and tooth stimulation. This complex feeding mode switch is regulated by the neurotransmitter serotonin in a previously uncharacterised role, a process that appears conserved across several predatory nematode species. Furthermore, we investigated the effects of starvation, prey size and prey preference on P. pacificus predatory feeding kinetics, revealing predation to be a fundamental component of the P. pacificus feeding repertoire, thus providing an additional rich source of nutrition in addition to bacteria. Finally, we found that mouth form morphology also has a striking impact on predation, suppressing predatory behaviour in the narrow mouthed form. Our results therefore hint at the regulatory networks involved in controlling predatory feeding and underscore P. pacificus as a model for understanding the evolution of complex behaviours., (© 2015. Published by The Company of Biologists Ltd.)

The citrus mealybug, Planococcus citri, is a highly destructive pest of citrus, occurring only in the aerial parts of plants. Humidity will be one of the key factors to consider when using entomopathogenic nematodes (EPN) as biological control agents. Different adjuvants can be added to suspensions of EPNs, to improve control as a foliar application. An aqueous suspension containing Heterorhabditis zealandica and 0.3% Zeba® significantly increased P. citri mortality by 22% at 80% relative humidity (RH) with a temperature cycle starting at 22°C for 14 h and 11°C for 11 h. The same polymer formulation was tested for Steinernema yirgalemense and mortality of P. citri increased by 21% at 60% RH and by 27% at 80% RH. The addition of Nu-Film-P® and Zeba® to H. zealandica suspensions did not significantly retard application run-off on citrus leaves. The combination of Nu-Film-P® and Zeba®, however, was able to significantly retard sedimentation, increasing the average number of nematodes deposited on 2-cm2 leaf discs by 10 nematodes. In an aqueous suspension, nematodes settle rapidly to the bottom, resulting in an uneven distribution of nematodes. Xanthan gum, at a concentration of 0.2%, was highly effective at retarding sedimentation, with 72% of the initial nematode number still in suspension after 1 h. Zeba®, at a concentration of 0.3%, despite not being as effective as Xanthan gum, nevertheless still retarded sedimentation significantly. This is the first report of the potential of Nu-Film-P® and Zeba® to improve EPN performance against P. citri when used above ground in citrus orchards.

Parasites that are carried by invasive species can infect native taxa, with devastating consequences. In Australia, invading cane toads (Rhinella marina) carry lungworm parasites (Rhabdias pseudosphaerocephala) that (based on previous laboratory studies) can infect native treefrogs (Litoria caerulea and L. splendida). To assess the potential of parasite transmission from the invader to the native species (and from one infected native frog to another), we used surveys and radiotelemetry to quantify anuran microhabitat use, and proximity to other anurans, in two sites in tropical Australia. Unsurprisingly, treefrogs spent much of their time off the ground (especially by day, and in undisturbed forests) but terrestrial activity was common at night (especially in anthropogenically modified habitats). Microhabitat overlap between cane toads and frogs was generally low, except at night in disturbed areas, whereas overlap between the two frog species was high. The situations of highest overlap, and hence with the greatest danger of parasite transmission, involve aggregations of frogs within crevices by day, and use of open ground by all three anuran species at night. Overall, microhabitat divergence between toads and frogs should reduce, but not eliminate, the transmission of lungworms from invasive toads to vulnerable native frogs.

Ambush foragers must employ a long-distance dispersal strategy to maximize reproductive success in the absence of hosts. This hypothesis was tested by comparing lateral dispersal of the ambusher, Steinernema carpocapsae , and the cruiser, Heterorhabditis bacteriophora , nematodes from infected host cadavers in autoclaved, silt-loam soil in large microcosms (0.05-1.5 m(2)) with or without vegetation in the absence of hosts. Dispersal was estimated by taking soil cores (5 × 2 cm in diameter) from the microcosms at different intervals (6-240 hr) and distances (3.8-61 cm) from the infected host cadavers and baiting with Galleria mellonella larvae. The numbers of baited larvae killed and the numbers of infective juveniles (IJs) penetrated in dead baits were counted to compute the percentage of IJs dispersed from the source cadavers, based on the emergence potential and penetration efficiency of the 2 species, and analyzed. Vegetation enhanced dispersal of both species but more so for H. bacteriophora . Although the pattern of dispersal differed spatio-temporally for the 2 species, average population displacement was similar (∼6 cm/day). A majority of the S. carpocapsae population ambushed close to the source cadaver (<3.8 cm), whereas a majority of H. bacteriophora population dispersed between 7-12 cm away from the source cadaver. About 4% of the S. carpocapsae population dispersed faster than the fastest H. bacteriophora , reaching 30-61 cm, compared to only 2% of the H. bacteriophora population dispersing this far. This use of 'sprinters' for long-distance dispersal may represent an adaptive dispersal strategy by the otherwise ambush forager S. carpocapsae in the absence of hosts.

