Benomyl: Effectiveness against the microsporidian Nosema heliothidis in the corn earworm,Heliothis zea

The fungicide benomyl was studied as a possible antimicrobial agent for obtaining Nosema heliothidis-free laboratory colonies of Heliothis zea. Newly hatched, transovarially infected larvae were placed on artificial diets containing 250, 500, or 1000 ppm benomyl. While late-stage larvae were found to be free of Nosema spores, low-level infections were found in pupae and newly emerged adults. The reduced intensity of infection in adults reared as larvae on treated diets was not correlated with a significant reduction in the efficiency of transovarian transmission. The chemical effect of benomyl was manifested by aberrant spores and vegetative stages and a rapid reduction in the number of microsporidian stages. However, small, isolated centers of infection in various host tissues resulted in a rapid resurgence of the microsporidiosis in pupae and adults. Thus, at the concentrations tested, benomyl was not effective in eliminating infection by N. heliothidis in H. zea. A discussion of the necessity for careful evaluation of the apparent suppression of microsporidioses by antimicrobial agents is also presented.     

Susceptibility of larvae ofHeliothis zea,H. virescens,andH. armigera [lep.: Noctuidae] to 3 baculoviruses

The pattern of virulence (based on inclusion bodies) for 3 baculoviruses ofHeliothis, i.e. a unicapsid, nuclear polyhedrosis virus (HzSNPV); a multicapsid, nuclear polyhedrosis virus (HaMNPV); and a granulosis virus (HaGIV) was the same (HzSNPV>HaMNPV>HaGIV) for 3 species ofHeliothis. Based on numbers of nucleocapsids, however, the HaGIV was ca 2X more virulent than the HaMNPV for larvae ofH. virescens, (F.), and the HaMNPV was about 6X more virulent than the HaGIV for larvae ofH. armigera (Hübner). The fastest rate of larval mortality was obtained with HzSNPV. Although the mortality rate for HaGIV was faster than that of HaMNPV forH. virescens andH. armigera, it was slower than that of HaMNPV for larvae ofH. zea (Boddie). The pattern of susceptibility ofHeliothis species to HzSNPV and HaMNPV wasH. zea>H. virescens>H. armigera. Differences in susceptibility of the least susceptible species (H. armigera) and the most susceptible species (H. zea) to HzSNPV was ca. 1.6 X. Larvae ofH. zea, however, were ca. 4 to 6 X more susceptible to HaMNPV than were larvae ofH. virescens orH. armigera. A different pattern of susceptibility was recorded for HaGIV when larvae were challenged with HzSNPV and HaMNPV. Larvae ofH. virescens were ca. 20 and 35 X more susceptible to HaGIV than were larvae ofH. zea andH. armigera, respectively.     

Are stag beetles fungivorous?

Stag beetle larvae generally feed on decaying wood; however, it was unknown whether they can use wood-rotting fungi alone as food. Here, to clarify this, newly hatched larvae of Dorcus rectus (Motschulsky) (Coleoptera: Lucanidae) were reared for 14 days on artificial diets containing a fixed amount of freeze-dried mycelia of the following fungi: Bjerkandera adusta, Trametes versicolor, Pleurotus ostreatus, and Fomitopsis pinicola. The mean incremental gain in larval body mass was greatest on diets containing B. adusta, followed by T. versicolor, P. ostreatus, and F. pinicola. The growth rate of body mass correlated positively with mycelial nitrogen content of the different fungi. It also correlated positively with the mycelial content of B. adusta in the diet. Addition of antibiotics to diets with mycelia nearly halved larval growth, indicating that larvae were able to use fungal mycelia as food without the assistance of associated microbes although the microbes positively affected larval growth. Four newly hatched larvae reared on artificial diets containing B. adusta mycelia developed to the second instar in 21-34 days; and one developed to the third (=final) instar. This study provides evidence that fungi may constitute the bulk of the diet of D. rectus larvae.     

