Morphological and molecular characterization of a new microsporidium, Janacekia tainanus n. sp. from the adipose tissue of Kiefferulus tainanus (Diptera: Chironomidae) in China.

We reported a new microsporidium Janacekia tainanus n. sp. from the adipose tissue of the midge Kiefferulus tainanus Kieffer, 1912 collected from a eutrophic pond in Daye city, Hubei Province, China. Infected chironomid larvae with hypertrophied adipose tissue exhibited porcelain-white. All developmental stages possessed large nuclei. The earliest stages observed were diplokaryotic meronts which were in direct contact with the host adipocyte cytoplasm. Diplokaryotic meronts developed into sporonts with the deposition of electron-dense coagulum on their surface. Multinucleate sporogonial plasmodia developed into uninucleate sporoblasts by the rosette-like division. Mature spores were oval and monokaryotic, measuring 6.14 ± 0.27 (5.65-6.67) µm long and 3.71 ± 0.12 (3.43-3.98) µm wide. Bipartite polaroplast consisted of a narrow anterior lamella and a wide posterior lamella. Isofilar polar filaments coiled 13-17 turns and arranged in one row. The exospore was thin and of no stratification, but remarkably covered with tubular secretions. The electron-lucent endospore was thick and measured 145-352 nm wide. Phylogenetic analysis based on the obtained SSU rDNA sequence indicated that the present species clustered closely with Jirovecia sinensis, a species with rod-shaped mature spores isolated from the coelomocytes of Branchiura sowerbyi. Consistent with the previous result, the monophyletic clade of Jirovecia-Bacillidium-Janacekia was sister to Pseudonosema clade and then collectively nested within Clade V of Class Aquasporidia sensu Vossbrinck and Debrunner-Vossbrinck (2005). The novel species did not form an independent monophyletic lineage with the congener, Janacekia debaisieuxi. Based on the morphological characters and ultrastructural features, as well as SSU rDNA-inferred phylogenetic relationships, a new species in the genus Janacekia, Janacekia tainanus n. sp. was designated. This is the first report of aquatic arthropod-infecting microsporidia in China.

Prevalence and dynamics of the zoosporic pathogen Catenaria uncinata in a natural population of the midge Glyptotendipes lobiferus.

A qPCR assay specific for zoospores of Catenaria uncinata, a fungal parasite in eggs of the midge Glyptotendipes lobiferus, was developed and used in parallel with traditional microscopic methods in a season-long study of a C. uncinata/G. lobiferus association in a local pond. Twenty-six consecutive weekly collections of egg masses were screened with a microscope to obtain percentages of infection and mortality in organogenetic egg masses and weekly water samples were processed by absolute quantification using qPCR to obtain estimates of zoospore density. Overall, 36.0% of G. lobiferus egg masses were infected to varying degrees and 11.2% of eggs were killed by C. uncinata. Continuous infection of egg masses occurred during a 6-wk period in May-June and a 7-wk period in September-October. Infection by C. uncinata was absent during a 10-week interval between periods of infection. Abrupt declines in zoospore density occurred during both infection periods and occurred only when water temperatures met or exceeded the viability threshold for zoospores (⩾31.0°C). The episodic death of zoospores during weeks in which egg infection and mortality levels were continuous likely resulted from distribution of zoospores throughout the water column and a temperature gradient in which zoospores sampled near the surface were subjected to lethal temperatures while non-sampled zoospores at lower depths were provided low temperature sanctuary. The hiatus of infection during the 10-week interval was likely due to lethal temperatures throughout the water column as average water temperatures exceeded 31.0°C over the period. A positive correlation between weekly zoospore densities obtained from qPCR and levels of infection/mortality in egg masses obtained from counts with a microscope supports the use of the qPCR assay alone in future studies that can rapidly and accurately determine parasite presence, prevalence and geographical range.

Evaluation of a commercial Bacillus thuringiensis var. israelensis formulation for the control of chironomid midge larvae (Diptera: Chironomidae) in establishing rice crops in south-eastern Australia.

