Freezing and Desiccation Tolerance in Entomopathogenic Nematodes: Diversity and Correlation of Traits

The ability of entomopathogenic nematodes to tolerate environmental stress such as desiccating or freezing conditions, can contribute significantly to biocontrol efficacy. Thus, in selecting which nematode to use in a particular biocontrol program, it is important to be able to predict which strain or species to use in target areas where environmental stress is expected. Our objectives were to (i) compare inter- and intraspecific variation in freeze and desiccation tolerance among a broad array of entomopathogenic nematodes, and (ii) determine if freeze and desiccation tolerance are correlated. In laboratory studies we compared nematodes at two levels of relative humidity (RH) (97% and 85%) and exposure periods (24 and 48 h), and nematodes were exposed to freezing temperatures (-2°C) for 6 or 24 h. To assess interspecific variation, we compared ten species including seven that are of current or recent commercial interest: Heterorhabditis bacteriophora (VS), H. floridensis, H. georgiana, (Kesha), H. indica (HOM1), H. megidis (UK211), Steinernema carpocapsae (All), S. feltiae (SN), S. glaseri (VS), S. rarum (17C&E), and S. riobrave (355). To assess intraspecific variation we compared five strains of H. bacteriophora (Baine, Fl1-1, Hb, Oswego, and VS) and four strains of S. carpocapsae (All, Cxrd, DD136, and Sal), and S. riobrave (355, 38b, 7-12, and TP). S. carpocapsae exhibited the highest level of desiccation tolerance among species followed by S. feltiae and S. rarum; the heterorhabditid species exhibited the least desiccation tolerance and S. riobrave and S. glaseri were intermediate. No intraspecific variation was observed in desiccation tolerance; S. carpocapsae strains showed higher tolerance than all H. bacteriophora or S. riobrave strains yet there was no difference detected within species. In interspecies comparisons, poor freeze tolerance was observed in H. indica, and S. glaseri, S. rarum, and S. riobrave whereas H. georgiana and S. feltiae exhibited the highest freeze tolerance, particularly in the 24-h exposure period. Unlike desiccation tolerance, substantial intraspecies variation in freeze tolerance was observed among H. bacteriophora and S. riobrave strains, yet within species variation was not detected among S. carpocapsae strains. Correlation analysis did not detect a relationship between freezing and desiccation tolerance.     

Dauer juvenile longevity and stress tolerance in natural populations of entomopathogenic nematodes: is there a relationship?

Qualitative and quantitative genetic analysis of life span in experimental adult animals predicts that resistance to stress and longevity are positively correlated, but such studies on field populations of animals are rare. We tested this hypothesis using dauer juveniles of 15 natural populations of the entomopathogenic nematode, Heterorhabditis bacteriophora, collected from diverse localities. Dauer juvenile longevity at 25 degrees C in autoclaved tap water and tolerance to major environmental stresses including heat (survival at 40 degrees C for 2 h), ultraviolet (UV) radiation (original virulence remaining after exposure to 302 nm UV for 5 min), hypoxia (survival at approximately 0% dissolved O2 at 25 degrees C for 96 h), and desiccation (survival in 25% glycerol at 25 degrees C for 72 h) differed significantly among populations. Intrinsic dauer juvenile longevity, defined as the number of weeks to 90% mortality (LT90) estimated using probit analysis of nematode survival data at 25 degrees C varied between 6 and 16 weeks among populations. Longevity was most strongly correlated with heat followed by UV and hypoxia tolerance, respectively, but showed no correlation with desiccation tolerance. The strong positive correlation of longevity with heat tolerance was further confirmed through principal components analysis which showed almost identical variance for heat and longevity. Among the stress factors, only UV tolerance was positively correlated with heat and hypoxia tolerance. Differences in longevity and stress tolerance in nematode populations isolated from a single 200 m2 grassland locality further support another hypothesis that population structure of heterorhabditid nematodes is highly fragmented, thus suggesting the existence of metapopulation dynamics.     

