Temperature effects during development on a polyembryonic parasitoid and its host

Ambient temperature can influence development through effects on metabolic rate and by inducing physiological stress. In this study, we assessed temperature effects on a host–parasitoid interaction and on the body size and brood size of emerging wasps. By examining the development at two different temperatures of the koinobiont parasitoid, Copidosoma bakeri, and its host, Agrotis ipsilon, we asked: (1) Does the growth response to temperature by A. ipsilon depend on whether the moth caterpillar is parasitized? (2) Does the allocation pattern of body size and brood size in C. bakeri change with temperature? To answer these questions, we exposed A. ipsilon larvae parasitized by C. bakeri to high or low non-lethal temperatures when A. ipsilon was in early or late larval stages and measured their development time and body mass for all four treatment combinations. We also examined the brood size and body mass of emerging wasps. Whether parasitized or not, A. ipsilon larvae decreased development time, but generally did not decrease final body mass, at the higher temperature. When parasitized A. ipsilon was exposed to the higher temperature only late in their development, enlargement of the host by the parasitoid was reduced. C. bakeri brood size significantly increased when the higher temperature was applied early in host development. We did not detect a shift with temperature in the allocation pattern of the size–number trade-off for wasp offspring, suggesting that this trade-off relationship may be under selection strong enough to yield insensitivity to temperature.     

Cryptosporidium parvum: the course of Cryptosporidium parvum infection in C57BL/6 mice co-infected with the nematode Heligmosomoides bakeri

The effects of Heligmosomoides bakeri infection on the course of a concurrent Cryptosporidium parvum infection were studied in C57BL/6 mice. Mice were initially infected with 80 L₃ of H. bakeri and then challenged with 10⁴ oocysts of C. parvum, administered during the patent period of the nematode infection (28 day post H. bakeri infection). The number of C. parvum oocysts excreted in the feces and the number of adult H. bakeri in the small intestine were monitored during the experiment. Concurrent H. bakeri infection resulted in a prolonged course of infection with C. parvum. The intensities of both parasite infections were higher in co-infections. We also investigated the cellular immune response at 14 and 42 days post infection C. parvum. During infection with C. parvum there was an increase in production of IFN-γ and IL-12 but co-infection with H. bakeri inhibited IFN-γ secretion. The present study is the first to demonstrate that infection with H. bakeri markedly exacerbates the intensity of a concurrent C. parvum infection in laboratory mice and also affects immune effectors mechanisms in co-infection with H. bakeri.     

Biological parameters of the predatory mite Cheletomimus bakeri (Acari: Cheyletidae) feeding on Tetranychus cinnabarinus (Acari: Tetranychidae)

Developmental periods of egg, larva and nymphal stages and fecundity as well as predation of Cheletomimus bakeri (Acari: Cheyletidae) feeding on Tetranychus cinnabarinus (Acari: Tetranychidae) were evaluated at different temperatures (15, 20, 25, 30 and 35°C) at 65 ± 10% relative humidity and 16 L: 8D in the laboratory. The development periods of C. bakeri from egg through adult decreased significantly when the temperature was increased from 20°C to 35°C. Egg and total development periods of C. bakeri at 20, 25, 30 and 35°C were 13.86, 7.98, 5.07, 4.08 days and 58.66, 41.51, 21.21, 22.92 days, respectively. The highest numbers of total and daily egg production were found at 20°C and 30°C, respectively. Net reproductive rate (R₀ = 13.29), mean generation time (T = 88.30), gross reproductive rate (GRR = 17.46) and doubling time (DT = 23.66) were the highest at 20°C. The intrinsic rate of increase (r_m = 0.0592) and finite capacity for increase (λ = 1.061) for C. bakeri were the highest at 30°C. Predation of C. bakeri increased throughout the range of prey densities. The highest consumption number of C. bakeri feeding on T. cinnabarinus males per day was 4.63, 4.70 and 4.60 when confined to 40, 80 and 160 individuals, respectively. Our data suggest that C. bakeri does not appear to have much promise for the control of spider mites because of the characteristics of the predator such as slow development period, poor searching capacity and low intrinsic rate of increase.     

