Larval recognition by Temnothorax longispinosus and T. ambiguus hosts of the slave-making ant Protomognathus americanus revisited: colony-level referent ensures conspecific preference

Slave-making ants exploit the labour of their own or another species. Temnothorax ambiguus and T. longispinosus are both ant species that serve as hosts of the obligatory slave-making ant Protomognathus americanus and are facultative slave-makers themselves. We offered laboratory colonies of T. ambiguus and T. longispinosus a series of choices among different larval types to better understand their brood discrimination abilities. Workers of both species preferentially accepted nestmate over non-nestmate larvae. Both species preferentially retrieved unrelated conspecific larvae over congeneric allospecific larvae, and T. ambiguus workers consumed more allospecifics than conspecifics. When presented with conspecific versus P. americanus larvae, both species manifested a clear bias towards conspecific larvae in terms of earlier retrieval and reduced cannibalism. That workers did not prefer P. americanus larvae over conspecific larvae as documented in previous research likely reflects the fact that in the present study, subject workers had access to the entirety of their colony as a referent during rearing and at the time of testing, as they would in nature. Moreover, reciprocal contact between P. americanus and conspecific larvae increased acceptance of the slave-maker larvae, but did not appear to lessen the acceptability of conspecific larvae. This suggests that transfer of cues through contact may be sufficient to alter the recognition signature of P. americanus larvae increasing acceptability by their hosts.     

Differential susceptibility of parasitized and nonparasitized larvae of Trichoplusia ni to a nuclear polyhedrosis virus

Nonparasitized second-instar larvae of Trichoplusia ni were twice as susceptible (at the LD₅₀ level) to the singly enveloped T. ni nuclear polyhedrosis virus as those parasitized by Hyposoter exiguae (Hymenoptera: Ichneumonidae). The LD₅₀ values for nonparasitized and parasitized larvae were 1.58 × 10³ and 3.16 × 10³ polyhedra/ml of diet, respectively. The LD₉₅ value for parasitized larvae was approximateely 5 times higher than that for nonparasitized larvae. The slopes (b values) were 1.2 for parasitized larvae and 1.7 for nonparasitized larvae. The LT₅₀ values for parasitized larvae also were significantly longer than those for nonparasitized larvae. No significant difference was found between the food consumption of parasitized and nonparasitized T. ni larvae.     

The mode of transmission of Tipula iridescent virus: II. Route of infection

Tipula iridescent virus (TIV) was inoculated into Tipula oleracea larvae via different routes. It was found to be much more infective when it was injected into the hemocoel than when it was ingested by the larvae.T. oleracea embryos did not become infected when eggs were laid in agar containing TIV, and there was no evidence that larvae can become infected via the spiracles or that transovum transmission occurs. It is suggested that TIV is transmitted principally by cannibalism, including killing and ingesting infected larvae, and finding dead infected larvae and ingesting them. It is proposed that a new generation becomes infected by first-instar larvae feeding upon infected fourth-instar larvae which have survived from the previous generation.     

Effects of the parasitization of a braconid, Apanteles, on the blood of its host, Pieris

Parasitization of a braconid wasp, Apanteles glomeratus, of larvae of a common cabbage butterfly, Pieris rapae crucivora, caused changes in differential haemocyte count (DHC), total haemocyte count (THC), and encapsulative capacity against dead eggs of Apanteles in the fourth and fifth instar host larvae.However, no correlation could be found between the number of Apanteles eggs deposited and THC of the middle fourth instar host larvae or between the number of parasitoid larvae and specific gravity of the haemolymph from the late fifth instar host larvae.From the changes in DHC and in THC of both non-parasitized and parasitized Pieris larvae, an increase in the number of plasmatocytes of non-parasitized Pieris larvae in the early fourth instar period was supposed to be due to transformation of prohaemocytes into plasmatocytes, and a low population of plasmatocytes of parasitized larvae in the comparable period was assumed to be due to a suppression of transformation of prohaemocytes by some factor released from the parasitoid eggs.Failure of the parasitized fourth instar Pieris larvae to encapsulate injected dead eggs of Apanteles indicated that the parasitoid embryos were, in some way, actively inhibiting the encapsulation reactions of the host.The increase in THC of the parasitized fifth instar larvae could not be ascribed to a decrease in the volume of host haemolymph. Rather it could be interpreted by a suppression of adhesive capacity of haemocytes in the host haemocoel to tissue surfaces.Reduced encapsulative capacity of the parasitized fifth instar larvae might be attributed either to a depression of the adhesive activity of plasmatocytes resulting from a depletion of energy source for haemocytes in the host haemolymph by parasitization, or from an active suppression of adhesiveness of the plasmatocytes by secretions from ‘giant cells’ (teratocytes) originated from the parasitoid.     

