Parasites of firebugs in Austria with focus on the "micro"-diversity of the cosmopolitan trypanosomatid Leptomonas pyrrhocoris.

In this work, we investigated parasites of the firebug Pyrrhocoris apterus in Austria and demonstrated that in addition to the extensively studied Leptomonas pyrrhocoris, it can also be infected by Blastocrithidia sp. and by a mermithid, which for the first time has been characterized using molecular methods. This diversity can be explained by the gregarious lifestyle, as well as the coprophagous and cannibalistic behavior of the insect hosts that makes them susceptible to various parasites. In addition, we showed no tight association of the L. pyrrhocoris haplotypes and geographical locations (at least, considering the relatively small scale of locations in Austria) implying that the natural populations of L. pyrrhocoris are mixed due to the mobility of their firebug hosts.

Biological control of the stink bug Nezara viridula (Heteroptera: Pentatomidae) by two parasitoids and their interaction in non-crop habitats: a simulation model.

Non-cultivated areas are resting, overwintering, feeding, and/or reproducing habitats for insects, and also places from where crop areas are colonized; thus, they are essential for understanding the biological control programs in agroecosystems. We developed a simulation model for a non-cultivated area of Buenos Aires province (Argentina), and we analyzed the control of Nezara viridula achieved by the action of two parasitoids: the oophagous Trissolcus basalis and the tachinid Trichopoda giacomellii, which attack older nymphs and adults. The model is a discrete time, deterministic, phenomenological, spatially homogeneous with a 1-week time interval simulation model, based on the age-structure and/or stage-structure of N. viridula and its two parasitoids. The host-parasitoid interactions were combined with a degree-day model affecting development times of T. giacomellii pupae and T. basalis pre-imaginal stages. The simultaneous attack of both parasitoid species enables the persistence of the system at low host densities, mediated by the functional response of the parasitoids, identified as population regulation factors. However, if only one parasitoid exists (i.e., only T. basalis or only T. giacomellii) the interaction N. viridula-parasitoid persisted but at higher density of N. viridula. These results explain the successful biological control of N. viridula after the introduction of T. basalis in the 1980s, when T. giacomellii was the only parasitoid present, unable to control N. viridula. Our model shows an indirect competition when both parasitoids are present: the attack of one of them diminished the potential number of hosts available to the other parasitoid species. In the field this interaction is obscured by the hibernation period which acted as a reset mechanism affecting the density and age/stage structure of all three populations. Our model was supported by field observations, and never exhibited the extinction of any of the parasitoids from the interaction.

Contrasting reproductive traits of competing parasitoids facilitate coexistence on a shared host pest in a biological control perspective.

BACKGROUND: Interspecific competition in insect parasitoids is an important ecological phenomenon that has relevant implications for biological pest control. To date, interspecific intrinsic (=larval) competition has been intensively studied, while investigations on extrinsic (=adult) competition have often lagged behind. In this study we examined the role played by parasitoid reproductive traits and host clutch size on the outcome of extrinsic competition between Trissolcus basalis (Wollaston) and Ooencyrtus telenomicida (Vassiliev), two egg parasitoids of the pest Nezara viridula (L). Laboratory experiments were conducted by allowing both parasitoid species to exploit an egg mass made of 10, 20, 30, or 40 hosts through single or simultaneous releases. Furthermore, under field conditions, egg masses consisting of 10 or 40 hosts were exposed in a tomato crop in order to validate laboratory investigation. RESULTS: The results show that the egg mass size is an important predictor of extrinsic competition in our study system as a higher proportion of T. basalis emerged from large egg masses, while O. telenomicida dominated in small egg masses. Analysis of reproductive traits of parasitoid species indicates that T. basalis has superior abilities in host exploitation compared with O. telenomicida. CONCLUSIONS: We found that contrasting reproductive traits of two competing egg parasitoid species facilitate coexistence on a shared stink bug host. This work also highlights the importance to consider extrinsic competitive interactions between parasitoid species in a biological control perspective. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Comparison of Yellow and Blue Sticky Cards for Detection and Monitoring Parasitoid Wasps of the Invasive Halyomorpha halys (Hemiptera: Pentatomidae).