Coiling is Not Essential to Anhydrobiosis by Aphelenchus avenae on Agar Amended with Sucrose

The roles of preconditioning and coiling upon entrance into anhydrobiosis by Aphelenchus avenae were tested via video-assisted analysis at 25²°C. Fourth-stage juveniles or young adults of A. avenae were individually placed on 5% agar containing 0.8 M sucrose. Nematodes became quiescent within 3 hr, then gradually resumed a low level of activity and assumed a coiled posture. High desiccation survival rate was recorded when nematodes were incubated on agar for more than 6 hr; the survival rates were 0%, 3%, 73%, and 92% for 0, 2, 6, and 12 hr on agar, respectively. All nematodes placed on agar for 24 hr or more revived after rehydration following desiccation. Once nematodes were on agar for a sufficient time, no difference in desiccation survival was observed between nematodes taking a coiled posture and those uncoiled artificially. Based on these results, exposure to osmotic stress for 6 hr can prepare A. aveae physiologically for anhydrobiosis, but coiling does not appear to be a physiological requirement for desiccation in survival.     

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.     

Viability and Virulence of Entomopathogenic Nematodes Exposed to Ultraviolet Radiation

Entomopathogenic nematodes (EPNs) can be highly effective biocontrol agents, but their efficacy can be reduced due to exposure to environmental stress such as from ultraviolet (UV) radiation. Our objectives were to 1) compare UV tolerance among a broad array of EPN species, and 2) investigate the relationship between reduced nematode viability (after exposure to UV) and virulence. Nematodes exposed to a UV radiation (254 nm) for 10 or 20 min were assessed separately for viability (survival) and virulence to Galleria mellonella. We compared 9 different EPN species and 15 strains: Heterorhabditis bacteriophora (Baine, fl11, Oswego, and Vs strains), H. floridensis (332), H. georgiana (Kesha), H. indica (HOM1), H. megidis (UK211), Steinernema carpocapsae (All, Cxrd, DD136, and Sal strains), S. feltiae (SN), S. rarum (17C&E), and S. riobrave (355). In viability assessments, steinernematids, particularly strains of S. carpocapsae, generally exhibited superior UV tolerance compared with the heterorhabditids. However, some heterorhabditids tended to be more tolerant than others, e.g., H. megidis and H. bacteriophora (Baine) were most susceptible and H. bacteriophora (Vs) was the only heterorhabditid that did not exhibit a significant effect after 10 min of exposure. All heterorhabditids experienced reduced viability after 20 min exposure though several S. carpocapsae strains did not. In total, after 10 or 20 min exposure, the viability of seven nematode strains did not differ from their non-UV exposed controls. In virulence assays, steinernematids (particularly S. carpocapsae strains) also tended to exhibit higher UV tolerance. However, in contrast to the viability measurements, all nematodes experienced a reduction in virulence relative to their controls. Correlation analysis revealed that viability among nematode strains is not necessarily related to virulence. In conclusion, our results indicate that the impact of UV varies substantially among EPNs, and viability alone is not a sufficient measure for potential impact on biocontrol efficacy as other characters such as virulence may be severely affected even when viability remains high.     

Dynamics of a Subterranean Trophic Cascade in Space and Time

Trophic cascades, whereby predators indirectly benefit plant biomass by reducing herbivore pressure, form the mechanistic basis for classical biological control of pest insects. Entomopathogenic nematodes (EPN) are lethal to a variety of insect hosts with soil-dwelling stages, making them promising biocontrol agents. EPN biological control programs, however, typically fail because nematodes do not establish, persist and/or recycle over multiple host generations in the field. A variety of factors such as local abiotic conditions, host quantity and quality, and rates of movement affect the probability of persistence. Here, we review results from 13 years of study on the biology and ecology of an endemic population of Heterorhabditis marelatus (Rhabditida: Heterorhabditidae) in a California coastal prairie. In a highly seasonal abiotic environment with intrinsic variation in soils, vegetation structure, and host availability, natural populations of H. marelatus persisted at high incidence at some but not all sites within our study area. Through a set of field and lab experiments, we describe mechanisms and hypotheses to understand the persistence of H. marelatus. We suggest that further ecological study of naturally occurring EPN populations can yield significant insight to improve the practice and management of biological control of soil-dwelling insect pests.     

