Agamermis sp. (Nematoda: Mermithidae) parasitizing Armadillidium vulgare (Crustacea: Isopoda) in Argentina.

The Mermithidae is a family of nematodes parasitic in many kinds of insects, spiders, leeches, crustaceans and other invertebrates throughout the world. While conducting an assay with entomopathogenic nematodes, we found Armadillidium vulgare (Crustacea: Isopoda) individuals to be infected with Agamermis sp., marking the fourth known discovery of a mermithid infection in the order Isopoda. In this work, we contribute with an 18S rDNA sequence of the isolated nematode and the morphological and morphometrical characterization of the juveniles.

Ovomermis sinensis parasitism arrests midgut replacement by altering ecdysone and juvenile hormone in Helicoverpa armigera larvae.

Many entomopathogens regulate the development of their insect hosts. However, the influence of mermithid nematodes on the development of their host remains unclear. In the current study, we provide insights into how Ovomermis sinensis parasitism affects the development of Helicoverpa armigera. We observed that O. sinensis arrests host development, as evidenced by the reduced body size and failure of Helicoverpa armigera to pupate. Moreover, midgut replacement of the host was significantly blocked by parasitism. Furthermore, juvenile hormone (JHIII) titers of the host were dramatically elevated by parasitism, but JH esterase (JHE) activities were strongly inhibited. By contrast, steroid hormone (20-hydroxyecdysone, 20E) titers of the host were significantly depressed by parasitism on days 4-6. The expression profiles of hormone-related genes in the host also showed similar patterns with the hormone titer. For this reason, rescue experiments were performed by injecting 20E and JHIII into developmentally arrested hosts. Notably, the midgut replacement of the host was rescued by the injection of 20E, whereas JHIII injection resulted in negative effects. Altogether, O. sinensis arrests H. armigera midgut replacement by reducing 20E and maintaining JH, thereby causing developmental arrests. Our study is the first report of the possible mechanism of mermithid nematodes in regulating insect development.

Behavior of Aedes albifasciatus (Diptera: Culicidae) larvae from eggs with different dormancy times and its relationship with parasitism by Strelkovimermis spiculatus (Nematoda: Mermithidae).

Aedine mosquitoes go through unfavorable periods as dormant eggs. However, extended dormancy times lead to a depletion of egg reserves, which might be partially compensated by changes in larval-feeding behavior. Changes in larval behavior may also be induced by parasitism by mermithids probably as a strategy to reduce the impact of the parasite on the host. The most abundant floodwater mosquito in southern South America is Aedes albifasciatus (Diptera: Culicidae), a species naturally parasitized by Strelkovimermis spiculatus (Nematoda: Mermithidae). This study aimed to evaluate the behavior of fourth-instar larvae of Ae. albifasciatus from eggs with 2, 4, and 6 months of dormancy, parasitized or not by S. spiculatus. To achieve this, eight categories of behavior were defined, and then, each individual was observed for 5 min, and its behaviors and their duration were recorded. The behaviors with the highest percentage of observation time were still (50.1%) and wriggle swimming (33.6%), while those with the lowest percentage of observation time were those involving active foraging (less than 8%). A great variability in the behaviors was observed among individuals within the same treatment. The type and duration of each behavior did not vary in relation to egg dormancy time or parasitism, but parasitism affected the level of larval activity. Parasitized larvae performed, on average, fewer behaviors than non-parasitized larvae. This result supports the hypothesis that S. spiculatus parasitism affects the behavior of Ae. albifasciatus larvae by reducing their level of activity.

Dive into the sea: first molecular phylogenetic evidence of host expansion from terrestrial/freshwater to marine organisms in Mermithidae (Nematoda: Mermithida).

We report the first mermithid nematode found to be parasitic in a marine tanaidacean crustacean. Ten host tanaidaceans were collected from a depth of 52 m in Otsuchi Bay, Iwate, Japan, north-western Pacific, and identified as a species in the tanaidid genus Zeuxo Templeton, 1840. Nematodes occurred in the host's body cavity; in one case, at least two individuals inhabited a single host. We provide a brief description and illustrations of the morphology of the nematode. In a phylogenetic reconstruction based on the 18S ribosomal RNA gene, the nematode nested in a clade otherwise containing mermithids from terrestrial or freshwater hosts, showing an expansion in host utilization in Mermithidae Braun, 1883 from terrestrial/freshwater hosts to a marine organism.