Comparison of in vivo infectivity of tissue-cultured nonoccluded virus and alkali-liberated occluded virus of Baculovirus heliothis

Feeding and intrahemocelic injection studies using tissue-culture-derived-nonoccluded virus (TCNOV) and occluded virus liberated by alkaline solution (ALOV) from polyhedral inclusion bodies were conducted with the single-embedded Heliothis nuclear polyhedrosis virus, Baculo-virus heliothis (H_zSEV). Comparisons of infectivity between ALOV and NOV were based upon the number of adminstered plaque-forming-units (PFU). There was little, if any, difference in infectivity between ALOV and TCNOV of H_zSEV when injected into 4th-instar larvae of Heliothis virescens. The LD₅₀, from the multiple dose injection studies, for ALOV and TCNOV was 6.5 ± 1.2 PFU per larva and 3.4 ± 0.9 PFU per larva, respectively. Injection of a single dose (5 PFU per larva) resulted in a larval mortality of 83.2 ± 3.4 and 62.6 ± 5.7% for ALOV and TCNOV of the H_zSEV, respectively. The LC₅₀ of ALOV and TCNOV, from the multiple-dose feeding tests, was 3.1 ± 0.4 PFU/cm² and 4.5 ± 0.9 PFU/cm², respectively. Feeding 24-hr-old larvae on virus-treated diets at a single dose (50.0 PFU/cm²) resulted in a 1.5-fold difference in percentage larval mortality between ALOV (91.0 ± 4.0%) and TCNOV (61.2 ± 3.0%). Counts of viral particles (VP), based upon electron microscopy, were 14.3 ± 2.6 × 10¹⁰ and 5.2 ± 1.1 × 10⁷ VP/ml for the ALOV and TCNOV, respectively. Thus, each larva ingesting or injected with one PFU received ca. 3500 × more VP of ALOV than in did of TCNOV.     

The effect of larval predators Thanasimus dubius (Coleoptera: Cleridae), produced by an improved system of rearing, against the southern pine beetle Dendroctonus frontalis (Coleoptera: Curculionidae)

The southern pine beetle, Dendroctonus frontalis (Coleoptera: Curculionidae) (SPB), is known to be a major bark beetle pest of pines throughout the southeastern United States. A common predator of bark beetles, Thanasimus dubius (Coleoptera: Cleridae), has been suggested to play a prevalent role on SPB dynamics. Evaluations of T. dubius have been limited by rearing methods; an artificial diet for larval T. dubius exists, and preservatives such as sorbic acid could help to maximize diet shelf-life and enhance the efficiency of the rearing system. The effects of sorbic acid at different concentrations (0%, 0.1% and 0.2%) in the larval diet for T. dubius were measured, and the effects of increased feeding time intervals (2-3 vs. 5 days) on predator performance evaluated. In addition, an experimental bioassay was conducted where newly hatched T. dubius larvae were released at four densities (0, 50, 100, and 200 per log) on pine logs infested by SPB. Sorbic acid in the diet reduced female fecundity (by 20-40%), but did not affect adult T. dubius size or longevity. However, using this preservative may not be necessary because it had no effect on the overall efficiency of the rearing system, while refreshing the larval diet every 5 days (compared with 2-3 days) did improve its efficiency, even without sorbic acid. The release of larval T. dubius resulted in a highly significant effect on the SPB ratio of increase (RI). This experiment was facilitated by the improvements in our rearing methods.     

