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.

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.

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 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.

A new genus and species of mermithid (Nematoda) emerging from chironomid imagos eclosing from the Mississippi River headwaters area in northern Minnesota.

Fibromermis roarki n. gen., n. sp. (Nematoda: Mermithidae) is described from the hemocoel of chironomid imagos of Ablabesmyia monilis (L.) eclosing from the headwaters of the Mississippi River at Gulsvig Landing in northern Minnesota in the summer of 2007. The new genus is described, distinguished from the most closely related tetrapapillate aquatic mermithid genus, and the new species described. Intensity and sex specificity of infection are recorded from 28 host adult chironomids of a single cohort. The new species represents the fourth tetrapapillate mermithid species described from the area and the third tetrapapillate genus.

Primeiro registro de larvas de Chironomus inquinatus Correia, Trivinho-Strixino & Michailova (Diptera, Chironomidae) vivendo no casco do cágado Phrynops geoffroanus Schweigger (Testudines, Chelidae) na região Neotropical / First record of Chironomus inquinatus larvae Correia, Trivinho-Strixino & Michailova (Diptera, Chironomidae) living on the shell of the side-necked turtle Phrynops geoffroanus Schweigger (Testudines, Chelidae)

Neste estudo reportamos pela primeira vez a ocorrência de larvas de Chironomus inquinatus Correia, Trivinho-Strixino & Michailova vivendo no casco do cágado Phrynops geoffroanus Schweigger, possivelmente em função da intensa acumulação de sedimento, em um rio poluído da região Neotropical.

In this study we report for the first time the occurrence of Chironomus inquinatus larvae Correia, Trivinho-Strixino & Michailova living on the shell of the side-necked turtle, Phrynops geoffroanus Schweigger, possibly by the intense accumulation of sediment, in a polluted river of the Neotropics.

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.

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.

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.

Proteocephalidean larvae (Cestoda) in naturally infected cyclopid copepods of the upper Paran River floodplain, Brazil.

The occurrence, prevalence and infection intensity of proteocephalidean larvae in naturally infected intermediate hosts of the Upper Paran River floodplain are reported. A total of 5,206 zooplanktonic and benthic organisms were analyzed, namely cyclopid (2,621) and calanoid (1,479) copepods, cladocerans (704), rotifers (307), chironomid larvae (41) and ostracods (54). Eight cyclopid copepods - two copepodids, one male and five females - comprising 0.3% of the cyclopid copepods examined, were naturally infected. The male infected belonged to a species of Paracyclops, and the females to Paracyclops sp., Thermocyclops minutus and Mesocyclops longisetus.

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.

Chromosomal polymorphism is associated with nematode parasitism in a natural population of a tropical midge.

A natural population of a tropical midge, Chironomus ramosus (Diptera: Chironomidae), was found to be polymorphic for a paracentric inversion (IV: 18C-19D). Based on the characteristic banding pattern of the fourth chromosome in the larval salivary gland polytene nuclei, individuals were classified as either structural homozygotes or heterozygotes. Isofemale lines were obtained and subsequently standard (S/S) and inversion (I/I) homozygotes were characterised by careful progeny testing in the laboratory. While exploring various biotic and abiotic factors that might be responsible for the maintenance of inversion polymorphism, we detected nematode (Family: Mermithidae) infections among the larval population. A detailed study indicated that the inversion polymorphism in the natural population of C. ramosus was apparently being maintained as a result of the selective pressure exerted by the nematode parasite. The corresponding pattern of increase and decrease in genotype frequencies and the relative fitness values indicated a selective advantage of inversion heterozygotes (S/I) over both homozygous types (S/S and I/I). Both empirical and experimental data suggest the strong heterotic nature of adaptation in this C. ramosus population towards nematode infection. This is the first report of its kind where inversion polymorphism has been shown to be associated with nematode parasitism.

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.

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.

The behavioral responses by larvae of the water mite Unionicola foili (Acari: Unionicolidae) to gravity, light and host chemicals: adaptations for transmission success?

The life cycle of the water mite Unionicola foili includes larvae that parasitize chironomid dipterans, and nymphs and adults that occur in association with the freshwater mussel Utterbackia imbecillis. The georesponses and photoresponses of larval U. foili in the presence of chemical substances from chironomids and mussels both prior to (= pre-chironomid larvae) and after their parasitism with chironomids (= post-chironomid larvae) were examined in the laboratory. The responses of larval U. foili to light and gravity changed during larval ontogeny and varied depending on the chemistry of the medium in which they were examined. Pre-chironomid larvae preferentially responded to water modified by chironomids given that negative phototaxis and positive georesponses were elicited only in the presence of water that had been modified by these hosts. Post-chironomid larvae exhibited host-induced negative phototaxis only in the presence of mussel-modified water. Post-chironomid larvae were consistently geopositive when exposed to water modified by mussels or chironomids. The pattern of responses exhibited by larval U. foili to light and gravity are interpreted as adaptations for locating potential hosts in the field.