The invasive Halyomorpha halys (Stål) is a significant agricultural and urban nuisance pest in many parts of the world. In North America, biological control of H. halys by parasitoid wasps in the families Scelionidae and Eupelmidae has shown promise. An effective technique for detection and monitoring native and exotic parasitoids is the deployment of yellow sticky cards; however, yellow cards also attract nontarget arthropods, reducing efficiency and accuracy of parasitoid screening. This study sought to identify an alternative yet effective trapping technique by comparing the number of target parasitoid wasps [Anastatus spp. Motschulsky (Hymenoptera: Eupelmidae), Telenomus spp. Haliday (Hymenoptera: Scelionidae), and Trissolcus spp. Ashmead (Hymenoptera: Scelionidae)] and arthropod bycatch on yellow and blue sticky cards deployed in urban, orchard, and vegetable landscapes in northern Utah from late May to early October in 2019 and 2020. Yellow sticky cards captured 54-72% more target parasitoids than blue cards from June through August in all three landscape types in both years; however, a positive correlation in parasitoid capture indicated blue cards detect target parasitoids, just in fewer numbers. Both card colors detected adventive Trissolcus japonicus (Ashmead) (Hymenoptera: Scelionidae) in initial findings of 2019, and in expanded locations of 2020. Furthermore, blue cards captured 31-48% less Diptera and nontarget Hymenoptera than yellow cards in both years across all three landscapes, translating to reduced card processing time and impacts to beneficial insect populations. Our results suggest that blue vs yellow sticky cards offer an alternative monitoring technique to survey for H. halys parasitoids.

Re-collection and identity of Ooencyrtus californicus (Hymenoptera: Encyrtidae), and its new synonym, Ooencyrtus lucidus.

Girault (1917) very briefly described and diagnosed the encyrtid wasp species Ooencyrtus californicus Girault (Hymenoptera: Encyrtidae) with the following limited data (p. 22): "Sacramento, California, from bug eggs on Pinus sabiniana, September". The first author examined its two syntypes, poorly mounted on a slide, of which only parts of the four antennae and a slightly damaged fore wing remain (Triapitsyn et al. 2020). They concluded that this species was better considered a nomen dubium until fresh specimens could be collected from the same host plant in or near the type locality. In July 2019, an effort was made to re-collect O. californicus from foothill (or gray) pine, Pinus sabiniana (Pinaceae), in the Oakhurst area of Madera County, California, USA, where this pine is abundant, but no specimens were captured. At the same time, a very similar insect was collected from sentinel eggs of the invasive stink bug Bagrada hilaris (Burmeister) in Riverside, California. This insect was cultured in the laboratory as part of a B. hilaris biological control program. Due to the poor preservation of the type specimens and in the absence of genetic evidence, it was impossible to positively and properly attribute specimens of this newly collected parasitoid to O. californicus. Therefore, to provide a much needed scientific name for this native egg parasitoid, it was described as Ooencyrtus lucidus Triapitsyn Ganjisaffar in Triapitsyn et al. (2020). The two nominal species were separated based on minor differences in the proportions of the scape, pedicel, and first funicular segment of the female antenna (Triapitsyn et al. 2020). John S. Noyes, world expert on Encyrtidae and one of the reviewers of Triapitsyn et al. (2020), disagreed with the erection of O. lucidus as a new taxon based on the morphological similarities of the female antenna with that of O. californicus, so the description was published with the understanding that the two might be conspecific. But to settle this one way or another new specimens of O. californicus had to be collected to allow morphological and molecular analyses.

Characterization of a new cosmopolitan genus of trypanosomatid parasites, Obscuromonas gen. nov. (Blastocrithidiinae subfam. nov.).