Activity and Persistence of Steinernema carpocapsae and Spodoptera exigua Nuclear Polyhedrosis Virus against S. exigua Larvae on Soybean

Experiments were conducted to assess the effects of the entomopathogenic nematode Steinernema carpocapsae and Spodoptera exigua multinucleocapsid nuclear polyhedrosis virus (SeMNPV), alone and in combinations, on mortality of the beet armyworm, S. exigua, larvae on soybean. In 1991 tests, field-grown soybean plants were treated with S. carpocapsae at 0.3 and 0.6 nematodes/cm² of leaflet, SeMNPV at 20 and 40 polyhedral inclusion bodies (PIB)/cm², and all possible combinations. Treated leaflets were collected from plants and bioassayed with 5-day-old larvae. The combination of S. carpocapsae at 0.6 nematodes/cm² + SeMNPV at 40 PIB/cm² produced significantly higher larval mortality (61.7%) compared with either S. carpocapsae (24.8-35.1%) or SeMNPV (26.5-33.7%) alone. In 1992, similar tests were repeated using S. carpocapsae at 0.2 and 0.5 nematodes/cm², and SeMNPV at 14 and 35 PIB/cm². The combination of 0.5 nematodes/cm² + 35 PIB/cm² resulted in significantly higher larval mortality (64.0%) than either pathogen alone (41.5-49.0%). Steinernema carpocapsae and SeMNPV produced additive effects on beet arlnyworm mortality. Persistence of S. carpocapsae was 12-24 hours and SeMNPV was 96-120 hours on soybean.     

Growth and fatty acid composition of freshwater prawn, Macrobrachium rosenbergii, larvae fed diets containing various ratios of cod liver oil–corn oil mixture

A feeding trial was conducted to determine the effect of replacing costly cod liver oil with corn oil as a source of dietary lipid on the growth and fatty acid composition of the larval freshwater prawn, Macrobrachium rosenbergii de Man. Prawn larvae were weaned to artificial diets containing cod liver oil and corn oil either singly or in various combinations (2 : 1, 1 : 1, 1 : 2, w/w). Weaning to artificial diets from Artemia nauplii commenced at larval stage III with complete substitution by stage X. The reference group was reared solely on Artemia nauplii during the entire experiment. Incorporation of corn oil at 33–67% of dietary supplemental oil did not have significant effects on the post‐larval production. However, larvae fed with corn oil alone revealed a significantly lower post‐larval production compared to other experimental diets as well as to the reference group. No significant differences (P > 0.05) were observed in dry weight, protein and lipid concentration among larvae fed on various dietary treatments. Palmitic (16 : 0) and oleic/vaccenic (18 : 1) acids were the dominant saturated and monounsaturated fatty acids in larval tissues, respectively, whereas the polyunsaturated fraction was dominated by eicosapentaenoic (20 : 5n‐3) acid. The polyunsaturated fatty acid composition was dominated by n‐3 acids rather than n‐6 fatty acids. The fatty acid composition of the prawn in general reflected that of the diet. Larvae on diets containing higher concentrations of corn oil rich in linoleic (18 : 2n‐6) acid showed a higher concentration of this acid in their tissues. No evidence of de novo synthesis of linoleic (18 : 2n‐6) acid was found. Higher levels of stearic (18 : 0), arachidonic (20 : 4n‐6) and eicosapentaenoic (20 : 5n‐3) acids found in larvae as compared with those fed Artemia and artificial diets strongly indicated the larval ability in chain elongation and desaturation of palmitic (16 : 0), linoleic (18 : 2n‐6) or linolenic (18 : 3n‐3) acids, respectively. Despite a large variation of n‐3 to n‐6 ratios of the live and artificial diets, larval n‐3 to n‐6 ratios were relatively stable among different dietary treatments, possibly indicative of the importance of such a ratio in the larval fatty acid metabolism.     

Effect of varying dietary levels of ω6 polyunsaturated fatty acids during the early ontogeny of the sea scallop, Placopecten magellanicus

