Inhibition of the accumulation of viral polypeptides in the densonucleosis virus-infected midgut of the silkworm,Bombyx mori,reared at a supraoptimal temperature

Effect of a supraoptimal temperature on the accumulation of viral polypeptides in the midgut was examined by immunoblot analysis in the larvae of the silkworm, Bombyx mori, infected with Bombyx densonucleosis virus type 2. In the larvae reared continuously at 25°C, viral polypeptides were first detected in the midgut at 2 days postinfection (pi) and in the feces at 4 days pi. When the larvae inoculated per os with the virus for 24 hr at 25°C were immediately shifted to 35°C, there were no detectable viral polypeptides in both the midgut and feces throughout the experiment. In the infected larvae shifted from 25° to 35°C at 48 hr pi, viral polypeptides preexisting in the midgut decreased to an undetectable level within 48 hr after the temperature shift, and no viral polypeptides were detected thereafter. Viral polypeptides in the feces of these larvae became detectable at 48 hr (4 days pi) after the temperature shift, as in the larvae at 25°C, and disappeared by 96 hr (6 days pi). These results indicate that a supraoptimal temperature inhibits accumulation of viral polypeptides in the midgut. It is likely that inhibited production of viral polypeptides rather than enhanced discharge of the infected midgut cells is responsible for the inhibited accumulation of viral polypeptides in the midgut at 35°C.     

Effect of alteration of endocrine mechanism on the development of nuclear polyhedrosis in the isolated pupal abdomens of the silkworm, Bombyx mori

Cyasterone injection into isolated pupal abdomens of Bombyx mori in an arrested state of development results in a hastened death of the abdomens from NPV. The time taken by the isolated abdomens to die was shortened even when cyasterone injection was delayed until 72 hr after NPV inoculation. When abdomens of pupae inoculated with NPV were isolated at different times after larval-pupal ecdysis, those prepared after the critical period of molting hormone secretion died sooner than those prepared before the critical period. These results imply that activated metabolism plays an active role in NPV development, possibly due to the increased rate of synthesis of viral constituents.     

Serial passage of Heliothis zea singly embedded nuclear polyhedrosis virus in a homologous cell line

This paper describes the replication and serial passage of Heliothis zea nuclear polyhedrosis virus (NPV) in a H. zea cell line. It was demonstrated that long-term serial passages of the H. zea NPV in homologous host cell culture decreased both the total number of polyhedral inclusion bodies (PIBs) produced and the infectivity of the supernatant as measured by TCID₅₀. The growth curve indicated that infectious material was released from cells 24 hr postinfection (p.i.) and approached a maximal titer 3 days p.i. The kinetics of H. zea NPV decay at 4°, 27°, and 37°C were determined. Infectivity was not detected after 3 weeks at 37°C, but approximately 10^3.5 TCID₅₀/ml activity was still present after 3 and 8 weeks storage at 27° and 4°C, respectively. Electron microscopy confirmed the presence of single embedded virions in the inoculated cells.     

Infectivity of the components of a nuclear polyhedrosis virus of the silkworm, Bombyx mori

The polyhedra of the nuclear polyhedrosis virus (NPV) of Bombyx mori were dissolved under controlled dissolution with weak alkali. Five components were separated by centrifugation of the dissolved solution in a sucrose gradient. The assay of the separated components in B. mori pupae showed that two types of rods were almost equally, highly infectious, whereas the small spherical particles were much less infectious than the rods by an order of 4 log units. The infectious unit of this virus appeared to be the virus rod without the outer membrane, and the small spherical particle did not represent any infectious functional unit but was a degradation product of the viral internal substance and the outer membrane may not be essential for the virus infection in the pupal hemocoel.     

Factors influencing the pathogenicity and development of Mattesia trogodermae infecting Trogoderma glabrum larvae

Factors that regulate development of Mattesia trogodermae in Trogoderma glabrum were defined, and their quantitative effects were determined. The rate of and the extent to which spore formation proceeds is strictly governed by temperature. More spores are produced at 30° than at 25°C and very low numbers of spores are formed when the incubation temperature is 35°C. When insects are incubated at 35°C for 1–10 days and transferred to 30°C for the remainder of the 30-day experiment, spore production capacity gradually declines with increasing time at 35°C. Two hypotheses are proposed for this phenomenon. Larval size also regulates the extent of spore production, larger larvae having greater potential for spore development. This is not influenced by dosage. Spore production in pupae and adults was always retarded.Dosage and environmental conditions which influence the virulence of M. trogodermae were investigated. These studies show that rates of mortality are higher at higher temperatures. Low doses of spores result in longer LT50's than do high doses at 25° and 30°C. No differences in rates of mortality were found between different doses at 35°C.     

