In vitro Cultivation of the Entomogenous Nematode Filipjevimermis leipsandra

The mermithid nematode, Filipjevimermis leipsandra, was successfully cultivated to the preadult stage in Schneider''s Drosophila medium supplemented with 20% fetal bovine serum. Upon transfer to a solid substrate the preadults continued to develop into ovipositing adult females. Four molts were observed. The first molt occurred in the egg. The second occurred after 6-8 days in culture during which the very thin cuticle was shed completely. The third molt occurred after 18-20 days in culture; the cuticle was retained by the third-stage nematode. This stage was considered comparable to the preadult stage that emerges from host larva, Diabrotica spp. The fourth molt occurred within 12 days after the preadult was transferred from the liquid medium to a solid substrate. Adult females began ovipositing viable eggs 1-3 days after the final molt.     

Phocanema decipiens: Pathology in seals

Infective larvae of the anisakine nematode, Phocanema decipiens from cod (Gadus morhua), were fed to harbor seals (Phoca vitulina) and gray seals (Halichoerus grypus). At termination of the experimental infections, 6 of 8 harbor seals were infected with 34(6–64) P. decipiens, while the remaining 2 seals had relatively heavy infections of 250 and 547 nematodes, respectively; 10 of 11 gray seals were infected with 128 (68–229) P. decipiens, but only 2 nematodes were recovered from the 11th seal. Larvae and adults of P. decipiens occurred throughout the gastrointestinal tract, but mainly in the fundic portion of the stomach; the anterior extremities of the nematodes were embedded in mucosa and submucosa. Clusters of adult P. decipiens were associated with ulcerous gastric lesions in harbor seals and raised inflammatory areas in stomachs of gray seals. Singly occurring larvae from challenge transmissions were associated with raised inflammatory areas in the stomachs of both host species. Histological examinations revealed that the lesions and inflammatory areas were eosinophilic granulomata. Anisakis sp. larvae from the viscera of cod were also fed to 1 of the gray seals. Eighty of these larvae were subsequently found in association with a general inflammation in the cardiac area of the stomach in this seal. Natural anisakine infection were surveyed in 16 harbor and 53 gray seals from the Nova Scotia mainland. The natural incidence of P. decipiens was 62(5–177) in harbor seals and 577(11–1694) in gray seals. Clusters of adult P. decipiens were found in association with gastric lesions in 2 juvenile harbor seals; however, in gray seals, the nematodes neither occurred in clusters nor in association with gastric lesions.     

Phocanema decipiens: Growth,reproduction, and survival in seals

Captive harbor seals (Phoca vitulina) and gray seals (Halichoerus grypus) were fed infective larvae of Phocanema decipiens, an anisakine nematode from the flesh of Atlantic cod (Gadus morhus). Ova of P. decipiens were first detected in the feces of harbor seals 21(17–30) days after exposure; the patency period was 15 to 45 days. In gray seals, the prepatent period was 19(16–23) days; patency 20–60 days. By the sixth week of infection in harbor seals, mean body lengths of adult females and males of P. decipiens were 60.8(40.8–76.2) and 54.3(45.5–60.8) mm, respectively; mean fecundity of female nematodes was 156,000 ova. In infections of similar duration in gray seals, females and males of P. decipiens were 82.1(69.7–104.3) and 64.4(53.8–72.7) mm in length, respectively; mean fecundity of females was 366,000 ova. In sensitizing infections in harbor seals, 28% of P. decipiens survived to early patency (Days 25–30) while only 9% of the nematodes survived to midpatency (Days 35–45). In sensitizing infections in gray seals, 56% of P. decipiens survived to early patency (Days 20–30) and 48% survived to midpatency (Days 35–50). Seals with existing or recent P. decipiens infections resisted reinfection; <50% of the nematodes in challenge infections in gray seals survived to Day 3 and <10% survived to patency. Growth of the nematodes, however, was not retarded in the challenge infections and resistence to reinfection subsided when seals were maintained anisakinefree for 2–6 months after loss of prior natural or experimental infections. Natural anisakine infections were surveyed in 16 harbor and 53 gray seals from the Nova Scotia mainland. The mean incidence of P. decipiens was 62(5–177) in harbor seals and 577(11–1694) in gray seals; incidence varied seasonally and with age of host. Adult females of P. decipiens from harbor seals were 64.0(49.2–79.8) mm in length and contained 1.68(0.87–2.73) × 10⁵ ova; females from gray seals were 78.3(62.3–92.1) mm in length and contained 2.39(0.69–4.39) × 10⁵ ova.     