This study investigated the effects of selected microalgal diets on growth, survival and fatty acid composition of sea scallops, Placopecten magellanicus, from egg hatching to postlarvae. Scallops were offered Pavlova lutheri (MONO) and Pavlova sp. (strain CCMP 459, P459) in combination with Chaetoceros muelleri (CHGRA) and P459 alone. These diets exhibited varying proportions of ω6 docosapentaenoic acid (22:5ω6, DPA) which represented 8.9% of total fatty acids in P459 compared to 5.4% in P459-CHGRA and only 0.5% in MONO-CHGRA. For experiment I, larvae were fed three diets (MONO-CHGRA, P459-CHGRA or P459 alone) for 35 days after fertilization. For experiment II, larvae were fed P459-CHGRA for 21 days and split among the three diets until 39 days post-fertilization. For experiment III, 4-day-old larvae were exposed to P459-CHGRA until they become postlarvae. MONO-CHGRA failed to produce pediveligers when fed to early veligers but yielded competent larvae when fed to late veligers. This may be related to the low ω6 DPA content of this diet. High accumulation of DPA by early veligers suggests a nutritional requirement for this fatty acid. Unialgal P459 supported lower pediveliger production and lower growth rate than other diets containing CHGRA. This may be the effect of higher levels of energetic substrates in CHGRA-containing diets. Finally, P459-CHGRA yielded the best survival and growth rate during the whole larval period. All three experiments showed marked arachidonic acid (20:4ω6, AA) accumulation in early veligers (4 to 21 days) and in pediveligers (32 to 43 days). Accumulation of AA during metamorphosis might reflect stimulation of immune functions due to pathogen infection and degradation of larval organs or the role of AA in cell signalling processes associated with tissue reorganization. Early veligers markedly accumulate DPA likely in response to low docosahexaenoic acid (22:6ω3, DHA) concentrations in the three tested diets. The potential role and effect of DPA as a substitute for DHA is discussed.     

Effect of varying levels of dietary essential fatty acid during early ontogeny of the sea scallop Placopecten magellanicus

This study investigated the biological effects of various dietary essential fatty acids levels to sea scallop larvae, Placopecten magellanicus. Scallop larvae were fed three diets from D-veliger to settlement. Diet A consisted of Isochrysis sp. and Pavlova lutheri, diet B was a mix of Isochrysis sp. and Chaetoceros muelleri and diet C consisted of the same two species, but C. muelleri was grown under silicate deprivation to alter the fatty acid composition. Pediveligers (28 days old) were sampled prior to settlement for fatty acid analysis, growth measurement and survival assessment. Survival and settlement success were measured at the end of the experiment (day 40). Our results show that feeding regime greatly influenced larval size, settlement and fatty acid composition. Diet A was severely deficient in arachidonic acid (20:4n-6, AA), leading to the poorest larval growth, survival and lipid content. Nevertheless, larvae fed diet A selectively accumulate AA by a factor three compared to the dietary amount. Shell size of 28-day-old larvae was positively correlated with AA content and negatively correlated with eicosapentaenoic acid (20:5n-3, EPA)-AA ratio, thus suggesting that these two variables are of major interest for the optimisation of larval growth in sea scallops. Finally, larvae fed diet C displayed 20% higher shell size at day 28 than larvae fed diet A and B, likely in relation to the dietary amount of saturated fatty acid (SFA). However, the moderate survival and settlement success of these groups of larvae might be associated with a relative deficiency in docosahexaenoic acid (22:6n-3, DHA). This study underlines that the overall balance between polyunsaturated fatty acid (PUFA) needs to be considered to adequately fed sea scallop larvae.     

Behavioral responses of leafroller larvae to apple leaves and fruit

Larvae of Epiphyas postvittana and Planotortrix octo were released onto branches cut from apple trees, and allowed to colonize a range of types of artificial nests. Both species exhibited similar strong preferences for nests comprising leaf-leaf or leaf-fruitlet combinations, followed by nests comprising leaf-plastic leaf, leaf-plastic fruitlet or plastic leaf-fruitlet combinations. Nests involving fresh plant material alone (shoot, fruitlet or leaf alone) were also colonized to a lesser extent, but no larvae were found on nests consisting of plastic leaves or fruitlets alone or in combinations of the two. In another experiment, more E. postvittana larvae colonized nests with leaf and fruitlet combinations, compared to leaf and glass ball, or leaf and treated wax ball combinations, where the wax had been in contact with fresh apples or fruitlets. Choice tests, conducted using larval traps, showed that larvae were caught in traps baited with odors collected and released by wax which had been in contact with mature apples and leaves. Chloroform extracts from apple skin also caught larvae in choice tests. These results suggest that both physical and chemical cues are important to leafroller larval establishment.     