The Effects of Hot Water Treatments on Survival of Heterodera schachtii

Cysts of Heterodera schachtii were treated in a water bath at constant temperatures ranging from 45 - 62.5 C for 1 sec to 28 hr. Treated and untreated cysts were incubated 8 weeks in sugarbeet root diffusate at 24 C to measure emergence of surviving larvae. Within the temperature range of 49 - 54 C, the minimum lethal temperature was proportional to the log time of treatment. No larvae emerged from cysts exposed 10 sec at 60 C. Although treatment of cysts for 8 hr at 45 C significantly reduced emergence, increasing the treatment period to 28 hr did not completely suppress emergence.     

Gene flow between an endangered endemic iguana, and its wide spread relative, on the island of Utila, Honduras: when is hybridization a threat?

The island endemic Ctenosaura bakeri was listed as critically endangered by the IUCN Redlist Assessment in 2004, 7 years after it was recognized as a distinct species. C. bakeri occupies a portion of Utila, a small continental island located off the northern coast of Honduras. Habitat destruction and over-harvesting are among the top threats facing this species. In addition, morphological evidence of hybridization was recently documented, raising the concern that gene flow from the common and widely distributed C. similis could threaten the genetic distinctiveness of C. bakeri. We show that hybridization occurs only at low levels and is not a current threat to C. bakeri. All ctenosaurs captured for this study were identified to species level without difficulty; none had intermediate or mosaic phenotypes. Sequence analysis of mitochondrial and nuclear markers revealed only two individuals with introgressed genotypes. Molecular analysis of the previously described hybrid showed it to be heterozygous for C. bakeri and C. similis alleles. Hybridization between these two species is possible and occurs occasionally in the wild, and the rate of hybridization could increase if habitat destruction or changes in relative abundance increase the probability of interbreeding. However, the level of gene flow indicated by current data is too low to threaten C. bakeri with genetic swamping or deleterious fitness effects.     

Effect of temperature and photoperiod on the induction of diapause in the mite Metaseiulus occidentalis

Diapause in a predaceous mite, Metaseiulus occidentalis, from a Californian vineyard population is a photoperiodically induced, facultative, adult reproductive diapause in females. The laboratory-determined critical photophase at 19°C was estimated at 11·2 hr. At 16°C, the critical photophase under laboratory conditions was approximately 11·6 hr. Temperature influenced the photoresponse of M. occidentalis so that diapause was entirely averted at temperatures of 22, 25, and 30°C. Aestival diapause at higher temperatures and long photophases was lacking. Development was continuous under constant darkness at all the temperatures tested. Diapause termination in laboratory-reared mites occurred spontaneously under the inductive conditions. Under constant 19°C temperatures, females responded to photophases so that diapause was terminated most rapidly under a 16 hr photophase (in 18·6 days); the 12 and 8 hr photophases, at this temperature, were next in their effectiveness, with 27·9 and 73·0 days, respectively, required for termination.     

Studies on the Host Range of Xiphinema bakeri and Its Pathogenicity to Raspberry

Thirty-one kinds of plants representing 12 families were tested for host suitability to Xiphinema bakeri. Sixteen supported a significant population increase but only members of the Rosaceae and Solanaceae were severely damaged. Eight of the 12 weed species tested were good hosts; Mouse-ear chickweed allowed the greatest population increase of all plants tested. Populations of X. bakeri declined under selected members of the Cruciferae and Cucurbitaceae more than in fallow soil after 12 weeks. Numbers of X. bakeri as low as one per 5 cc of soil reduced root and top growth of raspberry 40-50%. Where 100 and 500 nematodes per 10.5 cm-diameter pot were used the mean weight of roots was reduced 54% and 77%, the tops 59% and 78% and the linear growth 48% and 78%, respectively. This is the first report of an ectoparasitic nematode pathogenic to raspberry.     

Patterns of primary spore discharge of Entomophthora spp from the blue green aphid,Acyrthosiphon kondoi

Experiments were conducted to study the effects of time, temperature, and light regime on primary spore formation at 100% RH for the three major pathogens of Acyrothosiphon kondoi. Only small differences were detected between the continuous light and continuous dark regimes. Entomophthora obscura produced between 6 and 10 × 10³ primary spores mostly during the first 48 hr. Total primary spore production was similar at the five temperatures tested from 5° to 25°C. Entomophthora planchoniana produced large numbers of primary spores (about 5 × 10⁴ per aphid) only at temperatures between 10° and 20°C. The majority of primary spores were formed during the first 24 hr. Primary spore production with Entomophthora nr. exitialis ranged from about 10⁵ per aphid at 5° and 10°C to 3 or 4 × 10⁵ at 15° to 25°C, with most spores being formed during the first 48 hr. It is suggested that rainfall is more likely to be important for transmission of E. obscura and E. nr. exitialis than for transmission of E. planchoniana, and that E. obscura is likely to be the most important pathogen during cool or cold weather.     