The expanding phylogenetic tree of trypanosomatid flagellates (Kinetoplastea: Trypanosomatidae) contains a long-known and phylogenetically well-supported species-rich lineage that was provisionally named as the 'jaculum' clade. Its members were found in representatives of several unrelated families of heteropteran bugs captured in South and Central America, Europe, Africa, and Asia. However, this group resisted introduction into the culture, a needed prerequisite for its proper characterization. Here we describe four new cultivable species, which parasitize various parts of their hosts' intestine, including the thoracic and abdominal part of the midgut, hindgut, and Malpighian tubules. Morphologically, the cultured flagellates vary from relatively short stumpy promastigotes to long slender leptomonad cells. Some species form straphangers (cyst-like amastigotes) both in vivo and in vitro, initially attached to the basal part of the flagellum of the mother cell, from which they subsequently detach. To formally classify this enigmatic monophyletic cosmopolitan clade, we erected Obscuromonas gen. nov., including five species: O. modryi sp. nov. (isolated from the true bug host species Riptortus linearis captured in the Philippines), O. volfi sp. nov. (from Catorhintha selector, Curaçao), O. eliasi sp. nov. (from Graptostethus servus, Papua New Guinea), O. oborniki sp. nov. (from Aspilocoryphus unimaculatus, Madagascar), and O. jaculum comb. nov. (from Nepa cinerea, France). Obscuromonas along with the genus Blastocrithidia belongs to the newly established Blastocrithidiinae subfam. nov.

The Enhanced Expression of Cruzipain-Like Molecules in the Phytoflagellate Phytomonas serpens Recovered From the Invertebrate and Plant Hosts.

Phytomonas serpens is a protozoan parasite that alternates its life cycle between two hosts: an invertebrate vector and the tomato fruit. This phytoflagellate is able to synthesize proteins displaying similarity to the cysteine peptidase named cruzipain, an important virulence factor from Trypanosoma cruzi, the etiologic agent of Chagas disease. Herein, the growth of P. serpens in complex medium (BHI) supplemented with natural tomato extract (NTE) resulted in the increased expression of cysteine peptidases, as verified by the hydrolysis of the fluorogenic substrate Z-Phe-Arg-AMC and by gelatin-SDS-PAGE. Phytoflagellates showed no changes in morphology, morphometry and viability, but the proliferation was slightly reduced when cultivated in the presence of NTE. The enhanced proteolytic activity was accompanied by a significant increase in the expression of cruzipain-like molecules, as verified by flow cytometry using anti-cruzipain antibodies. In parallel, parasites incubated under chemically defined conditions (PBS supplemented with glucose) and added of different concentration of NTE revealed an augmentation in the production of cruzipain-like molecules in a typically dose-dependent way. Similarly, P. serpens recovered from the infection of mature tomatoes showed an increase in the expression of molecules homologous to cruzipain; however, cells showed a smaller size compared to parasites grown in BHI medium. Furthermore, phytoflagellates incubated with dissected salivary glands from Oncopeltus fasciatus or recovered from the hemolymph of infected insects also showed a strong enhance in the expression of cruzipain-like molecules that is more relevant in the hemolymph. Collectively, our results showed that cysteine peptidases displaying similarities to cruzipain are more expressed during the life cycle of the phytoflagellate P. serpens both in the invertebrate and plant hosts.

Parasitism Risk and Infection Alter Host Dispersal.

Dispersal determines the spatial dynamics of host-parasite assemblages, particularly during invasions and disease epidemics. The risk of parasitism may create an incentive for dispersal, but infection is expected to reduce dispersal ability, which may alter the host's dispersal response to biotic stressors, including population density. We measured the dispersal of a semiaquatic insect (Notonecta undulata) in aquatic mesocosms in which we manipulated the presence of ectoparasitic Hydrachnidia mites and infected conspecifics. We found that parasitism risk increases host dispersal propensity. Using a flight assay, we determined that parasite infection reduces host dispersal ability. Finally, we used a mark-release-recapture study to investigate the joint effects of both parasitism risk and parasite infection on host dispersal in a natural, spatially structured population. We found that parasitism risk reduced dispersal probability, eliminated positive density-dependent dispersal, and increased dispersal distance. Infection had no effect on dispersal in the natural population. Our results show that parasites can both increase and decrease the movement rates of their hosts, depending on the ecological context, and can alter the host's dispersal response to other biotic stressors. Future studies should consider the consequences of this double-headed impact of parasites for landscape connectivity, population persistence, and host-parasite coevolution.

A novel endosymbiont-containing trypanosomatid Phytomonas borealis sp. n. from the predatory bug Picromerus bidens (Heteroptera: Pentatomidae).