Entomopathogenic Nematode Production and Application Technology

Production and application technology is critical for the success of entomopathogenic nematodes (EPNs) in biological control. Production approaches include in vivo, and in vitro methods (solid or liquid fermentation). For laboratory use and small scale field experiments, in vivo production of EPNs appears to be the appropriate method. In vivo production is also appropriate for niche markets and small growers where a lack of capital, scientific expertise or infrastructure cannot justify large investments into in vitro culture technology. In vitro technology is used when large scale production is needed at reasonable quality and cost. Infective juveniles of entomopathogenic nematodes are usually applied using various spray equipment and standard irrigation systems. Enhanced efficacy in EPN applications can be facilitated through improved delivery mechanisms (e.g., cadaver application) or optimization of spray equipment. Substantial progress has been made in recent years in developing EPN formulations, particularly for above ground applications, e.g., mixing EPNs with surfactants or polymers or with sprayable gels. Bait formulations and insect host cadavers can enhance EPN persistence and reduce the quantity of nematodes required per unit area. This review provides a summary and analysis of factors that affect production and application of EPNs and offers insights for their future in biological insect suppression.     

In vivo and In vitro Rearing of Entomopathogenic Nematodes (Steinernematidae and Heterorhabditidae)

Entomopathogenic nematodes (EPN) (Steinernematidae and Heterorhabditidae) have a mutualistic partnership with Gram-negative Gamma-Proteobacteria in the family Enterobacteriaceae. Xenorhabdus bacteria are associated with steinernematids nematodes while Photorhabdus are symbionts of heterorhabditids. Together nematodes and bacteria form a potent insecticidal complex that kills a wide range of insect species in an intimate and specific partnership. Herein, we demonstrate in vivo and in vitro techniques commonly used in the rearing of these nematodes under laboratory conditions. Furthermore, these techniques represent key steps for the successful establishment of EPN cultures and also form the basis for other bioassays that utilize these organisms for research. The production of aposymbiotic (symbiont–free) nematodes is often critical for an in-depth and multifaceted approach to the study of symbiosis. This protocol does not require the addition of antibiotics and can be accomplished in a short amount of time with standard laboratory equipment. Nematodes produced in this manner are relatively robust, although their survivorship in storage may vary depending on the species used. The techniques detailed in this presentation correspond to those described by various authors and refined by P. Stock’s Laboratory, University of Arizona (Tucson, AZ, USA). These techniques are distinct from the body of techniques that are used in the mass production of these organisms for pest management purposes.     

Anhydrobiosis in Five Species of Plant Associated Nematodes

Five species of nematodes - Hemicriconemoides pseudobrachyurum, Hemicycliophora conida, Macroposthonia ornata, Aphelenchoides ritzemabosi, and Psilenchus hilarulus -were desiccated to study their capacity to survive anhydrobiotically. Results indicate that the ability of the sheath to shrink quickly and its relatively loose attachment with the nematode body allow H. conida to survive longer than H. pseudobrachyurum; the survival of M. ornata was intermediate, A. ritzemabosi and P. hilarulus survived immersion in paraffin oil for 12 and 17 days, respectively. Both of these nematodes possess multiple contraction ability; i.e., coiling coupled with transverse and longitudinal folding of the cuticle. P. hilarulus is a new addition to the list of anhydrobiotic nematodes.     