Effects of photoperiod and temperature on diapause induction and termination in the swallowtail,Sericinus montelus

Abstract Sericinus montelus overwinters as diapausing pupae. In the present study, the effects of photoperiod and temperature on diapause induction and termination of diapause are investigated. The results obtained demonstrate that high temperature can reverse the effect of short day‐lengths on diapause induction. Under an LD 12 : 12 h photoperiod, all pupae enter diapause at 15, 20 and 25 °C, whereas all pupae develop without diapause at 35 °C. No pupae enter diapause under an LD 14 : 10 h photoperiod when the temperature is above 20 °C. Photoperiodic response curves obtained at 25 and 30 °C indicate that S. montelus is a long‐day species and the critical day‐length is approximately 13 h at 25 °C. At 25 °C, the duration of diapause is shortest when the diapausing pupae are maintained under an LD 16 : 8 h photoperiod and increases under LD 14 : 10 h and LD 12 : 12 h photoperiods. Under an LD 16 : 8 h photoperiod, the duration of diapause is shortest when the diapausing pupae are maintained at 25 °C, followed by 20 and 30 °C, and then at 15 °C. These results suggest that a moderate temperature favours diapause development under a diapause‐averting photoperiod in this species. The duration of diapause induced by an LD 12 : 12 h photoperiod is significantly longer at 25 °C than those at 15, 20 and 30 °C, and is shortest at 15 °C. At 25 °C, the duration of diapause induced by LD 6 : 18, LD 12 : 12 and LD 13 : 11 h photoperiods is similar and longer than 90 days. Thus, the diapause‐inducing conditions may affect diapause intensity and a photoperiod close to the critical day‐length has significant influence on diapause intensity in S. montelus.     

Functional differences between occluded and nonoccluded viruses of a nuclear polyhedrosis of the silkworm, Bombyx mori.

Comparative infectivity and virus neutralization studies on occluded and nonoccluded viruses of Bombyx mori nuclear polyhedrosis revealed that the infectious unit causing peroral infection differed from that causing hemocoelic infection. There were functional differences between the occluded (mainly virons with envelopes) and the nonoccluded virus (mainly virions without envelopes) preparations. The peroral infection was largely due to the virion with an envelope (peroral infectious unit), and the hemocoelic infection was due largely to the virion without an envelope (hemocoelic infectious unit). The apparent change of the virions with envelope to those without envelopes was detected as a slight increase in hemocoelic infectivity when the occluded virus was diluted and incubated at 4°C for more than 6 days.     

The effect of gypsy moth metamorphosis on the development of nuclear polyhedrosis virus infection

The development of nuclear polyhedrosis virus (NPV) infection in gypsy moth (Lymantria dispar) was studied before, during, and after host metamorphosis, and in larvae and pupae in the subsequent generation, to determine whether NPV ingested by late instars can replicate in host tissues through metamorphosis and whether it can be vertically transmitted to progeny. Individuals that survived sublethal dosages of NPV did not differ from undosed insects in pupal weight, fecundity, larval and pupal weight of progeny, or response of progeny to NPV challenge. No evidence of NPV infection or of abnormal histology was found in adult tissues examined by light microscopy and no virus was detected on the surface of eggs produced by NPV-treated moths. No NPV-caused mortality was recorded among undosed progeny of dosed or undosed parents. The progeny of dosed parents were neither more resistant nor more susceptible to LdMNPV than were progeny of undosed parents and lethal times did not differ between groups. Examination of larval, pupal, and adult tissues by DNA hybridization revealed that insects in which NPV DNA was detected died prior to adult eclosion. NPV was not detected in any hosts surviving to the adult stage. These results suggest that survivors of sublethal dosages of NPV avoid infection and are therefore incapable of vertically transmitting infectious virus to progeny.     