Ingestion rates and dietary lipids affect growth and fatty acid composition of Dendraster excentricus larvae

This study investigated the effect of single and mixed algal diets on growth, fatty acid composition and ingestion rates for Dendraster excentricus larvae. Larvae were assigned to three single algal diet treatments Isochrysis galbana, Dunaliella tertiolecta or Rhodomonas sp. and four mixed algal diet treatments D. tertiolecta and Rhodomonas, I. galbana and D. tertiolecta, I. galbana and Rhodomonas, D. tertiolecta, Isochrysis galbana and Rhodomonas sp. Small amounts (0.36-0.6%) of stearic acid (18:0) were seen in the three algae used but a relatively large percentage (7-25%) of this SAFA was found in Dendraster larvae. The alga D. tertiolecta had the highest percentage (51.7%) of the short chain polyunsaturated fatty acid (PUFA) linolenic acid 18:3(n-3) and trace amounts (0.02-0.14%) of the long chain PUFAs eicosapentanoic (EPA, 20:5(n-3) and docosahexanoic acids (DHA, 22:6(n-3)). However, sand dollar larvae demonstrated the ability to elongate and desaturate shorter chain (18 carbon) polyunsaturated fatty acids (PUFA) to longer chain (20 carbon) n-3 PUFA. Thus high levels of 18:3(n-3) in D. tertiolecta led to high levels of EPA and low levels of 18:3(n-3) in Dendraster larvae fed this diet. Rhodomonas sp. had the highest percentage of stearidonic acid (18:4(n-3), 38.14%) and EPA (10.6%). Despite high levels of 18:4(n-3) in Rhodomonas sp. this acid was absent or found at very low levels in larvae fed this alga, or any combination of this alga. I. galbana had the highest percentage of DHA(14.3%) but was almost devoid of EPA (0.43%). Although Rhodomonas sp. and I. galbana had high levels of EPA and DHA sand dollar larvae did not incorporate higher levels of these long chain PUFAs into their lipids compared to those fed the alga D. tertiolecta. Dendraster larvae synthesized a number of 20 and 22 carbon non-methylene interrupted dienes (NMID), with levels increasing with larval stage. Higher ingestion rates were observed for Dendraster larvae fed single algal diets (Rhodomonas sp. or D. tertiolecta) and lower ingestion rates for those fed mixed algal diets. The highest ingestion rates were for 8-arm Dendraster larvae fed the large alga Rhodomonas sp. presented as a single algal diet. When fed a combination of three algae, selection of particles varied slightly depending on stage with 8-arm larvae ingesting slightly more of the larger algal cell in the mixture than 6-arm larvae. The present study suggests that regardless of the ratios given larvae might have an optimum ratio of different sized particles at which they can feed. The mixed algal diet of I. galbana and D. tertiolecta was the best algal diet leading to significantly larger larvae with high survival and development to metamorphosis. The single algal diets of Rhodomonas sp. or Dunaliella tertiolecta were the second and third best algal diets based on growth and survival to metamorphosis.     

Larval feeding behavior of Dipel-resistant and susceptible Ostrinia nubilalis on diet containing Bacillus thuringiensis (Dipel ESTM)

Diet choice tests were conducted to examine the effects of Bacillus thuringiensis Berliner on larval feeding behavior of Dipel-resistant and susceptible strains of European corn borer, Ostrinia nubilalis (Hübner). Larvae (first through fourth instars) were presented an untreated standard diet and three diets incorporating different concentrations of a commercial formulation of B. thuringiensis, Dipel ES^TM. Significantly higher proportions of susceptible and resistant larvae were found on the control diet compared to the proportions recorded on any of the Dipel-treated diets. Soon after release (0.5 to 3 h), the occurrence of larvae on each of the diets was similar across different instars. The number of first and second instars on the control diet increased steadily over 72 h after release, when 43 to 75% of larvae were found on the control diet. The proportion of third and fourth instars on the control diet also increased initially, with an equilibrium occurring 12 to 24 h later when 30 to 40% of larvae were found on the control diet. Both strains appeared to avoid the Dipel-treated diets and their responses were similar over different Dipel concentrations. The Dipel-resistant strain showed higher ability to avoid the treated diets than the susceptible strain.     