Embryology and Life Cycle of Tylenchorhynchus claytoni Steiner, 1937 (Nematoda: Tylenchoidea)

Development of Tylenchorhynchus claytoni from unsegmented egg to hatching takes 135 hr at 22-25 C. The fourth molt lasts 5 to 6 days. During exsheathment the cast cuticle of the larva separated into two unequal parts, breaking near either the anterior or posterior end. The life cycle from egg to egg required from 31 to 38 days at 28 C on alfalfa seedlings and included four molts and four larval stages. Sexual differentiation was apparent in third-stage larvae.     

A Case of Anisakidosis Caused by Pseudoterranova decipiens Larva

Pseudoterranova decipiens larva is a rare cause of anisakiasis. Indeed, prior to the present study, there had been only 12 reports of larval P. decipiens infection in the Republic of Korea. In June 2011, an anisakid larva, 32.1 mm in length and 0.88 mm in width, and finally identified as the third stage larva of P. decipiens owing to the presence of an intestinal cecum but lacking ventricular appendage, was discovered in a 61-year-old woman during the course of endoscopy executed as a part of routine physical examinations. The patient had eaten raw a rockfish 13 hr prior to the endoscopy, but showed no symptoms of anisakiasis. This paper is the 13th report of P. decipiens infection in Korea.     

Reproduction,Chromosome Number,and Postembryonic Development of Panagrellus redivivus (Nematoda: Cephalobidae)

Panagrellus redivivus (L.) T. Goodey reproduced amphimictically; the sexual primordia of males had nine chromosomes, those of females had ten. Eggs contained five chromosomes, sperm four or five. There were four molts, all after hatching. The sexes could be separated at the second molt by development of a lobe of somatic cells in the gonad, anteriorly in males, posteriorly in females. The lobe in males reflexed posteriorly at the third molt and joined the rectum at the fourth molt. Third molt females had a thickened vaginal primordium and at the fourth molt the spermathecal and uterine primordia were evident. The uterus elongated enormously in the adult. The 15 ventral chord nuclei between esophagus and rectum in the first stage increased to approximately 63 during the first molt; specialized nuclei, not evident until the third molt, participate in vaginal lining formation in fourth molt females. Sperm were first produced at the late fourth molt. Eggs, not produced until after copulation, hatched within the uterus.     

Brain-independent development in the moth Sesamia nonagrioides

The caterpillars of Sesamia nonagrioides developing under long-day (LD) photoperiod pupate in the 5th or 6th instar whereas under short day (SD) conditions they enter diapause and undergo several extra larval molts. The diapause is terminated within 1-3 instars upon transfer of SD larvae to the LD conditions. Brain removal from the 6th instar larvae promotes pupation followed by imaginal development; however, one third of the SD larvae and 12% of the LD larvae debrained at the start of the instar first undergo 1-2 larval molts. The incidence of larval molts is enhanced by the brain implants. Exclusively pupal molts occur in the LD larvae debrained late in the 6th instar. Decapitation elicits pupation in both LD and SD larvae, except for some of the 4th and 5th and rarely 6th instar that are induced to a fast larval molt. The pupation of decapitated larvae is reverted to a larval molt by application of a juvenile hormone (JH) agonist. No molts occur in abdomens isolated from the head and thorax prior to the wandering stage. Abdomens isolated later undergo a larval (SD insects) or a pupal (LD insects) molt. Taken together the data reveal that in S. nonagrioides (1) several larval molts followed by a pupal and imaginal molt can occur without brain; (2) an unknown head factor outside the brain is needed for the pupal-adult molt; (3) brain exerts both stimulatory and inhibitory effect on the corpora allata (CA); (4) larval molts induced in CA absence suggest considerable JH persistence.     