Here we describe the new trypanosomatid, Phytomonas borealis sp. n., from the midgut of the spiked shieldbugs, Picromerus bidens (Linnaeus), collected in two locations, Novgorod and Pskov Oblasts of Russia. The phylogenetic analyses, based on the 18S rRNA gene, demonstrated that this flagellate is a sister species to the secondary monoxenous Phytomonas nordicus Frolov et Malysheva, 1993, which was concurrently documented in the same host species in Pskov Oblast. Unlike P. nordicus, which can complete its development (including exit to haemolymph and penetration into salivary glands) in Picromerus bidens, the new species did not form any extraintestinal stages in the host. It also did not produce endomastigotes, indispensable for transmission in other Phytomonas spp. These observations, along with the fact that P. bidens overwinters at the egg stage, led us to the conclusion that the examined infections with P. borealis were non-specific. Strikingly, the flagellates from the Novgorod population contained prokaryotic endosymbionts, whereas the parasites from the second locality were endosymbiont-free. This is a first case documenting presence of intracellular symbiotic bacteria in Phytomonas spp. We suggest that this novel endosymbiotic association arose very recently and did not become obligate yet. Further investigation of P. borealis and its intracellular bacteria may shed light on the origin and early evolution of endosymbiosis in trypanosomatids.

Kairomones from Euschistus heros egg masses and their potential use for Telenomus podisi parasitism improvement.

Telenomus podisi Ashmead (Hymenoptera: Scelionidae) is the most important egg parasitoid of Euschistus heros (Fabricius) (Hemiptera: Pentatomidae), and its successful parasitism is related to their searching ability to find suitable hosts under a complex chemical environment using host-reliable cues. Thus, the objective of this study was to elucidate chemical substances on the external layer of E. heros eggs and report its potential kairomonal activity on T. podisi. We tested female wasps in olfactometer system to synthetic compounds obtained from a chemical identification of E. heros egg masses. The synthetic blend was also evaluated in parasitism tests under laboratory and semi-field conditions. We identified 31 substances from egg surface extracts, including monoterpenes, aldehydes and alkanes. Among those compounds, a synthetic solution including camphene, ß-pinene, limonene and benzaldehyde-induced chemotaxic behaviour on the wasps and increased the parasitism on E. heros eggs, either in laboratory or semi-field test, suggesting its potential use to T. podisi manipulation and parasitism improvement.

Parasitism of Adult Pentatomidae by Tachinidae in Soybean in the North Central Region of the United States.

Stink bugs (Hemiptera: Pentatomidae) are agricultural pests of increasing significance in the North Central Region of the United States, posing a threat to major crops such as soybean. Biological control can reduce the need for insecticides to manage these pests, but the parasitism of stink bugs by Tachinidae (Diptera) is poorly characterized in this region. The objective of this study was to evaluate the rate of parasitism of stink bugs by tachinids over 2 yr from nine states across the North Central Region. Parasitism was assessed by quantifying tachinid eggs on the integument of stink bug adults. Parasitism rates (i.e., percent of adult stink bugs with tachinid eggs) were compared across stink bug species, states, stink bug sex, and years. The mean percent parasitism of stink bugs by tachinids was about 6% across the region and did not differ among stink bug species. Mean percent parasitism was significantly higher in Missouri than in northern and western states. In addition, male stink bugs had significantly higher mean percent parasitism than females. Stink bug species commonly found in soybean in the region showed some parasitism and are therefore potentially vulnerable to oviposition by these parasitoids. This is the first study to characterize the level of parasitism of stink bugs by tachinids across the North Central Region.

If host is refractory, insistent parasite goes berserk: Trypanosomatid Blastocrithidia raabei in the dock bug Coreus marginatus.