Neoaplectana carpocapsae: Movements of nematode populations on a thermal gradient

To gain information on factors which could affect the nematode Neoaplectana carpocapsae's, dispersion and infection of insect larvae in the field, nematode populations were tested on a thermal gradient (0.5 C/cm). Infective juveniles grown at 15, 20, and 25 C migrated on the gradient toward their respective growth temperatures when tested immediately after harvesting from their medium. This migration by juveniles, grown at 20 or 25 C, was altered within 12 hr by shifting incubation temperature. The nematodes' tendency to migrate toward warmer temperatures, when placed below incubation temperature, decreased during 6–7 days incubation at 20 or 25 C and the nematodes reversed direction: incubation at 2–5 C inhibited this reversal. Nematodes incubated at 20 C, then applied to a gradient zone at 12–13 C, had a greater tendency (P < 0.05) to remain aggregated in that zone within a 3-hr period than those applied at 17–18, 22–23, or 27–28 C. Their tendency to migrate from the 12–13 C zone was significantly (P < 0.05) increased by shifting them to 15 C for 14–138 hr.     

Two new species and new records for Pteris (Pteridaceae) from Colombia

A systematic review of the Colombian species of the tribe Pterideae (Pteridaceae) resulted in the discovery of two new species ofPteris:P. muricatopedata andP. albertiae, both in the Deflexa group. Two species are reported for the first time from Colombia:P. bakeri C. Chr. andP. lechleri Mett.     

Effect of temperature on growth and macromolecular biosynthesis in cryptococcus species

Cryptococcus neoformans, a pathogenic yeast, grows at temperatures between 25 and 37°C. However, the closely related non-pathogen C. albidus exhibits restricted growth at temperatures above ambient with little or no growth at 37°C. The inhibition of growth of the non-pathogen, as measured by turbidity, cell number, and per cent budding, is reversible after 48 hr at the non-permissive temperature (37°C). Growth cessation at 37°C is accompanied by a corresponding decrease in DNA synthesis, which is not observed in C. neoformans. RNA and protein synthesis in C. albidus and C. neoformans are only slightly affected at the elevated temperature. Degradation by nucleases does not seem to account for the differences found in this cumulative DNA synthesis in C. albidus at 25 and 37°C. These facts suggest that C. albidus may possess a thermo-sensitive defect in the machinery responsible for the initiation of DNA replication.     

Comparison of the Recovery of Escherichia coli from Frozen Foods and Nutmeats by Confirmatory Incubation in EC Medium at 44.5 and 45.5 C

The productivity of confirmatory EC broth for the isolation of fecal Escherichia coli was determined at 44.5 and 45.5 C. A variety of frozen pre-cooked foods and an assortment of nutmeats were examined after primary incubation in Lauryl Sulfate Tryptose (LST) broth. In 85.3% of the cases, the parallel tubes of EC broth incubated for 24 hr at 44.5 and 45.5 C gave rise to identical E. coli responses of positive, false positive, and negative. The remaining 14.7% of the reactions represent the qualitative difference between the two temperatures. The EC test at 45.5 C was more specific for E. coli, since two- to threefold fewer false positives were produced at this temperature than at 44.5 C. However, fecal E. coli recoveries were slightly higher (4%) at the lower temperature. Incubating the EC tubes from the interval of 24 to 48 hr gave rise to an additional 4.3% of E. coli recovery, but this was accompanied by an excessive production of false positives (75.9%), representing a 3.5-fold decrease in specificity. It is recommended that, in the confirmatory use of EC broth in the examination of frozen foods and nutmeats for the recovery of fecal E. coli, the test be made at 45.5 C in a water bath and limited to 24 hr of incubation only, to insure optimal specificity. During the study, a “fixed” productivity ratio was noted; E. coli⁺/LST⁺ equaled approximately one-fourth or 25%. The significance of this ratio is discussed.     

