Novel spirochetes isolated from mosquitoes and black flies in the Czech Republic

During the years 1999–2002, a total of 4,898 individuals of 26 species of hematophagous insects (4,149 mosquitoes, 583 black flies, and 166 tabanid flies) was examined for the presence of spirochetes using dark‐field microscopy. There was an overall recovery of spirochetes from the midguts of Culicidae and Simuliidae of 23.5% and 11.4%, respectively. Spirochetes were not detected in Tabanidae. Seven spirochetal strains have been successfully recovered from mosquitoes and black flies: BR149 (Culex pipiens), BR151 (Cx. pipiens), BR173 (Cx. pipiens), BR177 (Cx. pipiens), BR193 (Aedes cinereus), BR208 (Cx. pipiens), and BR231 (Simulium noelleri). The strains have been adapted to laboratory conditions (BSK‐H Complete medium). Their preliminary determination based on 16S rRNA gene sequencing has shown that they differ from the Lyme disease spirochete Borrelia burgdorferi sensu lato as well as other members of the Order Spirochaetales indicating novel bacterial species in the Family Spirochaetaceae.     

Some boundary layer characteristics of microhabitats occupied by larval black flies (Diptera: Simuliidae)

Microhabitats occupied by larvae of 12 species or complexes of black flies at 30 sites in western Canada were investigated for fluid flow parameters. An attached boundary layer was a feature of almost all microhabitats occupied by larvae of black flies. Although not measured directly, acceleration of water resulting from flow around an object appeared to alter flow characteristics and make microhabitats more favourable. This predictability in the flow characteristics of microhabitats occupied by larvae of black flies implies that the contagious dispersion of these larvae is a response to the heterogeneity of flow over a stream bed. It is suggested that microhabitats characterized by suitable flow characteristics may constitute a resource that limits population size. Larvae occupied microhabitats with both laminar and turbulent boundary layers, and were found in microhabitats with a wide range of water velocities. Mainstream flow parameters investigated were; water velocity, water depth, and whether flow was subcritical, critical or supercritical. Boundary layer parameters investigated were; whether flow was laminar or turbulent, and the point at which separation from the substratum occurred.     

Factors affecting the efficacy of three commercial formulations of Bacillus thuringiensis var. israelensis against species of European black flies

The efficacy of three commercial formulations of Bacillus thuringiensis var. israelensis against two European black fly species was determined. The effect of seston concentration, temperature and larval size on the efficacy of the formulations was also determined. Teknar‐WDC was found to be almost four times as effective as Bactimos‐WP though only slightly more efficient than Vectobac‐AS against Simulium variegatum gp. larvae. Small larvae ( < 2.5 mm) were more susceptible to both Teknar‐WDC and Vectobac‐AS than large larvae ( > 5 mm) though no difference was seen with Bactimos‐WP. High seston concentrations both before and during bioassays significantly decreased the efficacy of all formulations. Increased temperatures increased the efficacy of all formulations.     

Flow patterns around cocoons and pupae of black flies of the genusSimulium (Diptera : Simuliidae)

Flow patterns around structurally different cocoons and pupae of five species ofSimulium Latreille are described. Three features of the flow pattern common to all cocoons are; 1) a solenoidal vortex around the cocoon, 2) upward flow anterior (downstream) to the cocoon, and 3) one or two pairs of spiral-shaped vortices, which either touch or envelop the fill filaments of the pupa. The solenoidal vortex and the upward-spiralling, downstream vortices are common features of flow patterns around most bluff bodies submerged in a boundary layer. The proximity of the vortices to the fill filaments of all pupae suggests that these vortices are associated with gaseous exchange at the gill filaments.     