Feeding History Affects Intraguild Interactions between Harmonia axyridis (Coleoptera: Coccinellidae) and Episyrphus balteatus (Diptera: Syrphidae)

While the effect of several factors such as predator and prey size, morphology and developmental stage on intraguild predation (IGP) is widely investigated, little is known about the influence of diet on the occurrence and outcome of IGP. In the present study, the effect of the diet experienced during larval development on IGP between the ladybird Harmonia axyridis and the syrphid Episyrphus balteatus is investigated. Four diets were tested for H. axyridis: eggs of the Mediterranean flour moth Ephestia kuehniella, pea aphids, Acyrthosiphon pisum, in an ad libitum amount, pea aphids in a limited amount, and honey bee pollen. For E. balteatus only the two aphid diets were tested. First, experiments were performed to determine the quality of the various diets for development of both predators. Second, IGP experiments between H. axyridis and E. balteatus were performed both in Petri dishes and on potted pepper plants. The diet of both species influenced the incidence of IGP between H. axyridis and E. balteatus both in Petri dishes and on potted plants. In general, smaller larvae of H. axyridis (those fed on poor or restricted diet) fed more on hoverflies than large (well-nourished) ladybird larvae. Further, poorly nourished (smaller) larvae of E. balteatus were more susceptible to predation than well-fed (larger) hoverfly larvae. The observed effects were not only due to the lower fitness of larvae of both predators reared on an inferior quality diet but also to changes in predator behaviour. The results from this study show that IGP interactions are influenced by a multitude of factors, including feeding history of the organisms involved, and emphasize the importance of taking these factors into account in order to fully understand the ecological relevance of IGP.     

Optimization of Methodology for Rearing <Emphasis Type="Italic">Spodoptera albula</Emphasis> on Artificial Diet

Advances in techniques for rearing insects on artificial diets are fundamental to solving issues of basic and applied entomology. In this study, we evaluated the development of Spodoptera albula (Walker) (Lepidoptera: Noctuidae) on three artificial diets used for other species of Lepidoptera, at three larval densities, and two densities of adult couples housed in oviposition cages of two sizes, with the aim of optimizing methodology for rearing S. albula in the laboratory. Biological parameters were recorded from S. albula, and a fitness index was calculated based on the larval survival and duration and weight of pupae. The total and daily oviposition was recorded using 5 or 10 adult couples of S. albula housed in two cage sizes. Concentrations of total nitrogen and protein in the tested diets were determined. Development of S. albula was completed in all artificial diets; however, the diet used for rearing Anticarsia gemmatalis (Hübner) larvae was the most suitable for S. albula, yielding intermediate development time and higher survival relative to the other diets. Individualization of larvae favored S. albula development by producing overall greater weights of larvae and pupae, higher survival rates, and longer adult longevity. Cage size and number of couples per cage did not influence S. albula fecundity in the experiment conditions. Spodoptera albula can be satisfactorily reared on the artificial diet used for A. gemmatalis, using one larva per tube, and either density of adults at any cage size. Additional amendments are needed in the rearing methodology to achieve optimal conditions for larval development to adulthood.     