Selective breeding and characterization of a black mealworm strain of Tenebrio molitor Linnaeus (Coleoptera: Tenebrionidae)

Larvae of Tenebrio molitor Linnaeus are edible insects and are approved as a food ingredient in Korea. They are typically yellow; however, rare black larvae have been found in breeding boxes at insect farms. It is not clear whether black larvae represent a different species that invaded and hybridized with the yellow larvae of T. molitor or whether T. molitor shows intraspecific color variation. In this study, we characterized and identified black larvae for applications in industrial fields as well as accurate breeding and management. First, in a comparative analysis, we did not detect differences in the morphological characteristics of yellow and black larvae and adults, with the exception of larval body color. For accurate species identification, molecular analyses (p-distances and neighbor-joining) were performed based on partial COI sequences of 33 yellow and seven black larvae. Genetic divergence between yellow and black larvae ranged from 0.0% to 2.1%, revealing intraspecific variation. A neighbor-joining analysis strongly supported the classification of the two morphs as a single species. Black larvae were separated from yellow larvae and maintained by selective breeding. As a result, black larvae were completely fixed in the F2 generation (F1 = 96% and F2 = 100%). Yellow and black larvae showed no significant differences in developmental characteristics and fecundity. These findings improve our understanding of diversity within an important edible insect species and contribute to quality assurance in the food industry based on clear species identification.     

Quantifying horizontal transmission of Nosema lymantriae, a microsporidian pathogen of the gypsy moth, Lymantria dispar (Lep., Lymantriidae) in field cage studies

Nosema lymantriae is a microsporidian pathogen of the gypsy moth, Lymantria dispar that has been documented to be at least partially responsible for the collapse of L. dispar outbreak populations in Europe. To quantify horizontal transmission of this pathogen under field conditions we performed caged-tree experiments that varied (1) the density of the pathogen through the introduction of laboratory-infected larvae, and (2) the total time that susceptible (test) larvae were exposed to these infected larvae. The time frame of the experiments extended from the early phase of colonization of the target tissues by the microsporidium to the onset of pathogen-induced mortality or pupation of test larvae. Upon termination of each experiment, the prevalence of infection in test larvae was evaluated. In the experiments performed over a range of pathogen densities, infection of test larvae increased with increasing density of inoculated larvae, from 14.2 ± 3.5% at density of 10 inoculated per 100 larvae to 36.7 ± 5.7% at 30 inoculated per 100 larvae. At higher densities, percent infection in test larvae appeared to level off (35.7 ± 5.5% at 50 inoculated per 100 larvae). When larval exposure to the pathogen was varied, transmission of N. lymantriae did not occur within the first 15 d post-inoculation (dpi) (11 d post-exposure of test larvae to inoculated larvae). We found the first infected test larvae in samples taken 20 dpi (16 d post-exposure). Transmission increased over time; in the cages sampled 25 dpi (21 d post-exposure), Nosema prevalence in test larvae ranged from 20.6% to 39.2%.     

