Systematics and biogeography of the Gondwanan Orthocladiinae (Diptera: Chironomidae).

Restrictions to effective dispersal and gene flow caused by the fragmentation of ancient supercontinents are considered to have driven diversification and speciation on disjunct landmasses globally. Investigating the role that these processes have played in the development of diversity within and among taxa is crucial to understanding the origins and evolution of regional biotas. Within the chironomid (non-biting midge) subfamily Orthocladiinae (Diptera: Chironomidae), a group of genera that are distributed across the austral continents (Australia, New Zealand, South America) have been proposed to represent a relict Gondwanan clade. We used a molecular approach to resolve relationships among taxa with the aim to determine the relative roles that vicariance and dispersal may have played in the evolution of this group. Continental biotas did not form monophyletic groups, in accordance with expectations given existing morphological evidence. Patterns of phylogenetic relationships among taxa did not accord with expected patterns based on the geological sequence of break-up of the Gondwanan supercontinent. Likewise, divergence time estimates, particularly for New Zealand taxa, largely post-dated continental fragmentation and implied instead that several transoceanic dispersal events may have occurred post-vicariance. Passive dispersal of gravid female chironomid adults is the most likely mechanism for transoceanic movement, potentially facilitated by West Wind Drift or anti-cyclone fronts. Estimated timings of divergence among Australian and South American Botryocladius, on the other hand, were congruent with the proposed ages of separation of the two continents from Antarctica. Taken together, these data suggest that a complex relationship between both vicariance and dispersal may explain the evolution of this group. The sampling regime we implemented here was the most intensive yet performed for austral members of the Orthocladiinae and unsurprisingly revealed several novel taxa that will require formal description.

Chironomid midges as a cause of allergy in the Sudan.

Hypersensitivity to Chironomidae (non-biting midges) has been a problem in the Sudan since about 1927 and appears to be due to increased breeding of a single chironomid species, Cladotanytarsus lewisi (Freeman). Mass emergence of the midges in thought to be related to the larval diet of algae and diatoms, the numbers of which are greatly enhanced by the retention of plant nutrients in lacustrine conditions resulting from interruption to the natural flow of the Nile by the construction of dams. Immunological studies in allergic individuals using an allergen extract prepared from C. lewisi indicate that the concentration of specific immunoglobulin E ("allergic antibody") directed against C. lewisi is raised in patients with established hypersensitivity to the midge but not in control subjects. The concentration of specific IgE is also related to the severity of clinical symptoms. These results indicate that this widespread and important "man made" hypersensitivity in the Sudan has the features of well recognized immediate-type allergy commonly associated with pollens and other air-borne allergens.

New World Stictocladius Edwards (Diptera: Chironomidae).

The orthocladiine Chironomidae genus Stictocladius Edwards was described originally from South America. Although recognised subsequently as present also in Australia and New Zealand, the true diversity in the Neotropics has remained unclear. After more than a decade of collections of both isolated adults and aquatic immature stages, we can recognise several new taxa and associate some immature stages. Thus, we describe Stictocladius prati n. sp. as male, female, pupa and larva; Stictocladius acutus n. sp. and Stictocladius acrilobus n. sp. as male, female and pupa; Stictocladius fimbriatus n. sp. as male and female; Stictocladius fovigus n. sp. and Stictocladius nudiventer n. sp. as male and pupa; and Stictocladius privicalcar n. sp. and Stictocladius prostatus n. sp. each as male imago alone. The male and female of Stictocladius pulchripennis Edwards is redescribed and the pupa described. The male and female of Stictocladius flavozonatus Edwards and the male of Stictocladius calonotum Edwards are described. Five pupal types are described: Stictocladius sp. A (near S. acrilobus), Stictocladius sp. B (possibly S. calonotum), Stictocladius sp. C (near S. calonotum), Stictocladius sp. D (possibly S. flavozonatus) and Stictocladius sp. E with uncertain affinity. A larva from Chile of the Stictocladius 'sofour type' (Stictocladius sp. F) and an unreared larva from North America (Stictocladius sp. G) possibly belonging to S. acutus are described. Keys to named Neotropical male and female imagines of Stictocladius and to all pupal forms of Neotropical Stictocladius are provided. Some data concerning fourth instars of Stictocladius are presented. Means of differentiation from putative sister taxon Lopescladius are discussed.