Lipid nutrition during larval development of the parasitic wasp, Exeristes

The parasite Exeristes roborator grew rapidly and completed larval development on fatty acid free chemically defined diets. Dietary supplements of palmitoleate, oleate, linoleate, and linolenate were detrimental to parasite development with most larvae dying in the first instar. Palmitate supplements were also toxic, but a small percentage of larvae consistently completed development and survival and development time on diets supplemented with free stearate did not differ significantly from results obtained with fatty acid free diets. Supplements of a mixture of all six free fatty acids were as toxic as the unsaturated free fatty acids. Dietary supplements of the triglycerides, tripalmitin, tripalmitoleate, tristearin, and trioleate had no positive nutritional value for larval growth and development but were not detrimental.Development time was increased when the parasite was reared on fatty acid free diets lacking carbohydrate, but survival was not affected. The parasite, therefore, appears to have the ability to utilize dietary free amino acids as the sole energy source at this stage. Under these nutritional conditions, supplements of triglycerides did not replace the nutritional value of carbohydrate and some of the supplementary triglycerides were detrimental to larval survival.     

Drosophila adult and larval pheromones modulate larval food choice

Insects use chemosensory cues to feed and mate. In Drosophila, the effect of pheromones has been extensively investigated in adults, but rarely in larvae. The colonization of natural food sources by Drosophila buzzatii and Drosophila simulans species may depend on species-specific chemical cues left in the food by larvae and adults. We identified such chemicals in both species and measured their influence on larval food preference and puparation behaviour. We also tested compounds that varied between these species: (i) two larval volatile compounds: hydroxy-3-butanone-2 and phenol (predominant in D. simulans and D. buzzatii, respectively), and (ii) adult cuticular hydrocarbons (CHs). Drosophila buzzatii larvae were rapidly attracted to non-CH adult conspecific cues, whereas D. simulans larvae were strongly repulsed by CHs of the two species and also by phenol. Larval cues from both species generally reduced larval attraction and pupariation on food, which was generally—but not always—low, and rarely reflected larval response. As these larval and adult pheromones specifically influence larval food search and the choice of a pupariation site, they may greatly affect the dispersion and survival of Drosophila species in nature.     

Suitability of monotypic and mixed diets for Anopheles hermsi larval development

The developmental time and survival to eclosion of Anopheles hermsi Barr & Guptavanij fed monotypic and mixed diets of ten food types were examined in laboratory studies. Larvae fed monotypic diets containing animal detritus (freeze‐dried rotifers, freeze‐dried Daphnia pulicaria, and TetraMin® fish food flakes) and the mixotrophic protistan Cryptomonas ovata developed faster and survived better than larvae that were fed other monotypic diets. Survival to adulthood of larvae fed several concentrations of the diatom Planothidium (=Achnanthes) lanceolatum was poor (<13%) and larval development time was approximately twice that of larvae fed TetraMin® fish food flakes, the standard laboratory diet. Larvae fed monotypic diets containing prokaryotes (bacteria [Bacillus cereus] and cyanobacteria [Oscillatoria prolifera]) and brewer's yeast (Saccharomyces cerevisiae) failed to survive beyond the 1^st and 2^nd instar, respectively. Larvae fed only chlorophytes, single‐celled Chlamydomonas reinhardtii and filamentous Spirogyra communis, failed to complete larval development, regardless of the concentration tested. Cohorts fed a combination of food types (mixed diets) usually developed better than cohorts fed monotypic diets. Food types that failed to support complete development when fed alone often facilitated development to adulthood when fed in combination with food types containing >1% C₂₀ polyunsaturated fatty acids as total fat, but regardless of essential fatty acid content, algae that produced mucilage and filaments that sank out of the feeding zone were poor quality diets.     