Cuticular sensilla on newly hatched larvae of Cuterebra fontinella Clark (Diptera : Cuterebridae) and Hypoderma spp. (Diptera : Oestridae)

Cuticular sensilla on newly hatched larvae of Cuterebra fontinella Clark (Diptera : Cuterebridae), Hypoderma lineatum (de Vill.) and H. bovis (L.) (Diptera : Oestridae) were studied by scanning electron microscopy. Three basic types of sensilla were found on C. fontinella larvae, while only 2 were present on the larvae of Hypoderma spp. Sensilla were more numerous on the C. fontinella larvae than on the Hypoderma spp. larvae (210 vs 66). There were no noticeable differences between the sensory complements of H. lineatum and H. bovis. Comparison of the number and type of sensilla on the parasitic larvae with those on free-living muscomorph Diptera larvae indicate that increasing numbers and greater morphological diversity correlate with increasing requirements for location of a suitable environment in which to feed and develop.     

The comparative persistence of toxicity of Bacillus sphaericus strain 1593 and Bacillus thuringiensis serotype H-14 against mosquito larvae in different kinds of environments

Bacillus sphaericus strain 1593 and B. thuringiensis serotype H-14 were evaluated for persistence of toxicity against two species of mosquito larvae, Culex quinquefasciatus and Aedes aegypti, in a selected simulating plot in Bangkok. Both strains of bacteria demonstrated larvicidal activity towards both species of mosquito larvae. In tap water, the toxicity of B. sphaericus strain 1593 was found to be greater towards C. quinquefasciatus larvae than A. aegypti larvae, whereas the toxicity of B. thuringiensis serotype H-14 was found to be greater towards A. aegypti larvae than C. quinquefasciatus larvae. The persistence of toxicity of these two bacteria was found to be different. The lethal concentration of B. thuriengiensis H-14 against A. aegypti decreased from LC₉₀ to below LC₅₀ in about 15 weeks when tested in tap water. The decrease was faster in polluted water. The toxicity of B. sphaericus 1593 towards C. quinquefasciatus larvae persisted for at least 9 months in tap water and 6 months in polluted water. The multiplication of bacteria was indicated only in populations of B. sphaericus 1593 tested with C. quinquefasciatus larvae.     

Toxic effects of spore/crystal ratios of Bacillus thuringiensis on European corn borer larvae

Bioassays to determine LC₅₀ values of spores and crystals of four varieties of Bacillus thuringiensis grown on nutrient agar plates were carried out against neonate and 6-day-old European corn borer, Ostrinia nubilalis, larvae. The four bacterial varieties were equally toxic against the neonates, but only B. thuringiensis var. kenyae, var. galleriae, and var. kurstaki were toxic to 6-day-old larvae. B. thuringiensis var. tolworthi was inactive against 6-day-old larvae. Different ratios of pure spores and crystals of the bacteria also were tested against neonate and 6-day-old larvae. Pure spores are not pathogenic to neonates or 6-day-old larvae. Pure crystals were toxic to both ages of the larvae, but a combination of spores and crystals was necessary for maximum larval mortality.     

Differences in the susceptibility of five herbivore species and developmental stages to tomato resistance induced by methyl jasmonate treatment

We compared the susceptibility of five herbivores to tomato resistance induced by methyl jasmonate (MeJA) treatment. We tested for lethal effects against five herbivores (Spodoptera litura, Mamestra brassicae, Frankliniella occidentalis, Tetranychus urticae, and Henosepilachna vigintioctopunctata) at various MeJA concentrations. The mortality of all five herbivores increased significantly with increasing MeJA concentration. The 25 % lethal concentration was 0.03 μM for both first-instar larvae of S. litura and third-instar larvae of M. brassicae, 0.51 μM for third-instar larvae of S. litura, 0.76 μM for adult T. urticae, 2.4 μM for first-instar larvae of F. occidentalis, and 5.7 μM for first-instar larvae of H. vigintioctopunctata. Thus, the degree of susceptibility to MeJA-induced resistance of tomato was first-instar larvae of S. litura = third-instar larvae of M. brassicae > third-instar larvae of S. litura ≈ adult T. urticae > first-instar larvae of F. occidentalis > first-instar larvae of H. vigintioctopunctata. Mortality of first-instar larvae of M. brassicae was >90 % at all concentrations. Mortality of fourth-instar larvae of H. vigintioctopunctata (<7 %) was similar to that of the control at all MeJA concentrations. We also detected statistically significant weight loss of the surviving lepidopteran larvae, increased larval duration of F. occidentalis and H. vigintioctopunctata, and reduced egg production by T. urticae grown on MeJA-treated tomato, suggesting that the MeJA-induced resistance can control these herbivores, but effectiveness is different on different species and growth stage. Feeding by both M. brassicae and H. vigintioctopunctata larvae activated JA-inducible genes in tomato.     