Food acquisition responses of the suspension-feeding bivalve Placopecten magellanicus to the flocculation and settlement of a phytoplankton bloom

In situ technologies were employed to monitor suspended particle flocculation and floc settlement and utilization by a cohort of sea scallops (Placopecten magellanicus) during the 2000 spring phytoplankton bloom in Bedford Basin, Nova Scotia, Canada. The objectives were to determine the effect of bloom flocculation and settling on food acquisition and utilization by scallops, and to assess the potential role of flocculation in enhancing the bioavailability of trophic resources and particle-reactive contaminants to bivalve filter feeders. The development and flocculation of the phytoplankton bloom were monitored within the surface layer (10 m depth) by in vivo chlorophyll fluorescence and silhouette camera observations. Sedimentation rate, seston abundance and composition, and sea scallop functional responses were monitored at 20 m depth (below the bloom) to provide insight into the potential forcing of feeding and digestion processes by changes in the abundance, composition and properties of the ambient food supply. The bloom began in mid-March and median floc diameter at 10 m depth increased rapidly from 200 μm to greater than 400 μm between 21 and 28 March. Flocs were observed to be abundant in the surface layer up to 4 April. Daily vertical particle flux was high during the last week of March and declined to near zero by 1 April. Clearance rates of scallops held at 20 m depth were relatively high (average ± S.D.; 11.7 ± 4.0 L h^− 1) during the period of bloom settlement and declined rapidly to low levels (0.4 ± 0.9 L h^− 1) after 31 March. Average absorption efficiency also declined (0.88 ± 0.01 to 0.78 ± 0.05) after bloom settlement. Daily biodeposition rates by scallops were poorly correlated with temporal variations in the quantity (total particulate matter and chlorophyll a concentration) or quality (organic content) of seston available to the scallops, but were significantly correlated with sedimentation rate. Comparison of disaggregated inorganic particle size distributions for suspended particulate matter, settled particles, and scallop feces indicated that fine-grained particles (1 to 4 μm) were effectively ingested by sea scallops—an indication of whole floc ingestion. The settlement of flocs produced during the spring bloom appears to be important in regulating this species physiological energetics and for enhancing the bioavailablility of fine particles (including picoplankton) and particle-reactive contaminants.     

Molecular phylogeny of black flies (Diptera: Simuliidae) from Thailand, using ITS2 rDNA

The sequences of the second internal transcribed spacer (ITS2) of ribosomal DNA (rDNA) were determined for 40 black fly species from Thailand, belonging to 4 subgenera of the genus Simulium, namely Gomphostilbia (12 species), Nevermannia (5 species), Montisimulium (1 species), Simulium sensu stricto (21 species), and an unknown subgenus with one species (Simulium baimaii). The length of the ITS2 ranged from 247 to 308 bp. All black fly species had high AT content, ranging from 71 to 83.8%. Intraindividual variation (clonal variation) occurred in 13 species, ranging from 0.3 to 1.1%. Large intrapopulation and interpopulation heterogeneities exist in S. feuerboni from the same and different locations in Doi Inthanon National Park, northern Thailand. Phylogenetic relationships among 40 black fly species were examined using PAUP (version 4.0b10) and MrBAYS (version 3.0B4). The topology of the trees revealed two major monophyletic clades. The subgenus Simulium and Simulium baimaii were placed in the first monophyletic clade, whereas the subgenera Nevermannia + Montisimulium were placed as the sister group to the subgenus Gomphostilbia in the second monophyletic clade. Our results suggest that S. baimaii belongs to the malyschevi-group or variegatum-group in the subgenus Simulium. The molecular phylogeny generally agrees with existing morphology-based phylogenies.     

Multiple symbiotes in larval black flies (Diptera: Simuliidae): associations of trichomycete fungi with ichthyosporeans, mermithid nematodes, and microsporidia

Studies of the interactions between two or more species of symbiotic microorganisms in a single host animal have lagged behind those involving interactions between hymenopteran or dipteran parasitoids. We screened the larvae of four species of black flies (Cnephia ornithophilia, Simulium furculatum, Simulium innoxium, and Simulium vandalicum) to determine if the colonization and prevalence of trichomycete fungi were influenced by ichthyosporeans, mermithid nematodes, microsporidia, or other species of trichomycetes. Differences in the layering of the peritrophic matrix of microsporidian-infected larvae, compared with that of uninfected larvae, were associated with greater levels of colonization by the midgut-inhabiting trichomycete Harpella melusinae. Colonization levels of different species of trichomycetes were significantly correlated with one another in only 2 of 11 cases. Aside from the consistent attachment of the trichomycete Simuliomyces microsporus to the ichthyosporean Paramoebidium chattoni, we found little evidence that prevalence and colonization levels of trichomycetes were influenced by other symbiotes, including other trichomycetes. This trend might reflect the physical separation of trichomycetes, which are gut specific, from other symbiotes. In addition, any combinations of symbiotes that were wholly incompatible would not have been observed in our field-collected material.     