From Augmentation to Conservation of Entomopathogenic Nematodes: Trophic Cascades, Habitat Manipulation and Enhanced Biological Control of Diaprepes abbreviatus Root Weevils in Florida Citrus Groves

The use of entomopathogenic nematodes (EPN) for management of the root weevil, Diaprepes abbreviatus, in Florida citrus groves is considered a biological control success story and typically involves augmentation in which EPN are applied inundatively as biopesticides to quickly kill the pest. However, recent evidence indicates that efficacy of EPN applications in Florida citrus depends on soil type. They are very effective in the well drained coarse sands of the Central Ridge but often less so in poorly drained fine-textured soils of the Flatwoods. Moreover, groves on the Central Ridge can harbor rich communities of endemic EPN that might often suppress weevil populations below economic thresholds, whereas Flatwoods groves tend to have few endemic EPN and frequent weevil problems. Current research is examining the ecological dynamics of EPN in Florida citrus groves, the potential impact of EPN augmentation on soil food webs, especially endemic EPN, and whether habitat manipulation and inoculation strategies might be effective for conserving and enhancing EPN communities to achieve long-term control in problem areas. Conservation biological control could extend the usefulness of EPN in Florida citrus and be especially appropriate for groves with persistent weevil problems.     

Desiccation tolerance,longevity and seed-siring ability of entomophilous pollen from UK native orchid species

Background and Aims

Pollinator-limited seed-set in some terrestrial orchids is compensated for by the presence of long-lived flowers. This study tests the hypothesis that pollen from these insect-pollinated orchids should be desiccation tolerant and relatively long lived using four closely related UK terrestrial species; Anacamptis morio, Dactylorhiza fuchsii, D. maculata and Orchis mascula.

Methods

Pollen from the four species was harvested from inflorescences and germinated in vitro, both immediately and also after drying to simulate interflower transit. Their tolerance to desiccation and short-term survival was additionally assessed after 3 d equilibration at a range of relative humidities (RHs), and related to constructed sorption isotherms (RH vs. moisture content, MC). Ageing of D. fuchsii pollen was further tested over 2 months against temperature and RH, and the resultant survival curves were subjected to probit analysis, and the distribution of pollen death in time (σ) was determined. The viability and siring ability, following artificial pollinations, were determined in D. fuchsii pollen following storage for 6 years at –20 °C.

Key Results

The pollen from all four species exhibited systematic increases in germinability and desiccation tolerance as anthesis approached, and pollen from open flowers generally retained high germinability. Short-term storage revealed sensitivity to low RH, whilst optimum survival occurred at comparable RHs in all species. Similarly, estimated pollen life spans (σ) at differing temperatures were longest under the dry conditions. Despite a reduction in germination and seeds per capsule, long-term storage of D. fuchsii pollen did not impact on subsequent seed germination in vitro.

Conclusions

Substantial pollen desiccation tolerance and life span of the four entomophilous orchids reflects a resilient survival strategy in response to unpredictable pollinator visitation, and presents an alternative approach to germplasm conservation.     

The lethal and sub-lethal consequences of entomopathogenic nematode infestation and exposure for adult pine weevils, Hylobius abietis (Coleoptera: Curculionidae)

Entomopathogenic nematodes (EPN) frequently kill their host within 1-2 days, and interest in EPN focuses mainly on their lethality. However, insects may take longer to die, or may fail to die despite being infected, but little is known about the effects of EPN infection on insects, other than death. Here we investigate both lethal and sub-lethal effects of infection by two EPN species, Steinernema carpocapsae and Heterorhabditis downesi, on adults of the large pine weevil, Hylobius abietis. Following 12 h nematode-weevil contact in peat, S. carpocapsae killed a significantly higher proportion of weevils (87-93%) than H. downesi (43-57%) at all concentrations tested. Less than 10% of weevils were dead within 2 days, and weevils continued to die for up to 10 days after exposure (LT₅₀ of 3 days or more). In a separate experiment, live weevils dissected 6 days after a 24 h exposure to nematodes on filter paper harbored encapsulated and dead nematodes, showing that weevils could defend themselves against infection. Some live weevils also harbored live nematodes 6 days after they had been removed from the nematode infested medium. Feeding by weevils was not affected by infection with, or exposure to, either species of EPN. We discuss these results in relation to the use of EPN in biological control against H. abietis.     