The general architecture of black fly-parasite interactions: Parasitism in lotic systems at a continental scale.

We examined the general architecture of interactions between stream-dwelling larval black flies (Diptera: Simuliidae) and their common parasites in 1736 collections across North America. Mermithid nematodes (family Mermithidae), microsporidia (phylum Microsporidia), and the fungus Coelomycidium simulii Debaisieux (phylum Blastocladiomycota) infected larval black flies. We found similar continental distributions for these three parasite taxa across North America. At least one of these taxa was represented in 42.2% of all black fly collections. Species interactions in ecological networks typically imply that each link between species is equally important. By employing quantitative measures of host susceptibilities and parasite dependencies, we provide a more complete structure for host-parasite networks. The distribution of parasite dependencies and host susceptibilities were right-skewed, with low values indicating that most dependencies (parasites) and susceptibilities (hosts) were weak. Although regression analysis between host frequency and parasite incidence were highly significant, frequency analysis suggested that the distributions of parasites differ significantly among the four most common and closely related (same subgenus) species of hosts. A highly significant pattern of nestedness in our bipartite host-parasite network indicated that specialized parasites (i.e., those that interact with few host species) tend to occur as subsets of the most common hosts.

Sexually anomalous individuals of the black fly Simulium trangense (Diptera: Simuliidae) infected with mermithid parasites (Nematoda: Mermithidae).

Sexually anomalous individuals, typically intersexes or gynandromorphs, bear a mixture of male and female traits. Twelve sexually anomalous individuals of the black fly Simulium (Gomphostilbia) trangense Jitklang, Kuvangkadilok, Baimai, Takaoka & Adler were discovered among 49 adults reared from pupae. All 12 sexually anomalous adults were parasitized by mermithid nematodes, although five additional parasitized adults had no overt external anomalies. Sequence analysis of the 18S rRNA gene revealed that the mermithids, possibly representing a new species, are related to Mesomermis spp., with genetic distances of 5.09-6.87%. All 12 anomalous individuals had female phenotypical traits on the head, thorax, forelegs, midlegs, and claws, but male features on the left and right hind basitarsi. One individual had mixed male and female genitalia. The findings are in accord with the trend that mermithid infections are associated with sexually anomalous adult black flies.

Susceptibility of the floodwater mosquito Aedes albifasciatus from eggs of different dormancy times to the nematode parasite Strelkovimermis spiculatus.

This study aimed to evaluate the effects of egg dormancy times on susceptibility of larvae of the floodwater mosquito Aedes albifasciatus (Diptera: Culicidae) to parasitism by their natural enemy Strelkovimermis spiculatus (Nematoda: Mermithidae) and on their life history traits. Aedes albifasciatus eggs stored for 2, 4, 6, 8 and 10 months were hatched, and the larvae either exposed to S. spiculatus (treatment group) or not exposed (control group). Egg dormancy time had a negative effect on the retention of parasites, but no effect on the prevalence and intensity of parasitism or the melanization of nematodes. The survival to adulthood of control individuals decreased as dormancy time increased, whereas that of exposed individuals that remained uninfected was constant and low. A trend towards increasing development times with longer dormancy times was detected in the control group, but not in the exposed noninfected group. The results suggest nonconsumptive effects of parasites in exposed but not infected larvae from eggs with short dormancy times. In contrast, the relatively low fitness of larvae from eggs with long dormancy times regardless of their contact with the nematodes may be the result of the nutritional deprivation during the egg stage.

Small but large enough: structural properties of armless mitochondrial tRNAs from the nematode Romanomermis culicivorax.