Germination of Bacillus thuringiensis var. israelensis spores in the gut of Aedes larvae (Diptera: Culicidae)

In laboratory experiments, germination and growth of Bacillus thuringiensis israelensis in the gut of Aedes aegypti and A. vexans larvae (Culicidae: Diptera) was observed. The number of spores and vegetative cells in the gut of living larvae and in cadavers was estimated by plaing homogenized larvae on selective agar plates. The number of spores per gut increased in the first 40–140 min of exposure to a maximum, and decreased in the subsequent time, demonstrating spore germination in living larvae, moribunds, and in cadavers. Twenty-four hours after the death of the larvae, a minimal amount of spores, but an increased number of vegetative cells, was found in cadavers. In A. aegypti larvae, germination and growth of B. thuringiensis israelensis in the larval gut was photographically documented.     

Predation byXylocoris flavipes [Hem.: Anthocoridae]: Influence of stage,species and density of prey and of starvation and density of predator

When the efficacy of the predaceous bugXylocoris flavipes (Reuter) was tested in the laboratory as a biological control agent against stored-product insects, it attacked eggs more than larvae, pupae, or adults and early-stage larvae ofTribolium castaneum (Herbst) more than late-stage larvae. The predator also killed more larvae ofT. castaneum than larvae ofAttagenus megatoma (F.). The density of predator and prey regulated the capture rate of the predator. Predation byX. flavipes was uninfluenced by starvation for as much as 96 hr.     

Interaction of larval age and dietary formaldehyde on the susceptibility of tufted apple budmoth (Lepidoptera: Tortricidae)to Bacillus thuringiensis

Laboratory tests of Bacillus thuringiensis var. kurstaki (strain HD-1) against Platynota idaeusalis reared on a semisynthetic diet showed that the LC₅₀ values for neonates and 9-day-old larvae showed no significant difference in susceptibility; however, 14-day-old larvae were significantly more susceptible than neonates or 9-day-old larvae. There was no evidence of any interaction between 14-day-old larvae reared on the semisynthetic diets with and without formaldehyde and B. thuringiensis toxicity at any assessment reading. Likewise the toxicity of B. thuringiensis to 14-day-old larvae reared on apple leaves did not differ from the B. thuringiensis toxicity of 14-day-old larvae reared on semisynthetic diets with and without formaldehyde.     

The significance of diet in the growth and development of larvae of the blue crab,Callinectes sapidus rathbun,under laboratory conditions