Susceptibility of Heliothis armiger to a commercial nuclear polyhedrosis virus

The susceptibility of Heliothis armiger larvae of different ages to a commercial nuclear polyhedrosis virus (NPV), Elcar, was determined by bioassay. The median lethal dosage (LD₅₀) increased 150-fold during the first week of larval life at 25°C, i.e., during development to early fourth instar, but daily feeding rate and thus potential virus acquisition also increased. A linear relationship was determined between log LD₅₀ and larval length, indicating that larval length constitutes a useful index for estimating the susceptibility of larval populations. Median lethal times (LT₅₀s) were similar for larvae tested at ages of 0 to 7 days and ranged from 3.6 to 8.0 days at 30°C. The amount of virus produced in a single, infected neonate was equivalent to 1.4 × 10⁶ LD₅₀s for neonates, a 900,000-fold increase on the dose supplied. The data support the practice of directing the NPV against neonates, but, on the basis of larval susceptibility alone, the age of larvae at treatment may not always be critical.     

Effect of dose of cytoplasmic polyhedrosis virus on infection,mortality, development rate,and larval and pupal weights of the pink bollworm

Host-pathogen relationships were studied between the pink bollworm, Pectinophora gossypiella, and a cytoplasmic polyhedrosis virus (CPV). Results showed that the median effective dose (ED₅₀), the dose that infects half the test subjects, was 1.91 × 10² polyhedral inclusion bodies (PIB)/ml of diet. The median lethal dose (LD₅₀) was 1.72 × 10⁵ PIB/ml. Diagnosis for CPV infection was more reliable in adult pink bollworms than in late-instar larvae. Duration of the larval stage increased with viral dose, but duration of the pupal stage was not affected by CPV. Weights of infected male and female pupae were 23.7 and 24.0% less than those of untreated pupae, respectively. Pupal weights were not significantly influenced by increases in the viral dose. Weights of larvae of a given age decreased as dose increased. The effect of CPV on duration of the immature stages and on pupal weight was not significantly influenced by rearing temperatures between 25.0° and 32.5°C. Pupal weight of infected pink bollworms decreased as the duration of the larval stage increased.     

Evaluation of the single radial diffusion technique for detection of nuclear polyhedrosis virus (NPV) infection in Heliothis zea

The single radial diffusion test is an effective method for detection of nuclear polyhedrosis virus infection in Heliothis zea larvae. Virus antigens were detected in some instances 48 hr after the larvae were exposed to virus. Most larvae tested positively for virus antigens 72 hr after exposure to the virus. The tests could be read within 4 hr if the incubation temperature was 35°C, and within 24 hr at 22°C.     

Ethanol production from pineapple juice in Côte d'Ivoire with preselected yeast strains

Fresh pineapple juice was inoculated with 8 preselected yeast strains and incubated for high yield ethanol production at between 25°C and 35°C.The natural pH of the juice did not affect the growth of the strains or the alcohol yields. The higher ethanol concentrations were obtained at 30°C and 35°C in the ranges of 27.17 to 46.50 g/l and 31.38 to 54.65 g/l, respectively, with 82% and 89% as the highest yields when referring to the theoretical yield.The study indicated that strain CMI is the best performing strain at 30°C and 35°C according to yield and productivity.     

An RNA virus in Autographa californica nuclear polyhedrosis virus preparations: Incidence and influence on baculovirus activity

A small RNA virus infectious to Trichoplusia ni larvae (TRV) was observed as a contaminant of several Autographa californica nuclear polyhedrosis virus preparations (AcMNPV). The extent of contamination in various AcMNPV preparations was studied by means of serial enrichment passages through T. ni larvae and enzyme-linked immunosorbent assay (ELISA). TRV could not be detected by ELISA in the original preparation of AcMNPV polyhedra prepared in 1968 even after five enrichment passages. Antibody inactivation offers a possible prophylactic method against TRV but temperature inactivation (55°C) does not. Although TRV reduced larval weight, it had little or no effect on bioassays of AcMNPV to T. ni and Heliothis virescens.     