As adapter molecules to convert the nucleic acid information into the amino acid sequence, tRNAs play a central role in protein synthesis. To fulfill this function in a reliable way, tRNAs exhibit highly conserved structural features common in all organisms and in all cellular compartments active in translation. However, in mitochondria of metazoans, certain dramatic deviations from the consensus tRNA structure are described, where some tRNAs lack the D- or T-arm without losing their function. In Enoplea, this miniaturization comes to an extreme, and functional mitochondrial tRNAs can lack both arms, leading to a considerable size reduction. Here, we investigate the secondary and tertiary structure of two such armless tRNAs from Romanomermis culicivorax. Despite their high AU content, the transcripts fold into a single and surprisingly stable hairpin structure, deviating from standard tRNAs. The three-dimensional form is boomerang-like and diverges from the standard L-shape. These results indicate that such unconventional miniaturized tRNAs can still fold into a tRNA-like shape, although their length and secondary structure are very unusual. They highlight the remarkable flexibility of the protein synthesis apparatus and suggest that the translational machinery of Enoplea mitochondria may show compensatory adaptations to accommodate these armless tRNAs for efficient translation.

Adaptation of the Romanomermis culicivorax CCA-Adding Enzyme to Miniaturized Armless tRNA Substrates.

The mitochondrial genome of the nematode Romanomermis culicivorax encodes for miniaturized hairpin-like tRNA molecules that lack D- as well as T-arms, strongly deviating from the consensus cloverleaf. The single tRNA nucleotidyltransferase of this organism is fully active on armless tRNAs, while the human counterpart is not able to add a complete CCA-end. Transplanting single regions of the Romanomermis enzyme into the human counterpart, we identified a beta-turn element of the catalytic core that-when inserted into the human enzyme-confers full CCA-adding activity on armless tRNAs. This region, originally identified to position the 3'-end of the tRNA primer in the catalytic core, dramatically increases the enzyme's substrate affinity. While conventional tRNA substrates bind to the enzyme by interactions with the T-arm, this is not possible in the case of armless tRNAs, and the strong contribution of the beta-turn compensates for an otherwise too weak interaction required for the addition of a complete CCA-terminus. This compensation demonstrates the remarkable evolutionary plasticity of the catalytic core elements of this enzyme to adapt to unconventional tRNA substrates.

Molecular characterization of immune responses of Helicoverpa armigera to infection with the mermithid nematode Ovomermis sinensis.

BACKGROUND: Mermithid nematodes, such as Ovomermis sinensis, display a broad host range including some lepidopteran pests. Infective juveniles penetrate their host through the cuticle, complete their growth within the hemocoel and eventually kill the host upon their emergence. Hence, mermithid nematodes are considered potential biological control agents of insect pests. Our previous data indicate that the infection rate of O. sinensis on cotton bollworm (Helicoverpa armigera) is low, which may be largely due to the strong immune system of the host. However, current knowledge on the interactions of mermithid nematodes with their hosts and the mechanisms employed by hosts to defend themselves against mermithid nematodes is limited. RESULTS: Here, we investigated the response of H. armigera to O. sinensis infection. Parasitism by O. sinensis caused a sharp decline in the survival rate of H. armigera. The hemocytic phagocytosis ability, antibacterial activity, and phenoloxidase (PO) activity in plasma of H. armigera increased at 1 d post parasitism (dpp) but decreased at 3 dpp. Further, we investigated gene expression in the fat body of parasitized and non-parasitized H. armigera larvae at 1, 3, and 5 dpp using a digital gene expression system. In total, 41, 60 and 68 immune-related differentially expressed genes were identified at 1, 3, and 5 dpp, respectively. These genes encoded pattern recognition receptors (PRRs), antimicrobial peptides (AMPs), serine proteases (SPs), SP inhibitors, mucins and other immune-related proteins. The expression of most PRRs, AMPs, SPs, and mucins was upregulated in the fat body of larvae at 1 dpp, downregulated at 3 dpp, and then again upregulated at 5 dpp by O. sinensis. The increased expression of SP inhibitors may contribute to the inhibited PO activity at 5 dpp. CONCLUSIONS: This study demonstrates that parasitism by O. sinensis modulates the immune reaction of the host H. armigera by altering the expression of immune-related genes. Our data provide a basis for future investigation of the molecular mechanisms employed by the mermithid nematode O. sinensis to modulate the immunity of the host H. armigera. These data will also likely facilitate the improvement of success in parasitism of H. armigera by O. sinensis.