Pathogenic effect of entomopathogenic nematode-bacterium complexes on terrestrial isopods

In this study, we evaluated the effect of entomopathogenic nematodes (EPNs) Steinernema carpocapsae, Steinernema feltiae and Heterorhabditis bacteriophora, symbiotically associated with bacteria of the genera Xenorhabdus or Photorhabdus, on the survival of eight terrestrial isopod species. The EPN species S. carpocapsae and H. bacteriophora reduced the survival of six isopod species while S. feltiae reduced survival for two species. Two terrestrial isopod species tested (Armadillidium vulgare and Armadillo officinalis) were found not to be affected by treatment with EPNs while the six other isopod species showed survival reduction with at least one EPN species. By using aposymbiotic S. carpocapsae (i.e. without Xenorhabdus symbionts), we showed that nematodes can be isopod pathogens on their own. Nevertheless, symbiotic nematodes were more pathogenic for isopods than aposymbiotic ones showing that bacteria acted synergistically with their nematodes to kill isopods. By direct injection of entomopathogenic bacteria into isopod hemolymph, we showed that bacteria had a pathogenic effect on terrestrial isopods even if they appeared unable to multiply within isopod hemolymphs. A developmental study of EPNs in isopods showed that two of them (S. carpocapsae and H. bacteriophora) were able to develop while S. feltiae could not. No EPN species were able to produce offspring emerging from isopods. We conclude that EPN and their bacteria can be pathogens for terrestrial isopods but that such hosts represent a reproductive dead-end for them. Thus, terrestrial isopods appear not to be alternative hosts for EPN populations maintained in the absence of insects.     

Competition Between Entomopathogenic and Free-Living Bactivorous Nematodes in Larvae of the Weevil Diaprepes abbreviatus

Field and laboratory experiments were conducted to determine the degree to which free-living, bactivorous nematodes (FLBN) are able to competitively displace entomopathogenic nematodes (EPN) from insect cadavers. Two hundred larvae of the insect Diaprepes abbreviatus were buried at regular intervals during 2 years in experimental plots that were untreated or treated twice annually with Steinernema riobrave. Larvae were recovered after 7 days, and nematodes emerging from cadavers during the next 30 days were identified. The monthly prevalence of FLBN was directly related to that of S. riobrave (r = 0.38; P = 0.001) but was not related to the prevalence of the endemic EPN, S. diaprepesi, Heterorhabditis zealandica, H. indica, or H. bacteriophora (r = 0.02; P = 0.80). In a second experiment, treatment of small field plots with S. riobrave increased the prevalence of insect cadavers in which only FLBN were detected compared to untreated controls (30% vs. 14%; P = 0.052), and increased numbers of FLBN per buried insect by more than 10-fold. In the laboratory, sand microcosms containing one D. abbreviatus larva were treated with (i) the FLBN, Pellioditis sp.; (ii) S. riobrave; (iii) S. riobrave + Pellioditis; or (iv) neither nematode. Insect mortality was higher in the presence of both nematodes (57%) than when S. riobrave was alone (42%) (P = 0.01). An average of 59.2 Pellioditis sp. g^-1 insect body weight emerged in the presence of S. riobrave, whereas 6.2 nematodes g^-1 insect were recovered in the absence of the EPN (P = 0.01). Pellioditis sp. reduced the number of S. riobrave per cadaver by 84%; (P = 0.03), and per available insect by 82% (P = 0.001), compared to S. riobrave alone. Population size of S. diaprepesi was not affected by Pellioditis sp. in experiments of the same design. Faster development (P = 0.05) and nutrient appropriation within the insect cadaver by S. diaprepesi compared to S. riobrave may increase the fitness of the former species to compete with Pellioditis sp. The results of these studies demonstrate the potential of FLBN to regulate population densities of EPN and to dampen estimates of EPN-induced mortality of insect pests in the field.     