Pathogenicity of a nuclear polyhedrosis virus of the almond moth, Cadra cautella

Tests were conducted with neonate Cadra cautella larvae to determine the pathogenicity of a nuclear polyhedrosis virus. A bioassay on an agar base diet showed that concentrations of 0.25, 0.50, 2.00, and 4.00 polyhedra/mm² killed 27, 55, 87, and 92% of the test larvae, respectively. A study of the time of death showed that most larvae died on the 9th or 10th day after exposure to 4 polyhedra/mm² at 27°C. When larvae were exposed to 8, 16, 32, and 64 × 10³ polyhedra/g of bran diet, recorded mortalities were 18, 22, 48, and 80%, respectively. All the samples of virus in bran diet which were incubated at various temperatures for 7, 14, and 28 days remained stable at all test conditions except the sample incubated at 42°C for 14 days, and those held at 37° and 42° for 28 days. Larvae of C. cautella, Plodia interpunctella, Ephestia elutella, and Paramyelois transitella placed on a diet with 40 × 10³ polyhedra/g had mortalities of 75, 59, 16, and 4%, respectively. Light and electron microscopical examination of P. interpunctella cadavers showed that they were infected with a multiply occluded nuclear polyhedrosis virus.     

Defect in translation of poliovirus RNA at the restrictive temperature.

Translation of the RNA of LSc type 1 poliovirus was examined in vivo at the restrictive temperature (39 °C). During the first two hours of infection at 39 °C the levels of viral polyribosomes were 50% lower than at 35 °C (permissive temperature). During the third hour of infection at 39 °C, only 4 to 10% of the control levels of polyribosomes were observed. Three experiments indicate that the elongation of viral peptides was not occurring properly at 39 °C. First, cultures incubated at 39 °C during the third hour of infection with both [³⁵S]methionine and [³H]uridine exhibit a fourfold increase in the ratio of viral protein/viral RNA in the polyribosome region of sucrose gradients in comparison to controls kept at 35 °C. However, at both temperatures the relative size distribution of polyribosomes was similar. Second, the ratios of released protein/nascent protein after 90-second and 5-minute pulses with [³⁵S]methionine indicate that elongation of peptide chains was inhibited at 39 °C. Third, when initiation of synthesis of viral protein was blocked with 150 mM-NaCl, the polyribosomes disaggregated four to five times more rapidly at 35 °C than at 39 °C. The data indicate that translation of viral RNA is inhibited at the restrictive temperature because of a reduced rate of elongation of viral proteins. The reduced rate of peptide chain elongation at 39 °C was fully reversible when cultures were shifted to 35 °C in the presence of 150 mm-NaCl. The latter finding indicates a conformational change in viral protein at 39 °C.     

The effect of the weight of the seedling-derived tuber on subsequent clonal generations in a potato breeding programme

Cultures of Polymyxa graminis were maintained in roots of barley plants grown in sand at different temperatures using Wisconsin soil temperature tanks. At 17 – 20°C, the minimum time from inoculation with cystosori to the production of zoospores from the inoculated roots was 2 – 3 wk. At 11 – 20°C many zoospores were produced but the incubation period was longer at the lower temperatures. Above 20°C little fungal development occurred. The duration of motility of zoospores ranged from c. 1 h to > 24 h. Bovine serum albumen (BSA) prolonged motility but glycine and glucose had no effect or, at higher concentrations, were toxic. Zoospores were rapidly immobilised by zinc ions in solution at or above 10μg/ml. In some experiments BSA added to the zoospore suspension greatly increased transmission of barley yellow mosaic virus (BaYMV) while glucose, glycine and ovalbumen decreased it. When seedlings were incubated with zoospore suspensions for 24 h at different temperatures, BaYMV transmission was high (> 60%) at 10, 15 and 20°C but there was little at 5 or 25°C. In experiments to determine the time taken for zoospore penetration, seedlings were incubated in suspension for different periods of time and then rinsed in zinc sulphate solution to kill free zoospores. Between 3 and 3·5 h was needed for zoospores to establish infection. Transmission occurred equally to plants of various ages between 3 days and 7·5 wk.     

Nuclear polyhedrosis virus infection ofPseudoplusia includens [Lep.: Noctuidae] larvae: Effect on post larval stages and transmission

Treatment ofPseudoplusia includens (Walker) larvae in either the 4th, 5th or 6th stage with a nuclear polyhedrosis virus (NPV) did not affect the weight or sex ratio of surviving pupae. Pupal mortality was low but increased with the dosage of the NPV larvae had received. Also, mortality of pupae from larvae treated in the 5th or 6th stage was higher than from larvae treated in the 4th stage. Adult longevity was not affected by the virus. Fecundity and egg viability were reduced following most treatments of 6th stage larvae but not 4th or 5th stage larvae. Adults exposed to NPV at 3 dosages during the 6th larval stage were released in cages on soybean. Some of the progeny succumbed to NPV infection. Progeny mortality increased over a 7 week period of adult release, but did not differ significantly between treated and untreated groups by the end of the 7 week period.     