The antiteratogenic effects of hypo- and hyperthermia in trypan blue-treated chick embryos

Fertile White Leghorn chicken eggs (N = 174) were incubated under optimum conditions until the embryos had reached Hamburger-Hamilton stage 12 (about 48 hr). At that time, 20 μl of 1% trypan blue solution, dissolved in 0.85% NaCl (wt/vol) was injected through the yolk sac into the liquid yolk found just under the embryo. After injection, the eggs were separated into groups and returned to the incubator under control conditions (38°C), or at temperatures lower (35°C) or higher (41°C) than optimum.After an additional 24 hr of incubation, the embryos incubated at 35°C (N = 53) exhibited significantly fewer caudal hematomas (P < 0.02) than did embryos incubated at 38°C (N = 51). Similarly, embryos incubated at 41°C (N = 40) also exhibited significantly fewer caudal hematomas (P < 0.05) than did their corresponding (38°C) controls (N = 30). There was no significant difference between the 35°C group and their controls, or the 38°C group and their controls, in embryonic dry weight, dry weight of the area vasculosa, or crown-rump length. The only other significant difference detected between groups was a very slight but significant (P < 0.0005) decrease in Hamburger-Hamilton stage (0.4 stage unit) between embryos incubated at 35°C and the corresponding controls.Since incubation temperatures either above or below optimum result in a marked reduction in the teratogenic response to trypan blue treatment, we conclude that there exists a temperature optimum for the development of caudal hematomas.     

Bacteriological and Shelf-Life Characteristics of Canned, Pasteurized Crab Cake Mix

The bacteriological spoilage characteristics of a canned, pasteurized crab cake mix product stored at various temperatures were investigated. A large number of bacteria, both mesophilic and psychrophilic, survived the pasteurization process. Bacillus and Micrococcus were found to predominate when the product was stored at 30 C (86 F) and 18 C (64 F), whereas Alcaligenes predominated at 2 C (36 F). The product was found to be free of Escherichia coli. Bacterial counts, trimethylamine nitrogen, volatile reducing substances, and ammonia determinations were evaluated as indices of quality for the product. Close correlation was observed between bacterial counts, volatile reducing substance values, and organoleptic tests when the product was stored at 30 C (86 F). The shelf-life of the product was approximately 6 months at 2 C (36 F), 4 days at 18 C (64 F), and 27 hr at 30 C (86 F).     

Preservation by Freeze-Drying and the Stability of Virulence of Salmonella typhimurium

Differences in the ability to withstand freeze-drying were demonstrated among strains of Salmonella typhimurium. On the average, the number of viable cells in freeze-dried cultures stored at 5 C for 12 to 18 months was approximately one half as large as that found 24 hr after freeze-drying. The viability in samples stored at higher temperatures declined rapidly and was correlated with the dryness of the sample. The virulence for mice of three strains of S. typhimurium did not change appreciably when samples were kept for 1 or 2 years as freeze-dried samples stored at 5 C, or as agar cultures stored at 5 C or at room temperature.     

Influence of Chilling and Freezing Temperatures on Infectivity of Meloidogyne incognita and M. hapla

Egg masses and second-stage larvae of Meloidogyne incognita and M. hapla in soil were exposed to temperatures ranging from 20 to -8 C. Temperature was lowered in 2-day intervals to 16, 12, 8, 4, 0, -4, and -8 C, and the nematodes remained at 4, 0, -4, or -8 C for 18, 14, 10, or 6 days, respectively. Unhatched larvae of both species were more resistant to low temperatures than were embryonic stages. Within the eggs of M. incognita, 7.5% of embryos and 48% of larval stages survived 14 days at 0 C, whereas 9% of embryos and 90% of larval stages in the eggs of M. hapla survived 10 days at -4 C. Second-stage larvae of both species remained infective in sol.1 at 4 or 0 C, but were injured at -4 and -8 C. Infectivily of these larvae was lower in saturated soil than in soil at 51 cm moisture tension at all temperatures.     