Effect of insect cadaver desiccation and soil water potential during rehydration on entomopathogenic nematode (Rhabditida: Steinernematidae and Heterorhabditidae) production and virulence

We examined the influence of insect cadaver desiccation on the virulence and production of entomopathogenic nematodes (EPNs), common natural enemies of many soil-dwelling insects. EPNs are often used in biological control, and we investigated the feasibility of applying EPNs within desiccated insect cadavers. Desiccation studies were conducted using the factitious host, Galleria mellonella (Lepidoptera: Pyralidae, wax moth larvae) and three EPN species (Heterorhabditis bacteriophora ‘HB1’, Steinernema carpocapsae ‘All’, and Steinernema riobrave). Weights of individual insect cadavers were tracked daily during the desiccation process, and cohorts were placed into emergence traps when average mass losses reached 50%, 60%, and 70% levels. We tracked the proportion of insect cadavers producing infective juveniles (IJs), the number and virulence of IJs produced from desiccated insect cadavers, and the influence of soil water potentials on IJ production of desiccated insect cadavers. We observed apparent differences in the desiccation rate of the insect cadavers among the three species, as well as apparent differences among the three species in both the proportion of insect cadavers producing IJs and IJ production per insect cadaver. Exposure of desiccated insect cadavers to water potentials greater than −2.75 kPa stimulated IJ emergence. Among the nematode species examined, H. bacteriophora exhibited lower proportions of desiccated insect cadavers producing IJs than the other two species. Desiccation significantly reduced the number of IJs produced from insect cadavers. At the 60% mass loss level, however, desiccated insect cadavers from each of the three species successfully produced IJs when exposed to moist sand, suggesting that insect cadaver desiccation may be a useful approach for biological control of soil insect pests.     

Entomopathogenic Nematodes Are Not an Alternative to Fenamiphos for Management of Plant-Parasitic Nematodes on Golf Courses in Florida

With the cancellation of fenamiphos in the near future, alternative nematode management tactics for plant-parasitic nematodes (PPN) on golf courses need to be identified. The use of entomopathogenic nematodes (EPN) has been suggested as one possible alternative. This paper presents the results of 10 experiments evaluating the efficacy of EPN at managing PPN on turfgrasses and improving turf performance. These experiments were conducted at various locations throughout Florida over the course of a decade. In different experiments, different EPN species were tested against different species of PPN. Separate experiments evaluated multiple rates and applications of EPN, compared different EPN species, and compared single EPN species against multiple species of PPN. In a few trials, EPN were associated with reductions in certain plant-parasite species, but in other trials were associated with increases. In most trials, EPN had no effect on plant parasites. Because EPN were so inconsistent in their results, we conclude that EPN are not acceptable alternatives to fenamiphos by most turf managers in Florida at this time.     

Genome sequence and comparative analysis of a putative entomopathogenic Serratia isolated from Caenorhabditis briggsae

Background

Entomopathogenic associations between nematodes in the genera Steinernema and Heterorhabdus with their cognate bacteria from the bacterial genera Xenorhabdus and Photorhabdus, respectively, are extensively studied for their potential as biological control agents against invasive insect species. These two highly coevolved associations were results of convergent evolution. Given the natural abundance of bacteria, nematodes and insects, it is surprising that only these two associations with no intermediate forms are widely studied in the entomopathogenic context. Discovering analogous systems involving novel bacterial and nematode species would shed light on the evolutionary processes involved in the transition from free living organisms to obligatory partners in entomopathogenicity.