Rhabdiasidae Railliet, 1915 is a globally distributed group of up to 100 known species of nematodes parasitic in amphibians and reptiles. This work presents the results of a molecular phylogenetic analysis of 36 species of Rhabdiasidae from reptiles and amphibians from six continents. New DNA sequences encompassing partial 18S rDNA, ITS1, 5.8S rDNA, ITS2 and partial 28S rDNA regions of nuclear ribosomal DNA were obtained from 27 species and pre-existing sequences for nine species were incorporated. The broad taxonomic, host and geographical coverage of the specimens allowed us to address long-standing questions in rhabdiasid systematics, evolution, geographic distribution, and patterns of host association. Our analysis demonstrated that rhabdiasids parasitic in snakes are an independent genus sister to the rest of the Rhabdiasidae, a status supported by life cycle data. Based on the combined evidence of molecular phylogeny, morphology and life cycle characteristics, a new genus Serpentirhabdias gen. nov. with the type species Serpentirhabdias elaphe (Sharpilo, 1976) comb. nov. is established. The phylogeny supports the monophyly of Entomelas Travassos, 1930, Pneumonema Johnston, 1916 and the largest genus of the family, Rhabdias Stiles and Hassall, 1905. DNA sequence comparisons demonstrate the presence of more than one species in the previously monotypic Pneumonema from Australian scincid lizards. The distribution of some morphological characters in the genus Rhabdias shows little consistency within the phylogenetic tree topology, in particular the apical structures widely used in rhabdiasid systematics. Our data suggest that some of the characters, while valuable for species differentiation, are not appropriate for differentiation among higher taxa and are of limited phylogenetic utility. Rhabdias is the only genus with a cosmopolitan distribution, but some of the lineages within Rhabdias are distributed on a single continent or a group of adjacent zoogeographical regions. Serpentirhabdias, Entomelas and Pneumonema show rather strict specificity to their host groups. The evolution of the Rhabdiasidae clearly included multiple host switching events among different orders and families of amphibians as well as switching between amphibians and squamatan reptiles. Only a few smaller lineages of Rhabdias demonstrate relatively strict associations with a certain group of hosts., (Copyright © 2014 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.)

Heterorhabditis is a nematode found in the soil that is used as an important biological control agent against various organisms. However, few studies have been performed of its use against snails and the present study is the first to investigate the effect of experimental exposure of Bradybaena similaris to Heterorhabditis indica LPP1. Two groups of 16 snails were formed: the control group (not exposed) and the treatment, which was exposed for three weeks to infective juveniles (J3) of H. indica LPP1. The entire experiment was conducted in duplicate, using a total of 64 snails. After this period, the snails were dissected to collect the hemolymph to evaluate the possible physiological alterations, namely total proteins, uric acid and hemolymph urea, as well as the activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) as a result of the infection. The terrariums were analyzed on alternate days throughout the experiment to count the dead snails. Intense proteolysis was observed in the infected snails. An increase in the level of uric acid and reduction of the hemolymph urea content indicated that the infection by H. indica results in the inversion of the excretion pattern of the host snail. Variations in the aminotransferase activities were also observed, with the infected group presenting significantly higher values (p<0.05) than the control group for both ALT and AST. The exposure to H. indica LPP1 caused 55% mortality, with the highest rate observed in the first week after exposure (30%). These results suggest that the use of H. indica LPP1 is a feasible alternative for the biological control of B. similaris., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Heterorhabditis bacteriophora is an entomopathogenic nematode (EPN) which infects its host by accessing the hemolymph where it releases endosymbiotic bacteria of the species Photorhabdus luminescens. We performed a genome-wide transcriptional analysis of the Drosophila response to EPN infection at the time point at which the nematodes reached the hemolymph either via the cuticle or the gut and the bacteria had started to multiply. Many of the most strongly induced genes have been implicated in immune responses in other infection models. Mapping of the complete set of differentially regulated genes showed the hallmarks of a wound response, but also identified a large fraction of EPN-specific transcripts. Several genes identified by transcriptome profiling or their homologues play protective roles during nematode infections. Genes that positively contribute to controlling nematobacterial infections encode: a homolog of thioester-containing complement protein 3, a basement membrane component (glutactin), a recognition protein (GNBP-like 3) and possibly several small peptides. Of note is that several of these genes have not previously been implicated in immune responses., (Copyright © 2013 S. Karger AG, Basel)