Human infection by Pseudoterranova decipiens (Nematoda, Anisakidae) in Chile: report of seven cases

From 1997 to 1999, we identified seven human cases of infection by fourth stage larvae of Pseudoterranova decipiens in Chile. All identified larvae were coughed up by the patients. Subjects were 10-55 years old; five were female. Some patients complained of coughing, expectoration, pharyngeal pain, nausea or anal and nasal pruritus. Larvae of three patients were coughed up from 36 h to 7 days after having eaten raw (cebiche or sushi) or lightly fried fish. P. decipiens has a marine life cycle. Infective third stage larva develop to adult stage in pinniped mammals. The nematode eggs are voided with the host faeces and develop and hatch releasing third stage larvae. Some crustaceans and fish act as hosts of third stage larvae. Man is an accidental host for third or fourth stage larvae.     

Analysis of pattern formation in Neurospora perithecial development using genetic mosaics.

The growth and reproduction of the small nematode Caenorhabditis elegans has been studied using an electronic nematode counter recently developed in our laboratory. At 20°C, the usual growth temperature, size increases in a smooth sigmoidal manner with time, linear growth being most rapid around the time of the fourth molt and nearly ceasing by the end of the period of egg-laying. Growth of populations is highly synchronous; the small residual size heterogeneity is maximal at about the time of maximal growth. The four molts do not involve major interruption of growth, but they do entail slight shape changes (elongating upon escape from the old cuticle). Egg-laying begins shortly after the fourth molt, the rate rising rapidly at first, then more gradually to a peak followed by a relatively rapid fall. Comparable measurements at 16 and 25°C establish that these are acceptable limit temperatures for work with temperature-sensitive mutants, although egg yield is somewhat reduced and the kinetics of egg-laying are altered at 25°C. Developmental chronologies for all three temperatures are presented.     

Life Cycle and Mating Behavior of Belonolaimus longicaudatus in Gnotobiotic Culture

The life cycle of Belonolaimus longicaudatus was observed in vitro on excised roots of Zea mays. Roots were cultured on Gamborg''s B5 medium in petri dishes with 1.5% agar adjusted to pH 5.8 and incubated at 28 °C in darkness. Second-stage juveniles (J2) fed on the roots and started the second molt (M2) to the third-stage juveniles 2 days after inoculation (DAI). The third molt (M3) to the fourth-stage juveniles occurred 7 DAI, followed by the fourth molt (M4) to males 13 DAI or to females 14 DAI. Nematode gender differences were observed by the end of the fourth molt. The first male appeared 15 DAI and the first female 17 DAI, after which mating occurred. Males were attracted to females, and mating was observed. Mating was required for reproduction. Fertilized females began to lay eggs 19 DAI and continued egg laying without the further presence of males during a 90-day observation. All of the eggs hatched. Unfertilized females rarely laid eggs, and none of the eggs were able to hatch. Feeding took place between each molt and before egg deposition occurred. The first-stage juveniles molted in the eggs 4 days after deposition, and J2 hatched from eggs 5 days after egg deposition. The life cycle from J2 to J2 was completed in 24 days.     

Anguina plantaginis n. sp. Parasitic on Plantago aristata with a Description of Its Developmental Stages

Anguina plantaginis n. sp., parasitic on Plantago aristata, is described and illustrated. This new species is most closely related to A. klebahni, A. millefolii, A. mobilis, and A. moxae and is characterized as follows: moderate body size for the genus; absence of esophageal "storage organ"; postvulval uterine sac extending about 45% of vulva-anus distance; crustaformeria of young females longer than spermatotheca or uterus proper; spicules with 2 sclerotized thickenings; long, conical tail in both sexes, narrowing at about 1/6 of its length to peg-like tip; parasitic only on P. aristata. Two nematode generations that are morphologically similar but differ in body size develop in one plant gall. The postembryogenesis, studied with respect to morphological development of the larval stages, is similar to that of Ditylenchus. The sexes can be differentiated from the second molt on. The infective larva is the third stage, which is morphologically distinct from the regularly developing third-stage larva.     