A commercial formulation of Bacillus thuringiensis var. israelensis (B.t.i.) was evaluated for its potential to control chironomid midge larvae in newly sown rice crops in south-eastern Australia. Two replicated small-plot field trials were conducted using product application rates of 0.5-6 kg/ha. In trial 1 application rates between 2 and 6 kg product/ha all significantly (P<0.05) reduced populations of target Chironominae/Orthocladiinae by between 71% and 93% over the 19 day post-treatment monitoring period. Trial 2 was conducted using lower application rates (0.5-2 kg product/ha) and only the 2 kg product/ha rate significantly (P<0.05) reduced numbers of the target group (81% reduction) despite lower application rates resulting in target group suppression of 38-62%. Identification of larvae to species level from selected samples indicated that populations of Chironomus tepperi, the principal pest species that attacks the roots of rice seedlings, were reduced at all application rates; elimination of C. tepperi was achieved in trial 1 at an application rate of 2 kg/ha. Consistent with other studies, non-target Tanypodinae were not adversely affected by B.t.i., and in some treatments populations of Tanypodinae exceeded control levels by up to 73%. In the first trial, which was conducted under relatively high pest pressure, plant establishment was significantly (P<0.05) increased (120-157%) by Vectobac® WDG application rates of 2-6 kg/ha. No significant increase in plant establishment relative to the controls was identified in the second trial, when pest pressure was substantially lower and minimal damage occurred in the control bays. Overall, our results demonstrate that B.t.i. may be an economically viable alternative to broad-spectrum synthetic pesticides for the control of phytophagous midge larvae in establishing rice crops where members of the Chironominae, the group most susceptible to B.t.i., are the principal species of concern. The high specificity of B.t.i. for nematoceran Diptera should lead to reduced impacts on non-target organisms.

Ultrastructure and molecular phylogenetics of Helmichia lacustris, a microsporidium with an uncoiled isofilar polar filament.

The description of Helmichia lacustris Voronin (Parazitologiya 34:327-331 1998) is supplemented with morphogenesis and ultrastructure of the extrusion apparatus. Formation of the anterior (made up by rare short lamellae) and posterior (made up by spongy matter or small vesicles) regions of the polaroplast is preceded by granulated spheres and agglomerations of bean-like bodies, respectively. The anchoring disc is formed by an oval structure of moderate electron density, sometimes possessing a granular texture. The parasite development occurs within the cisterns of granular endoplasmatic reticulum (ER) of the host cell. Each group of spores is enclosed within a two-layered sheath, including the smooth inner membrane of the sporophorous vesicle and the outer ribosome-encrusted membrane (which originates from the host cell ER) of the parasitophorous vacuole. Two microsporidia, H. lacustris (GenBank accession number GU130406) and Euplotespora binucleata (GenBank accession number DQ675604) share 78.1% of 16S rRNA gene sequence similarity. Both parasites are characterized by an uncoiled isofilar polar filament. They form a cluster nested among terrestrial and aquatic microsporidia with well-developed coiled polar filaments, suggesting that an uncoiled polar filament in this species is a result of reduction, rather than a "primitive" character.

A new mermithid nematode in imagos of Cladopelma collator (Townes) (Diptera: Chironomidae) eclosing from waterways in northern Minnesota.

Octomyomermis connellyi n. sp. (Nematoda: Mermithidae) is described from the hemocoel of male and female imagos of Cladopelma collator (Townes) eclosing from Lake Ozawindib and from Gulsvig Landing in the upper Mississippi River in northern Minnesota. The species is distinguished from the other species in the genus. Included is a list of the other 9 adequately described members of the genus Octomyomermis that have been reported from the United States (California and Minnesota), Argentina, Russia, and Zambia. Bionomics and anomalies of the new species are addressed.

A new species of mermithid (Nematoda) in chironomids eclosing from a northern Minnesota bog.

Hydromermis haggardi n. sp. (Nematoda: Mermithidae) is described from the hemocoel of chironomid imagos eclosing from Lake Alice Bog in northern Minnesota in June and July 2007. The species is distinguished from the other 27 described members of the genus by the subventral mouth, tulip shape to anterior end of esophagus, colorless vesicular trophosome, single nonbifurcated spicule, short body lengths of both males and females, structure of the S-shaped vagina, decrease in body width at vulva, brevity of postparasitic stage, and the greater development of a posterior ventral protractor muscle in the males. An anomalous adult male bearing large oval structures in the posterior testis is reported. Evidence is presented for the developmental interaction of the new species with Lanceimermis palustris Johnson and Kleve, 2004.

A new mermithid nematode from Ablabesmyia monilis (L.) (Diptera: Chironomidae) in northern Minnesota.