The effect of bacterial and diatom biofilms on the settlement of the bryozoan Bugula neritina

Biofilms of marine bacteria and diatoms and their combinations were examined in laboratory choice assays to determine their effects on the attachment and successful metamorphosis of the larvae of the bryozoan Bugula neritina (Linnéus). The larval settlement in response to unfilmed surfaces, a natural biofilm (NBF) and adsorbed cells of three strains of bacteria, five strains of pennate diatoms and combinations of the two at different densities. Bacterial and diatom strains showed different effects on the larval settlement of B. neritina. Bacterial monospecific strains of an unidentified α-Proteobacterium and Vibrio sp. mediated the same percentage of settlement as a filtered seawater control. Biofilms of Pseudoalteromonas sp. caused significantly lower larval settlement. Larval settlement of B. neritina was negatively correlated with increasing densities of Pseudoalteromonas sp. The highest percentages of settlement were mediated by the biofilms of the diatom species Achnanthes sp., Amphora cofeaeformis, Amphora tenerrima, Nitzschia constricta and a 5-day-old natural biofilm, while the lowest settlement was found on a N. frustulum film. A three-way analysis of variance demonstrated that the density of bacteria and the presence of particular species of diatoms and bacteria in combined biofilms, significantly affected the settlement of B. neritina larvae. High settlement of larvae (50-90%) at all treatments indicated that B. neritina larvae are much more indiscriminate settlers than previously expected. Hence, using this species as a monitoring organism to trace ecologically relevant subtle changes of settlement cues in the natural environment should be carefully re-examined.     

Role of Bacillus thuringiensis Cry1 δ Endotoxin Binding in Determining Potency during Lepidopteran Larval Development

Five economically important crop pests, Manduca sexta, Pieris brassicae, Mamestra brassicae, Spodoptera exigua, and Agrotis ipsilon, were tested at two stages of larval development for susceptibility to Bacillus thuringiensis toxins Cry1Ac, Cry1Ca, Cry1J, and Cry1Ba. Bioassay results for M. sexta showed that resistance to all four Cry toxins increased from the neonate stage to the third-instar stage; the increase in resistance was most dramatic for Cry1Ac, the potency of which decreased 37-fold. More subtle increases in resistance during larval development were seen in M. brassicae for Cry1Ca and in P. brassicae for Cry1Ac and Cry1J. By contrast, the sensitivity of S. exigua did not change during development. At both larval stages, A. ipsilon was resistant to all four toxins. Because aminopeptidase N (APN) is a putative Cry1 toxin binding protein, APN activity was measured in neonate and third-instar brush border membrane vesicles (BBMV). With the exception of S. exigua, APN activity was found to be significantly lower in neonates than in third-instar larvae and thus inversely correlated with increased resistance during larval development. The binding characteristics of iodinated Cry1 toxins were determined for neonate and third-instar BBMV. In M. sexta, the increased resistance to Cry1Ac and Cry1Ba during larval development was positively correlated with fewer binding sites in third-instar BBMV than in neonate BBMV. The other species-instar-toxin combinations did not reveal positive correlations between potency and binding characteristics. The correlation between binding and potency was inconsistent for the species-instar-toxin combinations used in this study, reaffirming the complex mode of action of Cry1 toxins.     

Activation of the ambusher tick Rhipicephalus microplus (Acari: Ixodidae) exposed to different stimuli

The present study aimed to evaluate the behaviour of larvae of Rhipicephalus microplus exposed to different stimuli. A Y-olfactometer was positioned vertically and R. microplus larvae were exposed to environmental air, CO₂ alone, N,N-diethyl-3-methylbenzamide (DEET) alone, and CO₂ combined with the repellents DEET and (E)-2-octenal. Tests were also conducted with the olfactometer positioned horizontally; in this case, however, only CO₂ was tested. In all tests conducted with the Y-olfactometer positioned vertically, CO₂ activated R. microplus larvae even in the presence of DEET and (E)-2-octenal, although activation was lower when these repellents were used. In the absence of CO₂, larval behaviour against DEET was similar to that of the larvae in the control group. In the tests performed with the olfactometer positioned horizontally, the larvae had no significant response to the presence of CO₂. The larvae were not attracted to or repelled by any compound tested in either the vertical or horizontal position of the olfactometer. The lack of horizontal displacement, attraction or repellence may have been a result of the ambush behaviour of this tick species. However, when larvae were exposed to stimuli and the olfactometer was positioned vertically, the interference of attractant and repellent stimuli in larval behaviour was assessed.