Here we characterized the development of the trypanosomatid Blastocrithidia raabei in the dock bug Coreus marginatus using light and electron microscopy. This parasite has been previously reported to occur in the host hemolymph, which is rather typical for dixenous trypanosomatids transmitted to a plant or vertebrate with insect's saliva. In addition, C. marginatus has an unusual organization of the intestine, which makes it refractory to microbial infections: two impassable segments isolate the anterior midgut portion responsible for digestion and absorption from the posterior one containing symbiotic bacteria. Our results refuted the possibility of hemolymph infection, but revealed that the refractory nature of the host provokes very aggressive behavior of the parasite and makes its life cycle more complex, reminiscent of that in some dixenous trypanosomatids. In the pre-barrier midgut portion, the epimastigotes of B. raabei attach to the epithelium and multiply similarly to regular insect trypanosomatids. However, when facing the impassable constricted region, the parasites rampage and either fiercely break through the isolating segments or attack the intestinal epithelium in front of the barrier. The cells of the latter group pass to the basal lamina and accumulate there, causing degradation of the epitheliocytes and thus helping the epimastigotes of the former group to advance posteriorly. In the symbiont-containing post-barrier midgut segment, the parasites either attach to bacterial cells and produce cyst-like amastigotes (CLAs) or infect enterocytes. In the rectum, all epimastigotes attach either to the cuticular lining or to each other and form CLAs. We argue that in addition to the specialized life cycle B. raabei possesses functional cell enhancements important either for the successful passage through the intestinal barriers (enlarged rostrum and well-developed Golgi complex) or as food reserves (vacuoles in the posterior end).

Tachinid Fly Parasitism and Phenology of the Neotropical Red-Shouldered Stink Bug, Thyanta perditor (F.) (Heteroptera: Pentatomidae), on the Wild Host Plant, Bidens pilosa L. (Asteraceae).

Field and laboratory studies were conducted with the Neotropical red-shouldered stink bug Thyanta perditor (F.) (Heteroptera: Pentatomidae) aiming to evaluate parasitism incidence on adults by tachinid flies (Diptera: Tachinidae), which were raised in the laboratory for identification. Egg deposition by flies on adult body surface was mapped. In addition, nymph and adult incidence on the wild host plant black jack, Bidens pilosa L. (Asteraceae), during the vegetative and the reproductive periods of plant development was studied. Seven species of tachinid flies were obtained: Euthera barbiellini Bezzi (73% of the total) and Trichopoda cf. pictipennis Bigot (16.7%) were the most abundant; the remaining five species, Gymnoclytia sp.; Phasia sp.; Strongygaster sp.; Cylindromyia cf. dorsalis (Wiedemann); and Ectophasiopsis ypiranga Dios & Nihei added 10.3% of the total. Tachinid flies parasitism on T. perditor adults was significantly greater on the dorsal compared to the ventral body surface. On the dorsal surface, the pronotum was significantly preferred and the wings the least preferred site. No differences were observed on the number of tachinid fly eggs deposited on wings, considering the "under" and "above" sites. Results indicated a significantly greater number of nymphs on mature compared to immature seeds. Adults significantly preferred immature compared to mature seeds; both were less abundant on leaves/stems and inflorescences.

Impact of Nosema maddoxi on the survival, development, and female fecundity of Halyomorpha halys.

Nosema maddoxi Becnel, Solter, Hajek, Huang, Sanscrainte, & Estep, a microsporidian species native to the United States, has been found infecting the invasive brown marmorated stink bug, Halyomorpha halys (Stål). Microsporidian infections in insects often shorten lifespans, decrease fecundity, prolong development, and stunt growth. This study was conducted to determine the impact of N. maddoxi on H. halys fitness. Adult females (2 doses) and nymphs (1 dose) drank suspensions of N. maddoxi spores to promote infection. Adult females receiving a high dose died faster than the controls. Nosema maddoxi infections impacted female egg production and egg viability at both doses compared with the controls. Infections were transmitted to 34.9% of adult males caged with infected females. As the number of days after inoculation increased, infection intensity (# spores found within an infected individual) for both adult treatments transitioned from low-intensity to high-intensity. Infected nymphs died significantly sooner than the controls. Of the treated nymphs, 55.9% died before molting into the fourth instar and only 26.5% eclosed to adults. Nymphal development rate and size were not impacted by N. maddoxi infection. These results indicate that N. maddoxi infection can negatively impact the lifespan of adult females, female fecundity, egg viability, and nymphal survival, which we hypothesize would negatively impact H. halys population densities.

Effect of the Ages of Parasitoid and Host Eggs on Telenomus podisi (Hymenoptera: Platygastridae) Parasitism.