The influence of intrinsic and extrinsic factors on developmental parameters and their relationships in the marine isopod Idotea linearis (Crustacea)

Developmental and reproductive parameters and their relationships were studied in the marine isopod Idotea linearis. We hypothesized that (1) the temporal patterns of molting and growth undergo complex and sex-specific changes with age as well as with the onset of sexual maturation, and that (2) sexual maturation (and dependent parameters) is controlled by the photoperiod. Both males and females were singly cultured in the laboratory at two alternative photoperiods (constant long and short days, respectively) from hatching until death. Males molted and grew throughout their life, showing a steady increase in stage duration and body size with each molt. Females, in contrast, showed much more complex modifications in molt chronology due to reproductive demands. There was some variability in the stage number, when females reached maturity. Reaching maturity early in the succession of molts was associated with smaller body size at maturity, smaller size of broods, but higher average number of broods per lifetime. Post-puberty molts in females occurred without further growth, and successive broods did not differ in size. The photoperiod strongly affected sexual maturation (and thus in turn molting and growth patterns) in females, while males remained completely unaffected by the photo regime.     

High Laccase2 expression is likely involved in the formation of specific cuticular structures during soldier differentiation of the termite Reticulitermes speratus

Termite soldiers are morphologically specialized for colony defense. Analysis of the mechanisms of soldier differentiation is important for understanding the establishment of termite societies. Soldiers differentiate from workers through a presoldier stage and have well-sclerotized and pigmented cuticles. These characteristics are important for nest defense and are likely to be caused by soldier-specific mechanisms of cuticular tanning. The molecular mechanisms leading to cuticular tanning have not been elucidated. Laccase2 (Lac2) plays important roles in this process in insects, and we hypothesized that Lac2 expression may be involved in soldier-specific cuticular tanning. We observed inner and outer head cuticle changes and compared the Lac2 expression patterns among three molts (worker–worker, worker–presoldier and presoldier–soldier) in the termite Reticulitermes speratus. Quantitative analyses of head cuticle colors showed that the color properties changed more conspicuously in presoldier–soldier molts than in the other two molts. Histological observations showed that the exocuticles of soldier heads were substantially thicker than those of worker and presoldier heads, underwent tanning before or just after ecdysis, and were pigmented at earlier time points than other molts. Finally, markedly higher Lac2 expression levels were observed just before and after ecdysis only in the presoldier–soldier molt. These results suggest that specific cuticular formation occurs in the exocuticles during soldier differentiation, and that the high level of Lac2 expression during the presoldier–soldier molt is related to soldier-specific cuticular tanning. We speculate that evolution of the regulatory mechanisms of Lac2 expression were important for the acquisition of soldier-specific cuticles.     

Biology of Octomyomermis muspratti,a parasite of mosquitoes as it relates to mass production

The first- and second-instar larvae of Culex pipiens quinquefasciatus were equally susceptible to parasitism by the mermithid nematode Octomyomermis muspratti; the third-instar hosts were significantly less susceptible. The yield of postparasitic nematodes was higher from hosts exposed as second instars then from hosts exposed as first or third instars at comparable levels of parasitism. Mortality of the host prior to emergence of the nematode occurred most frequently among first- and third-instar hosts. About 4% of the infected hosts exposed as first and second instars and 37% of the infected hosts exposed as third instars pupated prior to emergence of the postparasitic nematodes. The exposure of mosquitoes as second instars was the most efficient for mass rearing and resulted in the highest yields of postparasitic O. muspratti.     

MOLTING PHENOLOGY OF HARBOR SEALS ON TUGIDAK ISLAND, ALASKA

We documented the progression and timing of the annual molt of harbor seals on Tugidak Island, Alaska, from 1997 to 1999. In all years the timing of molting differed among age-sex classes. Yearlings molted first, subadults second, adult females third, and lastly adult males. Timing of molting was nearly identical in 1997–1998, whereas in 1999 molting occurred three to six days later for all age-sex classes except yearlings. Estimated dates when peak proportions of each age-sex class were molting ranged from 2 August (yearlings) to 2 September (adult males). The number of seals hauled out was positively related to the proportion of seals in the molt and negatively related to the proportion of seals in the postmolt. Population trend estimates, based on aerial counts conducted during a narrow window within the molting period, are likely biased toward certain age-sex classes. Statistical models used to estimate trend include covariates to help account for within-year variation in seal numbers, but do not account for compositional changes that occur during molting. Population modeling may elucidate the effects of within-year population structure on trend estimates. Monitoring molting phenology at additional sites is necessary to determine the extent of geographic variation in molting.     