Feeding behavior of larval black flies (Diptera: Simuliidae) with and without exposure to Bacillus thuringiensis var. israelensis.

Behavioral time budgets were constructed for larval black flies of the Simulium jenningsi and S. tuberosum groups in a recirculating flume. Larvae spent the majority (54-80%) of time filter feeding. Flick rates of the labral fans, which averaged 0.7-1.9 flicks/sec, did not differ significantly between regimens of 0 mg/l and 10 mg/l of food. Larvae consistently flicked the fan nearest the water surface significantly more often than the lower fan. They also tended to flick the right fan more than the left, although this trend was significant in only one of four treatments. Although handedness cannot be dismissed, the differential flick rate appeared to be related to the position of the fans relative to the water surface, possibly because of different particulate concentrations in the microcurrents received by the upper and lower fans. Cessation of feeding by larvae of the S. jenningsi group from rivers regularly treated with Bacillus thuringiensis var. israelensis was independent of exposure to this biological control agent. Of larvae that ceased to feed, the mean time to cessation of feeding after exposure to Bti was 12-18 s, sufficient time to ingest a lethal dose.     

Digestive system of larval black flies (Diptera: Simuliidae): structure and phylogeny

Abstract. Larval gut characters of 45 species of black flies, representing 17 subgenera and 15 genera, were studied for their utility in taxon diagnosis and phylogenetic inference. Phase-contrast and scanning electron microscopy revealed a wealth of new characters in the esophageal armature of the foregut. Larval gut morphology, particularly characters of the esophageal armature, varied significantly among taxa, permitting generic diagnoses. A cladogram inferred from 13 gut characters supported a sister-group relationship between Parasimulium and the remaining simuliids, but did not show a strong phylogenetic signal for the current, well-supported concept of tribal level monophyly.     

Divergence in larval responses to food plants between temperate and tropical populations of the black swallowtail butterfly

ABSTRACT.

• 1 Larvae of the black swallowtail butterfly, Papilio polyxenes Fabr., from New York and Costa Rica were reared under the same environmental conditions on Daucus carota L. and Spananthe paniculata Jacq. (Umbelliferae), which are the host plants most commonly used by the two populations, respectively. Feeding rates, growth rates and feeding efficiencies of the fifth instar larvae were measured by standard techniques.
• 2 Larvae from each population grew faster on the host-plant species from their region of origin than did larvae from the other population, though differences on S.paniculata were not significant. On D.carota the faster growth by New York larvae was attributable to greater efficiency. On S.paniculata, by contrast, the somewhat faster growth by Costa Rican larvae seemed to have resulted chiefly from faster feeding. The two populations have thus come to differ in metabolic and, probably, behavioural responses to host plants.
• 3 S.paniculata was a superior host plant for larvae of both populations, even though the total energy, nitrogen and water content of its foliage was similar to that of D.carota.
• 4 These results are consistent with the hypothesis that insects in the two populations have diverged in adaptation to their host plants following curtailment of gene flow between them as the species colonized Central America from the north.

     

Isolation and characterization of 11 microsatellite loci from the black fly, Simulium negativum (Diptera: Simuliidae)

Eleven polymorphic microsatellite loci were isolated and developed from the black fly, Simulium negativum, a member of the Simulium arcticum sibling species complex. The observed heterozygosity of the 11 loci ranged from 0.03 to 0.83. The number of alleles per locus ranged from 8 to 19. Significant linkage disequilibria were encountered only for the primer pairs BF7-1 with BF7-5 and BF6-32 with BF7-16. Presumably, these microsatellite loci can be used to study genetic structure within the entire S. arcticum complex.     

Search strategies of fruit flies in steady and shifting winds in the absence of food odours

Abstract. Flight behaviour by females of two species of fruit flies, Drosophila funebris and Drosophila immigrans , was videorecorded in a wind tunnel in still air and in wind with a constant or shifting direction. Flies which were deprived of food overnight took flight in the absence of food odours. Both species responded to the presented winds in agreement with two models that predict the shortest distance to an odour plume. According to these models, the shortest distance to an odour plume is travelled when insects fly at right angles to the wind with a steady direction. In winds shifting over more than 60, the shortest distance to an odour plume is achieved when insects fly parallel to the time-averaged wind direction. We propose a behavioural mechanism which accomplishes the observed flight directions taken by the two species of fruit flies in response to the tested wind regimes.     