Limnomermis steineri n. sp. (Nematoda: Mermithidae) is described from the hemocoel of male, female, and intersex imagos of Ablabesmyia (s. str.) monilis (L.) eclosing from Lake Ozawindib and Lake Alice Bog in northern Minnesota during summers 2000-2005. The species is distinguished from other described members of the genus. Paramermis rosea is returned to Limnomermis. A review of adequately described members of the genus is included. One species of the genus has been reported previously from the Nearctic Realm. Emphasis is placed on the need for host identification, larger number of specimens of both mermithid sexes, and information on the intensity of infections to enable more precise range comparisons in mermithid biometrics.

Hydromermis osami n. sp. (Nematoda: Mermithidae) from chironomids eclosing from northern Minnesota waterways.

Hydromermis osami n. sp. (Nematoda: Mermithidae) is described from the hemocoel of 2 species of Rheotanytarsus chironomid imagos eclosing from Lake Ozawindib and Gulsvig Landing in northern Minnesota during August 2002 and June and July 2006. The species is distinguished from the other 26 described members of the genus by the terminal mouth; oval, opaque, thick walled amphids, wider than long; acute posterior end; single spicule not bifurcated proximally; well-developed uterine and vulval limbs of the S-shaped vagina; absence of bursal sleeve; absence of nutrient vesicles in the trophosome; and an esophagus length over 40% of body length. Members of the new species emerge from the hosts as adults. The other described Hydromermis species, number of specimens, location, and known hosts are tabulated.

Annual Survey of Horsehair Worm Cysts in Northern Taiwan, with Notes on a Single Seasonal Infection Peak in Chironomid Larvae (Diptera: Chironomidae).

The life cycle of the freshwater horsehair worm typically includes a free-living phase (adult, egg, larva) and a multiple-host parasitic phase (aquatic paratenic host, terrestrial definitive host). Such a life cycle involving water and land can improve energy flow in riparian ecosystems; however, its temporal dynamics in nature have rarely been investigated. This study examined seasonal infection with cysts in larval Chironominae (Diptera: Chironomidae) in northern Taiwan. In the larval chironomids, cysts of 3 horsehair worm species were identified. The cysts of the dominant species were morphologically similar to those of Chordodes formosanus. Infection with these cysts increased suddenly and peaked 2 mo after the reproductive season of the adult horsehair worms. Although adult C. formosanus emerged several times in a year, only 1 distinct infection peak was detected in September in the chironomid larvae. Compared with the subfamily Chironominae, samples from the subfamilies Tanypodinae and Orthocladiinae were less parasitized. This indicates that the feeding behavior of the chironomid host likely affects horsehair worm cyst infections; however, bioconcentration in predatory chironomids was not detected.

Lanceimermis palustris n. sp. and Telomermis palustris n. sp. (Nematoda: Mermithidae) parasites of chironomid (Insecta: Diptera) adults from Lake Alice Bog in northern Minnesota.

In the summer of 2003, chironomid imagoes eclosing from Lake Alice Bog in Minnesota yielded mermithid nematodes of 2 new species. Lanceimermis palustris n. sp. is distinguished from the other 14 species of the genus by the subventral mouth position, thinness of the hook-shaped spicule, body index, and structure of the male tail muscles. Telomermis palustris n. sp. is distinguished from the only other species of the genus by the structure of the amphids, body length, maximum body width, length of the esophagus, and the presence of a minute larval terminal horn. Lanceimermis palustris n. sp. hosts were Chironomus maturus Johannsen and T. palustris n. sp. hosts were Tanytarsus mendax Kieffer, Paratanytarsus nr. dissimilis n. sp., and Micropsectra polita (Malloch). In both mermithid species, the percent of mermithid males per host increased with the intensity of the infection.

Strelkovimermis acuticauda n. sp. and Strelkovimermis buccalis n. sp. (Nematoda:Mermithidae) from adult chironomids (Diptera:Chironomidae) from Lake Itasca, Minnesota.

Specimens of 2 mermithid species were obtained from adults of undetermined species of chironomids emerging from Lake Itasca (Clearwater County), Minnesota. Both species have the characteristics of species in the genus Strelkovimermis. The new species are distinguishable from one another and from the 8 currently accepted species in the genus by their small size, shape of the posterior end, length of the buccal funnel, length of the cephalic papillae, location of the mouth, and morphology of the spicule region and vagina. Strelkovimrmis acuticauda n. sp. is unique in possessing an acute posterior end, an auxiliary protractor muscle, and a relatively long vagina, whereas Strelkovimermis buccalis n. sp. is distinguishable from other Strelkovimermis species by its thick cuticle and exceptionally long buccal funnel. The significance of the tail muscles is emphasized as a taxonomic feature to be used in describing future species of Strelkovimermis. A key to the 10 species of Strelkovimermis is provided.