Results

We report the complete genome sequence of a new member of the enterobacterial genus Serratia that forms a putative entomopathogenic complex with Caenorhabditis briggsae. Analysis of the 5.04 MB chromosomal genome predicts 4599 protein coding genes, seven sets of ribosomal RNA genes, 84 tRNA genes and a 64.8 KB plasmid encoding 74 genes. Comparative genomic analysis with three of the previously sequenced Serratia species, S. marcescens DB11 and S. proteamaculans 568, and Serratia sp. AS12, revealed that these four representatives of the genus share a core set of ~3100 genes and extensive structural conservation. The newly identified species shares a more recent common ancestor with S. marcescens with 99 % sequence identity in rDNA sequence and orthology across 85.6 % of predicted genes. Of the 39 genes/operons implicated in the virulence, symbiosis, recolonization, immune evasion and bioconversion, 21 (53.8 %) were present in Serratia while 33 (84.6 %) and 35 (89 %) were present in Xenorhabdus and Photorhabdus EPN bacteria respectively.

Conclusion

The majority of unique sequences in Serratia sp. SCBI (South African Caenorhabditis briggsae Isolate) are found in ~29 genomic islands of 5 to 65 genes and are enriched in putative functions that are biologically relevant to an entomopathogenic lifestyle, including non-ribosomal peptide synthetases, bacteriocins, fimbrial biogenesis, ushering proteins, toxins, secondary metabolite secretion and multiple drug resistance/efflux systems. By revealing the early stages of adaptation to this lifestyle, the Serratia sp. SCBI genome underscores the fact that in EPN formation the composite end result – killing, bioconversion, cadaver protection and recolonization- can be achieved by dissimilar mechanisms. This genome sequence will enable further study of the evolution of entomopathogenic nematode-bacteria complexes.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1697-8) contains supplementary material, which is available to authorized users.     

Factors affecting stress tolerance in recalcitrant embryonic axes from seeds of four Quercus (Fagaceae) species native to the USA or China

MethodsWater relations and survival of excised axes in response to water loss and cryo-exposure were compared for four Quercus species from subtropical China (Q. franchetii, Q. schottkyana) and temperate USA (Q. gambelii, Q. rubra).ConclusionsQuercus species adapted to arid and semi-humid climates still produce recalcitrant seeds. The ability to avoid freezing rather than drought may be a more important selection factor to increase desiccation tolerance. Cryopreservation of recalcitrant germplasm from temperate species is currently feasible, whilst additional protective treatments are needed for ex situ conservation of Quercus from tropical and subtropical areas.     

Control of the Oriental Fruit Moth, Grapholita molesta, Using Entomopathogenic Nematodes in Laboratory and Fruit Bin Assays

The oriental fruit moth (OFM), Grapholita molesta (Busck), which is among the most important insect pests of peaches and nectarines, has developed resistance to a wide range of insecticides. We investigated the ability of the entomopathogenic nematodes (EPN) Steinernema carpocapsae (Weiser), S. feltiae (Filipjev), S. riobrave (Cabanillas et al.), and Heterorhabditis marelatus (Liu and Berry) to control OFM under laboratory and fruit bin conditions. At a dosage of 10 infective juveniles (IJ)/cm² in the laboratory, S. carpocapsae caused 63%, S. feltiae 87.8%, S. riobrave 75.6%, and H. marelatus 67.1% OFM mortality. All four nematode species caused significant OFM larval mortality in comparison to the nontreated controls. Steinernema feltiae was used for the bin assays due to the higher OFM mortality it caused than the other tested EPN species and to its ability to find OFM under cryptic environments. Diapausing cocooned OFM larvae in miniature fruit bins were susceptible to IJ of S. feltiae in infested corner supports and cardboard strips. Treatment of bins with suspensions of 10 or 25 S. feltiae IJ/ml water with wetting agent (Silwet L77) resulted in 33.3 to 59% and 77.7 to 81.6% OFM mortality in corner supports and cardboard strips, respectively. This paper presents new information on the use of EPN, specifically S. feltiae, as nonchemical means of OFM control.