The following protistan diets were tested on blue crab larvae: the algae Isochrisis galbana Parke, Monochrisis lutheri Droop, Dunaliella sp., and an unknown mixture; and the ciliated protozoans Euplotes vannus Muller and Parauronema virginianum(2/1) Thompson. None of these diets resulted in development past the first zoea stage, although some apparently were ingested and delayed mortality as compared to unfed controls.The rotifer Brachionus plicatilis Müller sustained good survival through early zoea development; however, rotifer-fed larvae did not metamorphose to the megalopa. Larvae of the polychaete Hydroides dianthus (Verrill) sustained crab larvae throughout zoea development, resulting in 17% survival to metamorphosis. The percentage mortality per stage was significantly lower in polychaete-fed larvae when compared with rotifer-fed larvae during zoea stages III, VI, and VII. Mean intermolt duration varied between diet treatments during the first three stages, but showed no differences during later zoea development. In tests on groups of late stage sibling larvae, Artemia salina L. nauplii gave development to metamorphosis, whereas rotifers did not.All the diets so far tested on blue crab larvae are classified according to their ability to sustain development. It is demonstrated that the two diets which allow completed development, Hydroides dianthus larvae and Artemia salina nauplii, contain 2–3 times as much lipid per dry weight as do rotifers. A metabolic requirement for lipid late in development may be indicated. Invertebrate larvae derived from yolky telolecithal and centrolecithal eggs may be an important dietary component for brachyuran larvae.     

Influence of primary infection on the population dynamics of Nematospiroides dubius after challenge infections in mice

Dobson C., Sitepu P. and Brindley P. J. 1985. Influence of primary infection on the population dynamics of Nematospiroides dubius after challenge infections in mice. International Journal for Parasitology15: 353–359. Similar proportions of the inoculum of Nematospiroides dubius larvae reached sexual maturity by 14 days after administration of 50–400 larvae but more adult worms had been expelled by day 63 after infection from those mice infected with 50 vs 400 larvae. There was a significant correlation between time and worm expulsion for all inoculum size groups except for mice given 400 larvae.In mice reinfected with 100 larvae, after termination of primary infections derived from 10 through 400 larvae, more worms from the challenging dose were recovered from mice given greater compared with those given smaller numbers of larvae at primary infection. The N. dubius population size after challenge infection was correlated positively both with number of larvae administered as the primary infection and with the resultant population size during that infection. The serum anti-N. dubius antibody titres after reinfection were higher in mice given 400 compared with those given fewer larvae at primary infection, and the fecundity and female to male sex ratio of the N. dubius populations decreased in proportion to these antibody titres.Protective immunity against challenge N. dubius infection, in mice which had been drenched free of adult worms established from 400 larvae for 5 down to 1 weeks before reinfection, increased from 45% (1 week) to 80% (5 weeks). There was a negative correlation between the population size of N. dubius during challenge infection and the duration between anthelmintic treatment and challenge infection.     

Number of Malpighian Tubules in Larvae and Adults of Stingless Bees (Hymenoptera: Apidae) from Amazonia

The number of Malpighian tubules in larvae and adults of bees is variable. Larvae of Apis mellifera L. have four Malpighian tubules, while adults have 100 tubules. In stingless bees, this number varies from four to eight. The objectives of this study were to provide characteristics of the Malpighian tubules as well as to quantify their number in larvae and adults of six species of Meliponinae, Melipona seminigra merrillae Cockerell, Melipona compressipes manaosensis Schwarz, Melipona rufiventris Lepeletier, Scaptotrigona Moure, Frieseomelitta Ihering, and Trigona williana Friese. Malpighian tubules were dissected from larvae and adults, measured, quantified, and maintained in microtubes with Dietrich??s solution. The numbers of Malpighian tubules were constant only for larvae of M. rufiventris (four and eight) and Scaptotrigona sp. (four). The most frequent number of tubules in the Melipona group was seven and eight in larvae, and 70 and 90 in adults. In the Trigona group were four and 20 to 40, for larvae and adults, respectively. The results showed differences in the number of Malpighian tubules among the species analyzed and also between the larvae and adults of the same species. Despite the variation observed, species of the group Melipona always have a larger number and longer Malpighian tubules in both larvae and adults as compared to the Trigona group, which may indicate an evolutionary trend of differentiation between these groups.     