Economy of food utilization in a gregarious sawfly,Arge nigrinodosa Motschulsky (Hymenoptera: Argidae): Relative abundance of food in the natural habitat and behavioral pattern of larval feeding

Summary Relative abundance of natural food resource, leaves of wild rose, ofArge migrinodosa was evaluated by comparing the food quantity which the larvae composing a colony utilized throughout the larval stage with that which is available in the habitat. In this connection, behavioral pattern of larval feeding was also observed under natural conditions, and its ecological significance was discussed. It was quantitatively clarified that the larvae faced the severe shortage of food and had to disperse to other shoots with the expense of probably high risk. Under pressures of such insufficient supply of food, the larvae follow the most efficient pattern of feeding behavior that they move to the tip of shoot at first and feed regularly downward thereafter. Though individual larvae behaved completely independently of each other, the aggregation appeared to be formed and maintained because all the larvae from the same egg mass follow this fixed feeding pattern.     

Effects of Phaseolus vulgaris (Fabaceae) seed coat on the embryonic and larval development of the cowpea weevil Callosobruchus maculatus (Coleoptera: Bruchidae)

Bruchid beetles infest various seeds. The seed coat is the first protective barrier against bruchid infestation. Although non-host seed coats often impair the oviposition, eclosion and survival of the bruchid Callosobruchus maculatus larvae, morphological and biochemical aspects of this phenomenon remain unclear. Here we show that Phaseolus vulgaris (non-host) seed coat reduced C. maculatus female oviposition about 48%, increased 83% the seed penetration time, reduced larval mass and survival about 62 % and 40 % respectively. Interestingly, we found no visible effect on the major events of insect embryogenesis, namely the formation of the cellular blastoderm, germ band extension/retraction, embryo segmentation, appendage formation and dorsal closure. Larvae fed on P. vulgaris seed coat have greater FITC fluorescence signal in the midgut than in the feces, as opposed to what is observed in control larvae fed on Vigna unguiculata. Cysteine protease, α-amylase and α-glucosidase activities were reduced in larvae fed on P. vulgaris natural seed coat. Taken together, our results suggest that although P. vulgaris seed coat does not interfere with C. maculatus embryonic development, food digestion was clearly compromised, impacting larval fitness (e.g. body mass and survivability).     

Feeding behavior ofRhagoletis pomonella flies (Diptera: Tephritidae): Effect of initial food quantity and quality on food foraging,handling costs,and bubbling

Under seminatural conditions feeding and postfeeding behaviors of individual apple maggot flies, Rhagoletis pomonella(Diptera: Tephritidae), were recorded after flies were presented with yeast hydrolysate or sucrose droplets, varying in either concentration, amount of food solute, or total droplet volume. The objectives were (a) to establish, at a constant level of previous food deprivation, food ingestion thresholds in relation to food quality and quantity and (b) to study the effect of initial food quantity and quality on food handling time and subsequent food foraging behavior. For both carbohydrate and protein substrates, fly foraging time after feeding on a tree branchlet was positively related to total amount of food solute previously encountered on a leaf surface, though largely independent of food volume or concentration. The volume and concentration of food presented, however, significantly affected food handling and processing time and therefore foraging time. In fact, total branchlet residence time was more closely linked to food handling and processing time than to foraging time. Less time was needed for uptake of liquid than dry food, the latter requiring liquification by salivary secretion and eliciting considerable intermittent cleaning of mouthparts by feeding flies. Similar to the situation in other fluid feeders, uptake time in R. pomonelladecreased with increasing dilution, although below a threshold of a 30% concentration of solute, the rate of nutrient intake decreased rapidly. When the level of dilution and total volume of food ingested were great enough, engorged flies entered extended quiescent postfeeding periods during which they extrude orally droplets of liquid crop contents (bubbling). After this they reinitiated feeding, followed by more bubbling and feeding bouts. Multivariate logistic regression analysis suggested that bubbling behavior is determined by liquid food volume and degree of dilution, hunger, and temperature. Although thresholds triggering bubbling decreased with increasing temperature, higher temperature by itself did not result in bubbling behavior. This suggests that bubbling is not primarily a mechanism to achieve evaporative cooling as has been suggested but, rather, a behavior to eliminate excess water, thereby enabling engorged flies to continue feeding on diluted food sources.     