This study investigates the influence of parasitoid age and egg age of the hosts Euschistus heros (Fabricius) and Dichelops melacanthus (Dallas) on parasitism of Telenomus podisi Ashmead. Six separate bioassays were conducted: parasitism on eggs of E. heros (bioassay 1) and D. melacanthus (bioassay 2) by T. podisi females of different age (1, 5, and 10 days old); parasitism by T. podisi on eggs of different age (1, 2, 3, 4, and 5 days of embryonic development) of the hosts E. heros (bioassay 3) and D. melacanthus (bioassay 4); preference of T. podisi females for eggs at different embryonic developmental stages (eggs of 1, 2, 3, 4, and 5 days) of the hosts E. heros (bioassay 5) and D. melacanthus (bioassay 6). The age of T. podisi females and their hosts affected parasitism on both E. heros and D. melacanthus eggs. Overall, the parasitism rate was higher in older than younger parasitoids, independent of the tested host species, and host eggs between 1 and 3 days old were similarly parasitized. Thus, in T. podisi mass rearing facilities, it is recommended to use older adults (5 to 10 days old) as mother wasps to increase parasitism on the offered eggs. In addition, when hosts are completely absent in the field, or climatic conditions are unfavorable for release, mass-reared adults can be kept in the laboratory (25°C) for up to 10 days for later release in the field without any impairment of their subsequent parasitism performance.

The Fitness and Economic Benefits of Rearing the Parasitoid Telenomus podisi Under Fluctuating Temperature Regime.

Successful biological control requires detailed knowledge about the mass rearing conditions of the control agents in order to ensure higher quality of field-released insects. Thus, we investigated whether rearing fluctuating thermal condition would affect the fitness and costs of the parasitoid wasp Telenomus podisi Ashmead (a biocontrol agent used for controlling the Neotropical brown stink bug Euschistus heros (Fabricius)) when compared with parasitoid reared at constant temperature condition, which is commonly used in insect facilities. Parasitoids were reared under either constant (continuous exposure at 25 ± 2°C) or fluctuating temperature conditions (i.e., 30 ± 2°C during day and 20 ± 2°C at night) during four consecutive generations. Our results indicated that tested fluctuating temperature is more suitable for rearing of T. podisi as such temperature condition not only resulted in fitness benefits (e.g., shorter developmental time, longer female longevity, higher fecundity/fertility) but also reduced (approximately 23.5%) the estimated costs for producing the parasitoids. Furthermore, rearing T. podisi under fluctuating temperatures improved tolerance to low constant temperatures (i.e., 20°C) without changing the tolerance to constant high temperatures (30°C) in the fourth generation. Surprisingly, even parasitoids that developed under fluctuating thermal conditions performed better than those reared at constant temperature of 25°C. Collectively, our findings suggest that T. podisi reared under fluctuating thermal condition can tolerate better fluctuating temperatures that normally occur both during long periods of transport and in agricultural ecosystems, which will increase the quality and productivity of mass-reared T. podisi for inundative releases.

Natural infection by the protozoan Leptomonas wallacei impacts the morphology, physiology, reproduction, and lifespan of the insect Oncopeltus fasciatus.

Trypanosomatids are protozoan parasites that infect thousands of globally dispersed hosts, potentially affecting their physiology. Several species of trypanosomatids are commonly found in phytophagous insects. Leptomonas wallacei is a gut-restricted insect trypanosomatid only retrieved from Oncopeltus fasciatus. The insects get infected by coprophagy and transovum transmission of L. wallacei cysts. The main goal of the present study was to investigate the effects of a natural infection by L. wallacei on the hemipteran insect O. fasciatus, by comparing infected and uninfected individuals in a controlled environment. The L. wallacei-infected individuals showed reduced lifespan and morphological alterations. Also, we demonstrated a higher infection burden in females than in males. The infection caused by L. wallacei reduced host reproductive fitness by negatively impacting egg load, oviposition, and eclosion, and promoting an increase in egg reabsorption. Moreover, we associated the egg reabsorption observed in infected females, with a decrease in the intersex gene expression. Finally, we suggest alterations in population dynamics induced by L. wallacei infection using a mathematical model. Collectively, our findings demonstrated that L. wallacei infection negatively affected the physiology of O. fasciatus, which suggests that L. wallacei potentially has a vast ecological impact on host population growth.