A molecular war: convergent and ontogenetic evidence for adaptive host manipulation in related parasites infecting divergent hosts.

Mermithids (phylum Nematoda) and hairworms (phylum Nematomorpha) somehow drive their arthropod hosts into water, which is essential for the worms' survival after egression. The mechanisms behind this behavioural change have been investigated in hairworms, but not in mermithids. Establishing a similar mechanistic basis for host behavioural change between these two distantly related parasitic groups would provide strong convergent evidence for adaptive manipulation and insight into how these parasites modify and/or create behaviour. Here, we search for this convergence, and also contrast changes in physiology between hosts infected with immature and mature mermithids to provide the first ontogenetic evidence for adaptive manipulation by disentangling host response and pathology from the parasite's apparent manipulative effects. We used SWATH-mass spectrometry on brains of Forficula auricularia (earwig) and Bellorchestia quoyana (sandhopper), infected with the mermithids Mermis nigrescens and Thaumamermis zealandica, respectively, at both immature and mature stages of infection, to quantify proteomic changes resulting from mermithid infection. Across both hosts (and hairworm-infected hosts, from earlier studies), the general function of dysregulated proteins was conserved. Proteins involved in energy generation/mobilization were dysregulated, corroborating reports of erratic/hyperactive behaviour in infected hosts. Dysregulated proteins involved in axon/dendrite and synapse modulation were also common to all hosts, suggesting neuronal manipulation is involved in inducing positive hydrotaxis. Furthermore, downregulation of CamKII and associated proteins suggest manipulation of memory also contributes to the behavioural shift.

Let's go swimming: mermithid-infected earwigs exhibit positive hydrotaxis.

Certain species of parasites have the apparent ability to alter the behaviour of their host in order to facilitate the completion of their own life cycle. While documented in hairworms (phylum Nematomorpha), the ability for mermithid parasites (from the sister phylum Nematoda) to force hosts to enter water remains more enigmatic. Here, we present the first experimental evidence in a laboratory setting that an insect which normally never enters open water (the European earwig Forficula auricularia) will readily enter the water when infected with a mermithid nematode (Mermis nigrescens). Only adult mermithids appear capable of inducing this polarising shift in behaviour, with mermithid length being a very strong predictor of whether their host enters water. However, mermithid length was only weakly associated with how long it took an earwig to enter water following the beginning of a trial. Considering the evidence presented here and its alignment with a proteomic investigation on the same host-parasite system, this study provides strong evidence for adaptive behavioural manipulation and a foundational system for further behavioural and mechanistic exploration.

Behavior manipulation of mosquitoes by a mermithid nematode.

We examined manipulation of mosquito behavior by the parasitic mermithid nematode, Strelkovimermis spiculatus. This nematode species typically infects early instar host larvae and emerges after parasitic development to kill last-instar larvae. Parasitized adults, however, have occasionally been reported from field collections. We obtained low rates (1.7-11.5%) of parasitized adults in laboratory exposures only when Culex pipiens pipiens fourth-instar larvae nearing pupation were exposed to infective nematodes. This did not allow an adequate interval for parasitic development in immature host stages. Parasitized adult females in a multiple-choice assay were three times more likely to seek water than a blood source (63.1 vs. 20.5%), whereas uninfected females were twice as likely to seek blood than water (64%3.9 vs. 32.6%). This altered host behavior benefits the parasite by providing the only mechanism for dispersal and colonization of new host habitats while concurrently avoiding risks from the defensive behaviors associated with blood-feeding. Behavioral alternation in Cx. p. pipiens larval hosts was also examined using larvae infected as second instars to allow for a normal duration of parasitic development. As larvae neared pupation and parasite emergence, parasitized larvae became more spatially aggregated than unparasitized larvae. This altered host behavior benefits the parasite by providing a corresponding increase in post-parasite aggregation, which facilitates formation of large mating clusters and concomitantly reproductive success. Parasites derive fitness gains by overriding host autonomy, whereas hosts have zero fitness once parasitism is established, suggesting a coevolutionary response is inoperative and that the behavioral modifications may be adaptive.