Hydromermis contorta (Kohn) and Hydromermis pseudocontorta n. sp. from chironomids of Lake Itasca and Long Lake, Minnesota.

Hydromermis contorta (Kohn) and Hydromermis pseudocontorta n. sp. are described from chironomids in Lake Itasca and Long Lake, Minnesota, respectively. The former was recovered from adult females of Glyptotendipes paripes (Edwards) and the latter from fourth-instar larvae of Chironomus sp. Hydromermis pseudocontorta n. sp. resembles H. contorta in cephalic structures, overall size, and the presence of a restricted trophosome in the female. The terminal mouth, long uterine and vulvar limbs of the vagina, and the strongly chitinized brownish spicule of H. contorta contrast with the subventral mouth, short vaginal limbs, and the light yellow spicule of H. pseudocontorta n. sp. Both nematode species emerge from the host as sexually mature adults and both species give evidence of mating while in the host. The H. contorta described by Welch is designated as a new species, Hydromermis albionis n. sp.

Two new species of imagocidal Hydromermis (Nematoda: Mermithidae) from Endochironomus subtendens (Townes) (Diptera: Chironomidae).

Hydromermis viridis n. sp. and Hydromermis gastroviridis n. sp. are described from June and August emerging imagos, respectively, of Endochironomus subtendens (Townes) in Lake Itasca, Clearwater County, Minnesota. The position of the mouth, shape of the posterior end, structure and size of the spicule, absence of an excretory pore, arrangement of male tail musculature, and measurements separate the new species form other species of Hydromermis as well as from one another. The 2 species are placed in the genus Hydromermis though having a rounded tail and only 6 hypodermal cords at midbody. An anomalous double vagina condition is reported in H. viridis.

Survival and reproductive output of Chironomus tentans (Diptera: Chironomidae) in response to parasitism by larval Unionicola foili (Acari: Unionicolidae).

Parasitic associations between larval Unionicola foili and the dipteran Chironomus tentans were established in the laboratory, and the effects of these water mite larvae on survival and reproduction of C. tentans were determined. Longevity of parasitized insects was not significantly different from those that were unparasitized. Furthermore, there was no significant difference in the onset of oviposition between parasitized and unparasitized female chironomids. However, infected females oviposited significantly more eggs compared to uninfected controls. This study contradicts others indicating that water mite larvae severely reduce the survivorship and reproductive output of insects. The results are consistent with analyses indicating that degree of parasitism is not correlated with either duration of the parasitic phase or longevity of larvae following their parasitic association with chironomids. Larval U. foili apparently secure sufficient nutritional resources to complete their development, yet their hosts still exhibit fitness components expected for unparasitized individuals. Extreme site specificity exhibited by larval U. foili may limit the degree of parasitism on host insects and in doing so preclude larval mites from achieving infection levels that could negatively impact host survival and reproduction.

Host sex preferences and transmission success by the water mite Unionicola foili (Acari: Unionicolidae) parasitic on the midge Chironomus tentans (Diptera: Chironomidae).

This study examined whether ecoparasitic larval Unionicola foili exhibited a sex bias when infecting laboratory populations of the host insect Chironomus tentans and whether an association with male or female midges increased the likelihood of larval mites returning to the aquatic habitat. When laboratory populations of C. tentans were exposed to larval U. foili, there was a higher prevalence of mites among female hosts at emergence (17 of 30 males vs. 25 of 30 females infected by mites). However, there was no significant difference in the distribution or abundance of larvae among infected male (mean = 2.3 larvae per host) and female (mean = 2.6 larvae per host) midges. Larval mites parasitizing both male and female chironomids were more likely to return to water than could be expected by chance. Mite larvae infesting female C. tentans were more likely to return to water when female hosts deposited egg masses in water, suggesting that oviposition plays an important role in cueing larvae parasitizing female midges to detach. The mechanism responsible for increasing the likelihood that mites parasitizing male hosts return to water remains unclear. Future studies will address the possibility of parasite-mediated changes in host behavior.

Strelkovimermis amphidis n. sp. from chironomid adults emerging from Lake Itasca and Long Lake, Minnesota.