The contribution of the facultative feeding period to echinoid larval development and size at metamorphosis: a comparative approach

Planktotrophic invertebrate larvae have two dissociable stages during development, a facultative feeding period, whose length is determined by the amount of maternal provisioning in the egg, and an obligate feeding period, whose length is determined by the quantity of exogenous energy needed to reach metamorphic competence. Here we set out to experimentally test the impact of feeding during the facultative feeding period at two food concentrations (limiting and nonlimiting) on larval development time and juvenile quality. We used two closely related subtropical sand dollar species that differ in the quantity of maternal investment for these comparisons: Leodia sexiesperforata (large egg, long facultative feeding period) and Mellita tenuis (small egg, short facultative feeding period). We found that feeding during the facultative period accelerates development to metamorphosis only in M. tenuis and only at the high food ration. Feeding during the facultative feeding period had no effect on development time for M. tenuis at a food limiting concentration and for L. sexiesperforata at either food concentration. Furthermore, we found feeding during the facultative period to significantly increase quantity of carbohydrates and lipids at metamorphosis only for M. tenuis in nonlimiting food concentration. Thus, our data reveal a two-fold benefit of the facultative feeding period for a poorly provisioned species under high food conditions but little effect on a well-provisioned species. We discuss our results in reference to McEdward's [McEdward, L.R., 1997. Reproductive strategies of marine benthic invertebrates revisited: facultative feeding by planktotrophic larvae. Am. Nat. 150, 48-72] facultative feeding model.     

Patch selection by juvenile black surfperch (Embiotocidae) under variable risk: Interactive influence of food quality and structural complexity

The influence of risk on the selection of foraging patches by young-of-year black surfperch, Embiotoca jacksoni Agassiz, was investigated by laboratory and field experiments. These foragers harvest crustacean prey from a variety of benthic algal substrata. In field environments, patch types vary in two ways. First, substrata differ in structural complexity and probably afford different degrees of protection from predators. Second, substratum types vary in prey richness. There was no correlation between structural complexity and prey richness, and either or both factors could be a component of foraging patch value. Each patch is small and individual foragers are simultaneously confronted with arrays of patches encompassing the full range of variation in structure and prey richness. Furthermore, a major predator of young-of-year black surfperch, the kelp bass, Paralabrax clathratus (Girard), is patchily distributed in space and time. Thus similar arrays of patch types can be characterized by different levels of overall risk. Risk to foragers is dependent on light level as well as the presence and density of predators.The interplay between food quality and shelter potential in influencing patch choice was examined under different regimes of risk. Both laboratory and field experiments indicated patch preference was based primarily on food quality. However, the physical structure of a patch did become a component of patch choice as risk increased. The relative value of physical structure under high risk was dependent on the prey richness of a patch; food-poor substrata with high shelter potential remained unfavored even in situations of high risk.     

Taenia taeniaeformis: Increased cell growth and neutral mucus production in the gastric mucosa of the rat with a larval infection

Gross hyperplasia of the gastric mucosa and excessive mucus production in the stomach occur in rats heavily parasitized with larvae of Taenia taeniaeformis. In this study, a positive correlation between the number of larvae recovered from hepatic cysts and the weight of the stomachs of infected rats was found. By light microscopy, the hyperplasia was restricted to the glandular mucosa. Parietal and chief cells were very rare, and densely PAS-positive mucous cells were the major cell types in the hyperplastic stomach while, in comparison, alcian blue-positive cells were much fewer in number. The isolated gastric mucosa in organ culture had an increased [³H]thymidine incorporation rate in rats infected with T. taeniaeformis. The hexosamine concentration per milligram protein in the hyperplastic stomach mucosa was twice that in the control rat stomach mucosa. By electron microscopy, the apical cytoplasm of the mucous cells was found to be filled with small dark granules. These results indicate that the gastric hyperplasia is caused by stimulation of growth and major differentiation of stem cells to neutral mucus-producing cells.