Rarely reported, widely distributed, and unexpectedly diverse: molecular characterization of mermithid nematodes (Nematoda: Mermithidae) infecting bumble bees (Hymenoptera: Apidae: Bombus) in the USA.

Mermithid nematodes (Nematoda: Mermithida: Mermithidae) parasitize a wide range of both terrestrial and aquatic invertebrate hosts, yet are recorded in bumble bees (Insecta: Hymenoptera: Apidae: Bombus) only six times historically. Little is known about the specific identity of these parasites. In a single-season nationwide survey of internal parasites of 3646 bumble bees, we encountered six additional instances of mermithid parasitism in four bumble bee species and genetically characterized them using two regions of 18S to identify the specific host-parasite relationships. Three samples from the northeastern USA are morphologically and genetically identified as Mermis nigrescens, whereas three specimens collected from a single agricultural locality in the southeast USA fell into a clade with currently undescribed species. Nucleotide sequences of the V2-V6 region of 18S from the southeastern specimens were 2.6-3.0% divergent from one another, and 2.2-4.0% dissimilar to the nearest matches to available data. The dearth of available data prohibits positive identification of this parasite and its affinity for specific bumble bee hosts. By doubling the records of mermithid parasitism of bumble bee hosts and providing genetic data, this work will inform future investigations of this rare phenomenon.

Induction of hemocyte apoptosis by Ovomermis sinensis: Implications for host immune suppression.

The entomopathogenic nematode, Ovomermis sinensis, is a parasite of some common lepidopteran pests. O. sinensis is able to overcome the immune system of its hosts and eventually kill the hosts when it emerges. We provide insight into how the mermithid nematode overcomes the immune response of the host Helicoverpa armigera. Our results indicate that O. sinensis actively inhibits host immune responses as evidenced by hemocyte nodulation, spreading behavior, lysozyme activity and melanization. However, O. sinensis did not inhibit host immune responses through immune gene activation. Moreover, the research on the immune depressive strategies of O. sinensis revealed that the parasite did not inhibit host effector molecules, but did reduce the number of hemocytes. Flow cytometry analysis revealed that the host hemocytes were apoptotic within 24 h, and no hemocytes were present after 72 h. In addition, our in vivo and in vitro studies showed that the survival rate of O. sinensis was increased when hemocyte proliferation was inhibited. Our findings suggest that O. sinensis inhibited host immune responses by inducing apoptosis of host hemocytes.

Is the morphology of Culicoides intersexes parasitized by mermithid nematodes a parasite adaptation? A morphometric approach to Culicoides circumscriptus (Diptera: Ceratopogonidae).

Mermithidae is a family of endoparasitic nematodes known to cause intersexuality in arthropods. Intersexes of the genus Culicoides parasitized by mermithids have been the object of several studies aiming to describe their particular morphology. Culicoides intersexes are specimens with male genitalia and feminized sexually dimorphic structures, i.e. antennae, mouthparts and wings. To date, these specimens have only been described qualitatively and a quantitative approach supported by statistical analysis is lacking. Here we conduct morphometric analyses of sexually dimorphic structures in a sample of Culicoides circumscriptus that includes 34 intersexes with the aim of describing precisely the intersexual morphology. The morphology of antennae and the mouthparts was studied by multivariate statistical analysis of linear measures, and wing form by implementing geometric morphometrics techniques. While intersex wings proved to have a similar size to male wings, their shape was intermediate between males and females. However, when allometric shape variation was removed, the wing shape of intersexes was almost identical to that of females. The intersex antennae were morphometrically of the female type, especially when size variation was considered. In contrast, the measured mouthparts (the labrum and the third palpal segment) were halfway between males and females, even when body size was considered. Overall, the antennae and the wings showed a higher degree of feminization than the mouthparts. These findings indicate that the degree of feminization depends both on the morphological structure and on body size. Moreover, we propose that the feminization of the wings and antennae has an adaptive meaning for the parasite, which would favor female-like traits in order to access more easily its breeding sites, where the parasite has plenty of new hosts to infect. Female-like antennae would be beneficial to detect these sites, while having female-like wings would favor the host's capacity of dispersal to these sites.