Trissolcus japonicus (Hymenoptera: Scelionidae) Causes Low Levels of Parasitism in Three North American Pentatomids Under Field Conditions.

Trissolcus japonicus (Ashmead), an Asian parasitoid of Halyomorpha halys (Stål) (Hemiptera: Pentatomidae), was first detected in North America in 2014. Although testing in quarantine facilities as a candidate for classical biological control is ongoing, adventive populations have appeared in multiple sites in the United States, Canada, and Europe. Extensive laboratory testing of T. japonicus against other North American pentatomids and H. halys has revealed a higher rate of parasitism of H. halys, but not complete host specificity. However, laboratory tests are necessarily artificial, in which many host finding and acceptance cues may be circumvented. We offered sentinel egg masses of three native pentatomid (Hemiptera: Pentatomidae) pest species (Chinavia hilaris (Say), Euschistus conspersus Uhler, and Chlorochroa ligata (Say)) in a field paired-host assay in an area with a well-established adventive population of T. japonicus near Vancouver, WA. Overall, 67% of the H. halys egg masses were parasitized by T. japonicus during the 2-yr study. Despite the 'worst case' scenario for a field test (close proximity of the paired egg masses), the rate of parasitism (% eggs producing adult wasps) on all three native species was significantly less (0.4-8%) than that on H. halys eggs (77%). The levels of successful parasitism of T. japonicus of the three species are C. hilaris > E. conspersus > C. ligata. The potential impact of T. japonicus on these pentatomids is probably minimal.

Changes in vitellogenin expression caused by nematodal and fungal infections in insects.

This study examined the expression and role of vitellogenin (Vg) in the body of the firebug Pyrrhocoris apterus (Heteroptera, Insecta) during infection elicited by two entomopathogenic organisms, the nematode Steinernema carpocapsae and the fungus Isaria fumosorosea Infection by S. carpocapsae significantly upregulated Vg mRNA expression in the male body. The corresponding increase in Vg protein expression was also confirmed by electrophoretic and immunoblotting analyses. Remarkably, in females, the opposite tendency was noted. Nematodal infection significantly reduced both Vg mRNA and Vg protein expression levels in fat body and hemolymph, respectively. We speculate that infection of reproductive females reduces Vg expression to a level that is still sufficient for defense, but is insufficient for reproduction. This circumstance reduces energy expenditure and helps the individual to cope with the infection. Importantly, purified Vg significantly inhibited growth of Xenorhabdus spp., an entomotoxic bacteria isolated from S. carpocapsae. However, the effect of Vg against I. fumosorosea was not so obvious. The fungus significantly stimulated Vg gene expression in males; however, a similar increase was not recapitulated at the protein level. Nevertheless, in females, both mRNA and protein Vg levels were significantly reduced after the fungal infection. The obtained data demonstrate that Vg is probably an important defense protein, possibly with a specific activity. This considerably expands the known spectrum of Vg functions, as its primary role was thought to be limited to regulating egg development in the female body.

Development of Phytomonas lipae sp. n. (Kinetoplastea: Trypanosomatidae) in the true bug Coreus marginatus (Heteroptera: Coreidae) and insights into the evolution of life cycles in the genus Phytomonas.

Here we described a new trypanosomatid species, Phytomonas lipae, parasitizing the dock bug Coreus marginatus based on axenic culture and in vivo material. Using light and electron microscopy we characterized the development of this flagellate in the intestine, hemolymph and salivary glands of its insect host. The intestinal promastigotes of Phytomonas lipae do not divide and occur only in the anterior part of the midgut. From there they pass into hemolymph, increasing in size, and then to salivary glands, where they actively proliferate without attachment to the host's epithelium and form infective endomastigotes. We conducted molecular phylogenetic analyses based on 18s rRNA, gGAPDH and HSP83 gene sequences, of which the third marker performed the best in terms of resolving phylogenetic relationships within the genus Phytomonas. Our inference demonstrated rather early origin of the lineage comprising the new species, right after that of P. oxycareni, which represents the earliest known branch within the Phytomonas clade. This allowed us to compare the development of P. lipae and three other Phytomonas spp. in their insect hosts and reconstruct the vectorial part of the life cycle of their common ancestor.