The dynamics of predation on parasites within prey has received relatively little attention despite the profound effects this is likely to have on both prey and parasite numbers and hence on biological control programmes where parasites are employed. The nematode Phasmarhabditis hermaphrodita is a commercially available biological agent against slugs. Predation on these slugs may, at the same time, result in intraguild predation on slug-parasitic nematodes. This study describes, for the first time, predation by carabid beetles on slugs and their nematode parasites on both spatial and temporal scales, using PCR-based methods. The highest nematode infection levels were found in the slugs Deroceras reticulatum and Arion silvaticus. Numbers of infected slugs decreased over time and no infected slugs were found four months after nematode application. The density of the most abundant slug, the invasive Arion vulgaris, was positively related to the activity-density of the carabid beetle, Carabus nemoralis. Predation on slugs was density and size related, with highest predation levels also on A. vulgaris. Predation on A. vulgaris decreased significantly in summer when these slugs were larger than one gram. Predation by C. nemoralis on slugs was opportunistic, without any preferences for specific species. Intraguild predation on the nematodes was low, suggesting that carabid beetles such as C. nemoralis probably do not have a significant impact on the success of biological control using P. hermaphrodita.

Entomopathogenic nematodes (EPNs) respond to a variety of stimuli when foraging. In a laboratory investigation, we tested the chemotactic responses of 8 EPN strains (Steinernema and Heterorhabditis) to three mechanically damaged maize root compounds (linalool, α-caryophyllene and β-caryophyllene). We hypothesized that the EPN directional response to the tested volatile compounds would vary among the species and volatile compound and may be related to foraging strategies. The nematodes with an intermediate foraging strategy (Steinernema feltiae) proved to be less active in their movement toward volatile compounds in a comparison with the ambushers (Steinernema carpocapsae) and cruisers (Steinernema kraussei and Heterorhabditis bacteriophora); β-caryophyllene was found to be the most attractive substance in our experiment. The results of our investigation showed that the cruisers were more attracted to β-caryophyllene than the ambushers and intermediates. The foraging strategy did not affect the movement of the IJs toward the other tested volatile compounds or the control. Our results suggest that the response to different volatile cues is more a strain-specific characteristic than a different host-searching strategy. Only S. carpocapsae strain B49 displayed an attraction to linalool, whereas S. kraussei showed a retarded reaction to β-caryophyllene and α-caryophyllene in our experiment. The EPN strains showed only a weak attraction to α-caryophyllene, suggesting that this volatile compound could not have an important role in the orientation of IJs to the damaged roots of maize plants. These results expand our knowledge of volatile compounds as the cues that may be used by EPNs for finding hosts or other aspects of navigation in the soil., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Species of Haematoloechus and Rhabdias both are ubiquitous lung parasites of frogs, yet surprisingly little research has been conducted on the interactions between these worms. In a small Nebraska stream, 256 Rhabdias joaquinensis and 225 Haematoloechus complexus were found in the lungs of Lithobates (=Rana) blairi. Thirty-six of the 44 (82%) frogs were co-infected with H. complexus and R. joaquinensis whereas 5 (11%) frogs were infected with only 1 species and 3 (7%) frogs were uninfected. Tests for association between H. complexus and R. joaquinensis found a significant positive relationship between the 2 parasites. Significant positive relationships in intensity of infection were found when both lungs were combined but not when lungs were considered individually. Tests for association between H. complexus and R. joaquinensis found no relationship between the worms in the anterior and posterior portions of the lungs. These data suggest that H. complexus and R. joaquinensis do not competitively exclude each other from the lungs of L. blairi in southeastern Nebraska.

Lungworms of the cosmopolitan genus Rhabdias are among the most common parasites of amphibians and squamate reptiles. The present study used experimental infections, field studies, and a molecular phylogeny to determine the host specificity of 6 Rhabdias spp. that infect snakes and anurans from North America. The molecular phylogeny suggests Rhabdias ranae from Nebraska and Mississippi may represent separate, cryptic species. In addition, the phylogeny strongly supports separate clades for anuran and snake lungworms. Field studies and experimental infections indicate that snake lungworms are generalist snake parasites; however, laboratory experiments also suggest that lizards can be infected under some environmental conditions. Lungworms from anurans were found not to infect salamanders or reptiles, in nature or in the laboratory; anuran lungworm species ranged from strict host specificity, e.g., R. ranae from Nebraska, to relative generalist, e.g., Rhabdias joaquinensis from Nebraska. Overall, host specificity for species of Rhabdias does not provide support for the evolution of progressive specialization over time. For most species of lungworms, host specificity in nature appears to be limited by both ecological and physiological factors, which vary between species and their hosts. Furthermore, some lungworms, e.g., Rhabdias bakeri from Missouri, appear to be tracking host resources instead of host phylogenies, an example of ecological fitting.