Quantitative analysis of the metabolism in the isolated pupal abdomens of the silkworm, Bombyx mori, infected with nuclear polyhedrosis virus

The rate of oxygen uptake and the amount of several main cell constituents were determined in NPV-infected and uninfected isolated pupal abdomens of the silkworm, Bombyx mori. The oxygen uptake in uninfected isolated abdomens increases for the first 2 days due to the effect of water injection, but thereafter it remains almost unchanged at a relatively low level. When isolated abdomens are infected with an NPV, the oxygen uptake increases markedly from 3 days postinfection onward, being about three times that of uninfected isolated abdomens at late stages of infection. The amounts of cell constituents analyzed in the present study change little in uninfected isolated abdomens throughout the experiment. Infection of NPV leads to a marked accumulation of both DNA and RNA, and a loss of glycogen. The amount of protein, lipid, and sugar are scarcely affected by NPV infection. These results indicate that the activity of metabolism in uninfected isolated abdomens is not only low but also stable, and that infection of NPV results in an activation of host cell metabolism. It is considered that such isolated pupal abdomens provide an excellent opportunity to analyze both the process of NPV replication and the alteration of host cell metabolism resulting from NPV infection.     

Persistent Baculovirus Infection Results from Deletion of the Apoptotic Suppressor Gene p35

Infection with the wild-type baculovirus Autographa californica multiple nuclear polyhedrosis virus (AcMNPV) results in complete death of Spodoptera frugiperda (Sf) cells. However, infection of Sf cells with AcMNPV carrying a mutation or deletion of the apoptotic suppressor gene p35 allowed the cloning of surviving Sf cells that harbored persistent viral genomes. Persistent infection established with the virus with p35 mutated or deleted was blocked by stable transfection of p35 in the host genome or by insertion of the inhibitor of apoptosis (iap) gene into the viral genome. These artificially established persistently virus-infected cells became resistant to subsequent viral challenge, and some of the cell lines carried large quantities of viral DNA capable of early gene expression. Continuous release of viral progenies was evident in some of the persistently virus-infected cells, and transfection of p35 further stimulated viral activation of the persistent cells, including the reactivation of viruses in those cell lines without original continuous virus release. These results have demonstrated the successful establishment of persistent baculovirus infections under laboratory conditions and that their establishment may provide a novel continuous, nonlytic baculovirus expression system in the future.     

The effect of temperature on feeding,growth, and metabolism during the last larval stadium of the female house cricket, Acheta domesticus

Eighth instar female house crickets at 35°C developed faster, gained slightly more wet weight, and consumed less food, water, and oxygen than at 25°C. The duration of the 8th stadium at 25°C was 13 days (undisturbed), but was 14 days when disturbed by daily weighing. The duration of the 8th stadium at 30°C was 8 days and at 35°C was 6 days. During the first half of the 8th stadium at 25, 30, and 35°C, there was a high rate of food and water consumption resulting in statistically equal maximum dry weight achievement (124 mg). Respiratory quotients greater than one during this time indicated the conversion of ingested carbohydrate to fat. During the latter half of the 8th stadium, food and water consumption declined and the crickets lost weight. The period of weight loss was proportionally much longer at 25°C than at 30 or 35°C. Respiratory quotients lower than 1.0 during the latter half of the 8th stadium at 30 and 35°C indicated the metabolism of stored lipids. The respiratory quotient at 25°C never fell below 1.0, possibly because some food remained in the gut. The absorption efficiency was not influenced by temperature (25–35°C). Though the caloric content of the faeces was lower at 25°C than at 30 or 35°C, which correlated to the much longer time for food passage at 25°C than at 35°C, the difference in total calories egested was insufficient to alter the absorption efficiency. A longer period of reduced feeding and greater dry weight loss during the latter half of the 8th stadium at 25°C resulted in a lower metabolic efficiency at 25°C than at 30 or 35°C. Eighth instar crickets in response to a step-function transfer from 30°C–25 or 35°C showed an immediate (<1 hr) and complete metabolic adjustment which was not affected by the temperature history during the 7th stadium. House crickets did not exhibit temperature acclimation in the range 20–40°C, the metabolic rate being determined by ambient temperature. The Q₁₀ for oxygen consumption in the range 20–40°C was about 2.