Inequalities in body size among mermithid nematodes parasitizing earwigs.

Variation among body sizes of adult parasitic worms determines the relative genetic contribution of individuals to the next generation as it affects the effective parasite population size. Here, we investigate inequalities in body size and how they are affected by intensity of infection in Mermis nigrescens (Mermithidae: Nematoda) parasitizing the European earwig Forficula auricularia in New Zealand. Among a population of pre-adult worms prior to their emergence from the host, we observed only modest inequalities in body length; however, among worms sharing the same individual host, inequalities in body sizes decreased with increasing intensity of infection. Thus, the more worms occurred in a host, the more the second-longest, third-longest and even fourth-longest worms approached the longest worm in body length. This pattern, also known from another mermithid species, suggests that worms sharing the same host may have infected it roughly simultaneously, when the host encountered a clump of eggs in the environment. Thus, the life history and mode of infection of the parasite may explain the modest inequalities in the sizes achieved by pre-adult worms, which are lower than those reported for endoparasitic helminths of vertebrates.

First report of a mermithid nematode infecting the invasive Megacopta cribraria (Hemiptera: Plataspidae) in the United States.

Megacopta cribraria (Hemiptera: Plataspidae) has become a pest of soybean, Glycine max (L.), in the United States. While several natural enemies of M. cribraria have been reported, our study is the first to report nematodes beneath the pleural membranes in the abdominal cavities of adults. Morphological and molecular analyses suggest this nematode belongs to the family Mermithidae. This first report of a nematode infection in M. cribraria adds to the current inventory of enemies attacking this insect. Our observations provide a basis for future research to examine the impact of nematodes on M. cribraria mortality and to investigate their capacity to reduce populations.

Transmission of a pathogenic virus (Iridoviridae) of Culex pipiens larvae mediated by the mermithid Strelkovimermis spiculatus (Nematoda).

Little progress been made in elucidating the transmission pathway of the invertebrate iridescent virus (MIV). It has been proposed that the MIV has no active means to enter the mosquito larva. We have previously found that the presence of the mermithid nematode Strelkovimermis spiculatus is associated with MIV infection in Culex pipiens under field conditions. In the present study, we evaluated the transmission of MIV to C. pipiens larvae mediated by S. spiculatus and several factors involved in this pathway (mosquito instars, nematode:mosquito larva ratio, amount of viral inoculum). Our results indicate that S. spiculatus functions as an MIV vector to C. pipiens larvae and seems to be an important pathway of virus entry into this system. Moreover, TEM images of S. spiculatus exposed to the viral suspension showed no infections inside the nematode but showed that viral particles are carried over the cuticle of this mermithid. This explains the correspondence between MIV infection and the factors that affect the parasitism of S. spiculatus in C. pipiens larvae.

Factors influencing spatial variation and abundance of a mermithid parasite in sand hoppers.

The impact of parasites on host population dynamics depends on local abundance of the parasites, which may vary considerably across spatial scales. In sand hopper populations, mermithid parasites have major impacts on host dynamics, which may vary among spatially separated populations due to the sand hopper's wide, patchy distribution. The present study compared the abundance and biomass of a mermithid parasite (Thaumamermis zealandica Poinar et al., 2002) in sand hoppers (Bellorchestia quoyana (Milne-Edwards)) both within and among disconnected beaches. In addition, several variables were measured and tested as potentially important predictors of the parasite abundance and biomass. It was found that geographic isolation may only be responsible for minor differences in parasite populations compared with other factors. Host size was identified as the most important predictor of mermithid parasite abundance, but epibiont abundance, kelp patch mass and host density were poor predictors of abundance. These factors were also poor predictors of parasite biomass in hosts. This study further supports the notion that studies aiming to elucidate population dynamics or patterns should sample thoroughly across both spatial and temporal scales.