Biomonitoring and invertebrate taxonomy.

Biological monitoring requires identification of the fauna of the environment under study. Three prevalent methods have been used to alleviate the need for universal species-level identification: selection of indicator taxa, taxonomic reduction, and allocation to functional groups. Each method is discussed in relation to species recognition. It is suggested that the validity of all attempts to reduce data should be tested in the light of phylogenetic hypotheses.

Allergens of non-biting midges (Diptera: Chironomidae): a systematic survey of chironomid haemoglobins.

Various genera of non-biting midges (Diptera: Chironomidae) possess haemoglobins in larvae and adults. For certain species, these haemoglobins have been implicated in human allergic disease in several countries. The present study confirms and extends observations that haemoglobin is present in many species of Chironomidae, establishes that it is retained from the larval to the adult stage and shows that haemoglobin persists both in live and in dead dry flies. Previous suggestions that Chironomidae should be seen as important environmental and occupational allergens are clearly substantiated.

Further characterisation of allergens associated with hypersensitivity to the "green nimitti" midge (Cladotanytarsus lewisi, Diptera: Chironomidae).

Chironomid midges are small (2-15 mm) non-biting flies, characteristically seen swarming by water at dusk. Allergens of the "green nimitti" midge, Cladotanytarsus lewisi (Freeman) (Diptera: Chironomidae), a cause of widespread hypersensitivity in the Sudan, were isolated and partially characterized by Sephacryl S200 chromatography. The allergenicity of the fractions was identified by "rocket" autoradiography, RAST inhibition, skin "prick" tests, and the immunoblot technique. The fractions were further analysed by isoelectric focusing and SDS-PAGE. Two major allergens with pI's ranging from 4.3 to 6.0 were identified and had molecular weights of approximately 17,000 and 32,000 daltons, sizes compatible with their being monomeric and dimeric haemoglobins. Since chironomids occur in nuisance numbers worldwide and their haemoglobins have been shown to produce severe hypersensitivity reactions in man, they should be seen as an important potential cause of environmental and occupational allergy.

Immediate-type skin reactivity to extracts of the 'green nimitti' midge, (Cladotanytarsus lewisi), and other chironomids in asthmatic subjects in the Sudan and Egypt.

Skin 'prick' tests with an extract of the 'green nimitti' midge, Cladotanytarsus lewisi (Freeman), a potent airborne allergen associated with asthma and rhinitis in the Khartoum area, were undertaken on asthmatic subjects in several localities in the Sudan and Egypt in order to provide further information on the extent of this allergic problem. In the Sudan skin-test-positive individuals were identified in the Khartoum area, Kosti, Sennar, Wad Medani, Shendi and Atbara, and in Egypt at Aswan, Luxor and Qena. A number of asthmatic individuals giving a positive reaction to C. lewisi were also tested with extracts of seven sub-dominant species of Nilotic Chironomidae. There appeared to be a limited degree of cross-reactivity with these species, particularly Dicrotendipes fusconotatus (Kieffer), Procladius noctivagus (Kieffer) and Conchapelopia cygnus (Kieffer). The numbers of C. lewisi sensitive subjects giving positive skin-tests to these other species were ten out of 20, 12 out of 28 and 12 out of 29 respectively. In contrast, only five out of 31 reacted to Paracladopelma graminicolor (Kieffer), five out of 24 to Cladotanytarsus pseudomancus (Goetghebuer), three out of 32 to Nanocladius vitellinus (Kieffer) and two out of 28 to Cryptochironomus neonilicola (Freeman). These results indicate that Cladotanytarsus lewisi is a major source of chironomid allergen(s) in these asthmatic subjects and that hypersensitivity to C. lewisi is probably far more widespread than originally envisaged.

Partial characterization of allergens associated with hypersensitivity to the 'green nimitti' midge (Cladotanytarsus lewisi, Diptera: Chironomidae).