Mermithid nematodes, Strelkovimermis amphidis n. sp., emerged from chironomid imagos from Lake Itasca in Minnesota in the fall of 1996, 1997 and from Long Lake in the fall of 1998. The species is distinguished from the other 11 members of the genus by the long cephalic papillae, absence of an excretory pore, pointed termini in both sexes, large amphids, body diameter decrease at the vulva, long vagina, and the absence of lateral genital papillae. Strelkovimermis amphidis n. sp. is the fifth member of this genus recorded from Lake Itasca. The presence of and nature of the bursal sleeve is suggested as a useful distinguishing characteristic. The ratios involving spicule axis length, diameter of the body at the genital pore, and the length of the tail are also discussed in distinguishing species of Strelkovimermis. An expanded key to the species of Strelkovimermis is included.

Strelkovimermis papillosus n. sp. (Nematoda: Mermithidae), a parasite of chironomid (Insecta: Diptera) adults from the headwaters of the Mississippi River in northern Minnesota.

On 18 August 2002, chironomid imagoes of Rheotanytarsus sp. emerged from the upper Mississippi River in Minnesota and yielded distinctive mermithid nematodes of a new mermithid species. Strelkovimermis papillosus n. sp. is distinguished from the other 14 species of the genus by the presence of unusually large cephalic papillae encircling the mouth and forming a rosette with the mouth in the center and by the absence of a fixator muscle in the males. Additionally, both sexes have very acute posterior ends, long amphids, and a long stoma. Strelkovimermis is revised by eliminating nondiscriminating parameters and accommodating the 15 known species. Intrageneric characteristics useful in separating species of Strelkovimermis are listed. Intensity of infection and intensity of infection versus sex were determined from 41 hosts. Where known, the hosts and geographical distribution are given for all 15 Strelkovimermis species.

Strelkovimermis arkansensis n. sp. and Hydromermis welchi n. sp. from chironomid imagos.

Strelkovimermis arkansensis n. sp. and Hydromermis welchi n. sp. are described from chironomid adults in Arkansas and Minnesota, respectively. The host of the former is Glyptotendipes paripes (Edwards) and of the latter Polypedilum (Polypedilum) nubeculosum (Meigen). Strelkovimernis arkansensis n. sp. most closely resembles Strelkovimermis buccalis Johnson and Kleve, 1996 and is separated from that species by color in the living worms, thickness of the cuticle, and length of the cephalic papillae. Hydromermis welchi n. sp. is similar to Hydromermis gastroviridis Johnson and Kleve, 1997 and is separated from that species by color in the living worms, structure of the spicule, and number of genital papillae. A variant of H. welchi n. sp. is described. Molting in S. arkansensis was apparently facilitated by having both postparasitic males and females in the same vial.

[Changes in the monocercus cercomer in the cysticercoid cavity and the hemocoel of the nonspecific host]. / Izmeneniia tserkomera monotserkov v polosti tsistitserkoida i gemotsele nespetsificheskogo khoziana.

In order to elucidate the functional role of cercomer in larvae of the monocercus type their transplantation from the specific host Chironomus obtusidens to Gammarus lacustris was conducted. At early stages after the transplantation proceeds an increase in the functional activity of the tegument of follicles of the cercomer followed by their complete destruction in 3 or 4 days. On the surface of the exocyst membrane an adhesion of the host's haemocytes occurs, which becomes more distinct in 3-4 days when the process acquires a character of local encapsulation. Within the same period, in the places of haemocytes aggregation, a local resorption of the exocyst external membrane takes place. Later intensification of the host response to transplant is associated with the destruction of follicles of the cercomer. In one case the occurrence of follicles of the cercomer in the cavity of cysticercoid was observed that is caused by the microbe affection of the latter. In the zone of contact of the tegument of scolex and neck with follicles of the cercomer an increased secretion (the microapocrine type) of the tegument, disturbance of the microvillous tegument of the cercomer's follicles and their destruction are observed. Incompatibility of the tegument of definitive departments and cercomer, which arises during differentiation of larvae, is supposed to affect the formation of scolex invagination in the evolution of larvae of Hymenolepis.

A new mermithid species (Nematoda) from chironomid imagos eclosing from Lake Ozawindib in northern Minnesota.

Hydromermis biesboeri n. sp. (Nematoda: Mermithidae) is described from the hemocoel of imagos of chironomid host species eclosing from Lake Ozawindib in northern Minnesota in July and August 2008 and 2009. The new species is distinguished from the other 28 described species in the genus by terminal mouth, wide oval (male) and long oval (female) amphids with a dorsal commissure, tapered and rounded posterior ends, split base of the spicule, short protractor muscles in males, and by females having an S-shaped vagina with equal-length terminal limbs. The new species represents the 10th Hydromermis mermithid species described, or reported, from the region and the second from Lake Ozawindib.