Organisms that live in close association with other organisms make up a large part of the world's diversity. One driver of this diversity is the evolution of host-species specificity, which can occur via reproductive isolation following a host-switch or, given the correct circumstances, via cospeciation. In this study, we explored the diversity and evolutionary history of Acrostichus nematodes that are associated with halictid bees in North America. First, we conducted surveys of bees in Virginia, and found six halictid species that host Acrostichus. To test the hypothesis of cospeciation, we constructed phylogenetic hypotheses of Acrostichus based on three genes. We found Acrostichus puri and Acrostichus halicti to be species complexes comprising cryptic, host-specific species. Although several nodes in the host and symbiont phylogenies were congruent and tests for cospeciation were significant, the host's biogeography, the apparent patchiness of the association across the host's phylogeny, and the amount of evolution in the nematode sequence suggested a mixture of cospeciation, host switching, and extinction events instead of strict cospeciation. Cospeciation can explain the relationships between Ac. puri and its augochlorine hosts, but colonization of Halictus hosts is more likely than cospeciation. The nematodes are vertically transmitted, but sexual transmission is also likely. Both of these transmission modes may explain host-species specificity and congruent bee and nematode phylogenies. Additionally, all halictid hosts come from eusocial or socially polymorphic lineages, suggesting that sociality may be a factor in the suitability of hosts for Acrostichus., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Despite impressive advances in the broad field of insect innate immunity, our understanding of the molecular basis of insect immune responses to nematode infections remains incomplete. Here we have investigated the transcriptional induction of immune pathway genes in the fruit fly Drosophila melanogaster upon infection with the entomopathogenic (or insect pathogenic) nematodes Heterorhabditis bacteriophora and their mutualistic bacteria Photorhabdus luminescens, either collectively or separately. We show that in most cases, infection of wild-type adult flies with Heterorhabditis nematodes carrying or lacking mutualistic Photorhabdus bacteria results in the up-regulation of genes in the Toll, Imd, JAK/STAT, JNK and TGF-beta pathways. We also find that direct injection of Photorhabdus bacteria into flies fails to induce the transcription of antimicrobial peptide genes and stress-related genes in Drosophila. These results suggest that Heterorhabditis nematodes and their associated Photorhabdus bacteria employ distinct strategies to evade the Drosophila immune response and establish infection., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

The entomopathogenic nematode (EPN) Heterorhabditis bacteriophora is used in biological plant protection to control pest insects. In the past, several attempts targeted at an enhancement of the desiccation tolerance of EPN by genetic selection in order to improve their storage stability. The subsequent loss of improved beneficial traits after release of selection pressure has often been reported. In order to stabilize progress of selective breeding, selection during liquid culturing was tested against propagation in host insects. After release of the selection pressure, the tolerance was monitored over additional reproductive cycles in vivo and in vitro to compare the stability of the trait. Furthermore, it was tested whether the virulence of the selected strains would be impaired. Exposure to desiccation stress prior to propagation, in vivo or in vitro, both resulted in increasing desiccation tolerance. When selection pressure was released, the gained tolerance was lost again during in vivo production, whereas the tolerance was maintained at a high level when EPNs were cultured in liquid culture. In Heterorhabditis sp., liquid culture conditions produce highly homozygous, genetically stable inbred lines. The investigation provides easily applicable methods to improve and stabilize beneficial traits of heterorhabditid EPNs through selective breeding in liquid culture. Compared to nematodes from in vivo propagation, production in liquid media yielded EPN of higher virulence.

Planococcus citri, the citrus mealybug, is the most important species of mealybug known to infest citrus in South Africa. Various laboratory bioassays were conducted to determine the potential of entomopathogenic nematodes to control P. citri. Adult female P. citri were screened for susceptibility to six indigenous nematode species. P. citri was found to be most susceptible to Steinernema yirgalemense and Heterorhabditis zealandica, causing 97% and 91% mortality, respectively. The development of nematodes after infecting adult female P. citri showed both H. zealandica and S. yirgalemense were able to complete their life cycles inside the host. Further bioassays illustrated a linear relationship between mealybug mortality and the concentration of nematodes applied, with the highest level of control using a concentration of 80 infective juveniles (IJs)/insect. As nematodes would be used as an above-ground application to control P. citri in citrus orchards, available water is a major limiting factor. Insecticidal activity proved to be dependent on the available surface moisture after nematode application. The water activity (a(w)) bioassay indicated that S. yirgalemense to be two times more tolerant to lower levels of free water, with a(w50)=0.96 and a(w90)=0.99, compared to H. zealandica with a(w50)=0.98 and a(w)90=1.0. After application, nematodes have a limited time frame in which to locate and infect hosts, as the level of available free water gradually decreases, as trees dry out. S. yirgalemense proved able to locate and infect P. citri quicker than H. zealandica. Nematode activity was not significantly affected when exposed to 15°C, 20°C and 25°C. IJs were able to infect P. citri at an exposure time as short as half an hour. Results also showed that the first 2-4h post application is the most decisive time for establishing successful infection of mealybugs. This is the first report on the potential use of nematodes for the control of P. citri., (Copyright © 2012 Elsevier Inc. All rights reserved.)