Allergens in extracts of the 'green nimitti' midge, Cladotanytarsus lewisi Freeman (Diptera: Chironomidae), a cause of widespread hypersensitivity in the Sudan, were isolated and partially characterized by assays which depend on the binding of 125I-anti-IgE to allergen-IgE complexes. These methods included RAST inhibition, crossed radioimmunoelectrophoresis (CRIE) and rocket radioimmunoelectrophoresis (RRIE). Following Sephadex G100 chromatography the 'major peak' of allergenicity as determined by RAST inhibition, RRIE and SDS-PAGE was associated with molecules of approximately 17 000 daltons. The peak eluting at Vo contained material of molecular weight 66 000 daltons which also bound 125I-anti-IgE, but had only 61% of the activity of the 'major peak' by RAST inhibition. By isoelectric focusing and RRIE of fractions obtained by chromatofocusing with polybuffer exchanger 94, the 'major peak' was associated with multiple bands with a pI range of 3.5-5.5. These results indicate that the major allergens from C. lewisi are a group of closely related acidic peptides.

Evidence of allergic hypersensitivity to chironomid midges in an English village community.

Some chironomid (non-biting) midges contain potent human allergens, principally in the form of insect haemoglobins. In view of previously demonstrated immunological cross-reactivity between some different midge species, the possible role of British midges in allergic disease was studied. The chosen site, Farmoor near Oxford, is a village that has been subject to several annual nuisance midge swarms since the construction of an adjacent water-supply reservoir in 1964, which was extended in 1975. The incidence and severity of hypersensitivity in the community was assessed by questionnaire survey, skin-prick testing, radio-allergosorbent tests (RAST) and RAST inhibition, and enzyme-linked immunosorbent assays (ELISA) and ELISA inhibition, using extracts of locally caught adult midges (Tanytarsus sylvaticus, T. bathophilus and Chironomus anthracinus) and laboratory cultured larvae (C. riparius). These tests revealed a low but significant level of midge-related hypersensitivity with skin-prick results showing seasonal fluctuations, but this seasonality was not confirmed with an IgE RAST. A correlation between IgE levels and relevant symptoms was found (P less than 0.01) using RAST, but not between IgE and skin-test results. Enzyme-linked immunosorbent assay results did not correlate with any other parameter measured. The incidence and severity of allergic reactions to the midges tested in this community was low.

Evidence for haemoglobins as common allergenic determinants in IgE-mediated hypersensitivity to chironomids (non-biting midges).

Chironomids (non-biting midges) are known to cause IgE-mediated hypersensitivity in man. This study compares the cross-reactivity between the chironomid midge Cladotanytarsus lewisi ('green nimitti'), a widespread cause of allergy in the Sudan and Chironomus riparius (= thummi, CTT) where larvae are used as pet fish food and where haemoglobins were previously shown to be major allergens. As with C. riparius, immature forms of C. lewisi also contain allergenic material since skin test responses to larval, pupal and adult extracts were obtained in Sudanese individuals. Crossed radioimmunoelectrophoresis of the C. lewisi larval and pupal extracts indicate that they contain a higher proportion of the allergenic fractions than adults. Further evidence of common allergen determinants between C. lewisi and C. riparius were obtained by the demonstration of positive skin-prick tests, in Sudanese patients, to extracts of larval, adult and isolated haemoglobin extracts of C. riparius. Cross-reactivity between C. lewisi and C. riparius was also demonstrable by RAST inhibition studies. A dose-dependent inhibition was observed using both the C. lewisi adult midge RAST and the C. riparius haemoglobin RAST, the two respective antigens, and sera from individuals hypersensitive to either C. lewisi or C. riparius. Due to the immunological cross-reactivity found between these distantly related species, we conclude that chironomids should be seen as significant environmental and occupational allergens.

Persistence of hemoglobin allergenicity and antigenicity during metamorphosis of Chironomidae (Insecta: Diptera).

Chironomid hemoglobins are potent allergens. The allergenic and antigenic activities of these hemoglobins are studied with the help of RAST, RAST inhibition and double immunodiffusion. Human as well as rabbit antisera were used. It was shown that hemoglobins are the main antigenic/allergenic components in extracts of Camptochironomus tentans larvae. Furthermore, immunological cross-reactivity among larvae, pupae and adult midges of this species are shown to be due to the existence of hemoglobin antigenic determinants in all developmental stages of this insect.