Host age and pathogen dosage impact cyst morphogenesis in the invertebrate pathogenic alga Helicosporidium sp. (Chlorophyta: Trebouxiophyceae)

Helicosporidium sp. is a pathogenic alga that replicates in the hemolymph of various invertebrate hosts. Morphogenesis of the infectious life stage, the cyst, occurs in the infected host, but to date cannot be induced in vitro. Using larvae of the heterologous host Helicoverpa zea, we examined potential factors influencing pathogenicity and in vivo cyst production of the alga and the impact of infection on host survival. Factors tested were cyst dosage administered per os (ranging from 10² to 10⁵ cysts per larva) and host age at exposure (third, fourth, and fifth larval instar). Cyst production occurred between 7 and 13 days after treatment, regardless of host age at treatment. Increasing dosage increased both percent infection and mortality, but cyst production did not track the total infection response. Increasing host age at exposure mitigated dosage effects on infection and mortality and also elevated cyst production in later-treated larvae. Only the highest dosage produced a significant decrease in the overall time to death. Moderate cyst dosages and later host ages were most effective at regenerating Helicosporidium cysts.     

Regulation ofHelicoverpa zea larval behavior by the parasitoidEucelatoria bryani

The parasitoidEucelatoria bryani Sabrosky regulates the larval behavior of its hostHelicoverpa zea (Boddie). Parasitized third, fourth and fifth instars burrow into the soil 0.7–3.4 days earlier than unparasitized larvae that normally enter the soil to pupate at the end of the fifth and final larval instar. Parasitized third instars molt once then burrow as fourth instars, one instar earlier than normal. WhenE. bryani pupariated on the soil surface in the field, none survived to the adult stage. However,E. bryani adults emerged from 49.2% of hosts that had burrowed into the soil. By accelerating the timing ofH. zea burrowing behavior and causing host larvae to enter the soil before death,E. bryani ensures its pupariation in an environment with improved protection against natural enemies and lethal temperatures.     

Effect of Host Plant on the Infectivity of Sl MNPV to Spodoptera litura (Fabricius) (Lepidoptera: Noctuidae) Larvae

The susceptibility of Spodoptera litura to SlMNPV infection was markedly affected by phyto-chemicals ingested during the acquisition of viral inoculum on foliage of tomato and cauliflower. The LD₅₀ values computed for second, third and fourth instar larvae assayed on tomato leaves were 254, 819 and 23395 PIBs/larva, respectively whereas, it was 326, 1719 and 43843 PIBs/larva for respective instars when assayed on cauliflower leaves. Thus LD₅₀ values for second, third and fourth instar larvae were 1.28-, 2.09- and 1.87- fold lower, respectively in tomato leaves. Similarly, LT₅₀ values for second, third and fourth instar larvae assayed on tomato leaves were 7.1 and 7.5 days, respectively at inoculum dose of 2.7×10⁴ PIBs/larva whereas, it was 7.7 and 8.0 days for respective instars when assayed on cauliflower leaves at same inoculum. This result also showed that the S. litura were more susceptible on tomato leaves in comparison to cauliflower leaves as the time required for mortality was lower in tomato leaves. The possible biochemical bases for differential level of mortality of S. litura larvae on tomato and cauliflower crops needs to be investigated.     

Effects of Host Diet on Romanomermis culicivorax,a Mermithid Parasite of Mosquitoes

When larval mosquitoes (Aedes aegypti) infected with the mermithid nematode Romanomermis culicivorax were fed on a diet low in quantity or protein content or both, the number of postparasites which emerged from the hosts decreased and host mortality increased marginally. Parasitic development was prolonged and became asynchronous in nutritionally deprived hosts. Nematodes emerged from insects infected by more than one nematode before the remaining juveniles comprising such infections had completed parasitic development; this resulted in substantial reductions in postparasite nttmbers. Host development was retarded by low protein and/or reduced diets. Postparasites emerging from second and third instars were reduced in size and in the amount of stored nutriment compared to those recovered from hosts fed on a high protein diet ad libitum. A greater proportion of the rnermithids developed into males in hosts fed on reduced diets but not in hosts fed on low protein diets.