Silencing of acetylcholinesterase gene of Helicoverpa armigera by siRNA affects larval growth and its life cycle

RNA interference is an effective means of regulation of gene expression both in vitro and in vivo. We studied the effect of siRNA on larval development by selective targeting of the acetylcholinesterase (AChE) gene of Helicoverpa armigera. Chemically synthesized siRNA molecules were directly fed to H. armigera larvae along with the artificial diet. The siRNA treatment resulted in specific gene silencing of AChE and consequently brought about mortality, growth inhibition of larvae, reduction in the pupal weight, malformation and drastically reduced fecundity as compared to control larvae. Our studies suggest some novel roles for AChE in growth and development of insect larvae and demonstrate that siRNA can be readily taken up by insect larvae with their diet.     

Insecticidal activity of Bacillus laterosporus

Strains of Bacillus laterosporus demonstrated pathogenic activity for second-instar larvae of the mosquito, Culex quinquefasciatus, but failed to demonstrate detectable pathogenicity against larvae of the cabbage looper, Trichoplusia ni. Of 29 strains of the bacterium screened, 16 displayed pathogenicity for mosquito larvae. One of the most pathogenic strains, NRS 590, also demonstrated pathogenic activity for larvae of the mosquito, Aedes aegypti, and for larvae of the black fly, Simulium vittatum. The pathogenicity for Culex larvae was associated with the cell mass rather than with the culture supernatant. A suspension of ultraviolet irradiation-killed cells demonstrated no loss in pathogenic activity, an indication that the pathogenicity is toxin mediated. The toxic substance produced by NRS 590 was found to be resistant to heating at 96°C for 10 min. The toxin was not associated with the heat-resistant, bacterial endospore or with the associated paraspore since a suspension consisting primarily of spores was not toxic to mosquito larvae. Toxic activity in stationary phase cells of NRS 590 was associated with the cell's particulate fraction rather than with the soluble fraction.     

Larval diapause of the European corn borer, Ostrinia nubilalis: Further experiments examining its hormonal control

The possible involvement of juvenile hormone (JH) in controlling the mature larval diapause of the European corn borer, Ostrinia nubilalis, was examined using biological and chemical assays for JH titres, topical applications of JH mimic, and injections of 20-hydroxy-ecdysone. Bioassays of extracts of larval haemolymph showed that (1) 4th instar pre-diapausing larvae had a higher JH titre (ca. 1450 Galleria Units (GU)/ml) than equivalent non-diapausing larvae (ca. 340 GU/ml), and that (2) 5th instar pre-diapausing larvae contained a JH titre of ca. 320 GU/ml, which declined to ca. 90 GU/ml in newly-diapaused larvae. Chemical assasys carried out on extracts of whole larvae showed that early diapausing larvae contained an extremely low titre of JH. In addition, the application of JH mimic or 20-hydroxy-ecdysone or both agents to diapausing larvae failed to reveal the presence of a functional JH titre during diapause. The application of JH mimic to early 5th instar non-diapausing larvae produced moribund larval-pupal intermediates rather than supernumerary larvae. Our results, therefore, suggest that although JH may control some phases of diapause induction, it is not involved in maintaining diapause.     

Susceptibility of early larval stages of Pseudaletia unipuncta and Spodoptera exigua (Lepidoptera: Noctuidae) to the entomogenous nematode Steinernema feltiae (Rhabditida: Steinernematidae)

Early stages (neonate to 7- or 8-day-old larvae) of Spodoptera exigua and Pseudaletia unipuncta were exposed to the entomogenous nematode, Steinernema feltiae, at concentrations of 0, 10, 25, 60, 100, or 200 nematodes per larva. Larvae of both species were susceptible to nematode infections. However, neonate larvae of S. exigua were significantly less susceptible to nematode infection than 3- or 8-day-old larvae at or above 50 nematodes per larva. Mortalities of neonate larvae exposed to 50 or more nematodes ranged from 68 to 74% while mortalities of 3- and 8-day-old larvae ranged from 91 to 100%. The results with P. unipuncta showed similar trends as described for S. exigua, albeit at a lower mortality level and usually with no statistical differences. Mortalities of neonate larvae exposed to 50 or more nematodes ranged from 34 to 44% while mortalities of 7-day-old larvae ranged from 32 to 91%.