Cellular immune competence of a Drosophila mutant with neoplastic hematopoietic organs

In the Tum^l mutant of Drosophila melanogaster, the larval hematopoietic organs undergo neoplastic changes and release into circulation large numbers of blood cells. The lamellocytes, and to a lesser extent the plasmatocytes from which they are derived, are the cells that encapsulate various endogenous tissues and form melanotic tumors. The mutation is temperature sensitive, with maximum gene expression manifested at 29°C. The ability of Tum^l larvae to encapsulate eggs of the wasp parasite Leptopilina heterotoma is dependent not only on temperature, with host larvae much more immune reactive at 29°C than at lower temperatures (15° or 21°C), but also on the interval of time following infection when temperature shift experiments are performed. When the shift of parasitized larvae from 21° to 29°C is delayed by 18 hr the hosts are not as immune reactive as those shifted immediately after infection. Since Tum^l larvae are potentially highly immune reactive at the time of infection (with sufficient numbers of lamellocytes in circulation to encapsulate parasites), the low degree of immune competence in hosts shifted to 29°C after 18 hr or maintained at lower temperatures suggests that the increased capacity of blood cells to react against foreign surfaces is dependent on the cells acquiring new or altered recognition and adherence properties at 29°C. The 18-hr delay may provide the parasite with an opportunity to interfere with the acquisition of these specific cellular alterations. Differential hemocyte counts from parasitized larvae show abnormally low lamellocyte counts in susceptible hosts, indicating that successfully developing parasites interfere with the differentiation of hemocytes.     

Effects of Carbon Dioxide on Coccidian Oocysts from 6 Host Species*

SYNOPSIS Activation of sporozoites in oocysts of Eimeria acervulina (chicken), E. intricata (sheep), and E. scabra (swine) occurred after pretreatment in aqueous 0.02 M cysteine hydrochloride under an atmosphere of CO₂, followed by incubation in a trypsin-bile mixture. Sporozoites of E. stiedae (rabbit), E. bilamellata (squirrel), and Isospora canis (dog) became activated when incubated in trypsin and bile with or without prior CO₂-pretreatment of oocysts; however, when CO₂-pretreatment was used, activation of these species in trypsin and bile was greatly enhanced. For E. acervulina, 12% of the oocysts were activated after 4 hr CO₂-pretreatment and 10 hr incubation in trypsin and bile at 43 C; higher temperatures or longer pretreatment times did not cause greater activation. Eimeria intricata oocysts became activated after 1 hr pretreatment and 10 hr incubation in trypsin and bile at 37, 39 or 41 C, respectively. The highest activation (31%) occurred after 20 hr pretreatment and 10 hr incubation in trypsin and bile at 41 C. Ninety percent of E. scabra oocysts contained active sporozoites after 1 hr CO₂-pretreatment and 10 hr incubation in trypsin and bile at 37 C. At 39 or 41 C, 100% activation occurred with this species after similar pretreatment and treatment periods. With E. bilamellata, 64% activation occurred in nonpretreated oocysts incubated 10 hr in trypsin and bile at 41 C, whereas 100% activation occurred if oocysts were pretreated with CO₂ for 1 hr before treatment with trypsin and bile. Thirty-one, 35, and 36% of CO₂-pretreated E. stiedae oocysts were activated after 1 hr incubation in trypsin and bile at 37, 39 or 41 C, respectively, whereas 1, 2, and 20% activation occurred in nonpretreated oocysts incubated at the same temperatures. Sporozoites in 99-100% of I. canis oocysts were activated after 10 hr treatment in trypsin and bile with or without 1 hr CO₂-pretreatment at 23, 37, 39 or 41 C.     

Mutagenesis of Trichoderma reesei endoglucanase I: impact of expression host on activity and stability at elevated temperatures

Background

Trichoderma reesei is a key cellulase source for economically saccharifying cellulosic biomass for the production of biofuels. Lignocellulose hydrolysis at temperatures above the optimum temperature of T. reesei cellulases (~50°C) could provide many significant advantages, including reduced viscosity at high-solids loadings, lower risk of microbial contamination during saccharification, greater compatibility with high-temperature biomass pretreatment, and faster rates of hydrolysis. These potential advantages motivate efforts to engineer T. reesei cellulases that can hydrolyze lignocellulose at temperatures ranging from 60–70°C.

Results

A B-factor guided approach for improving thermostability was used to engineer variants of endoglucanase I (Cel7B) from T. reesei (TrEGI) that are able to hydrolyze cellulosic substrates more rapidly than the recombinant wild-type TrEGI at temperatures ranging from 50–70°C. When expressed in T. reesei, TrEGI variant G230A/D113S/D115T (G230A/D113S/D115T Tr_TrEGI) had a higher apparent melting temperature (3°C increase in T_m) and improved half-life at 60°C (t_1/2 = 161 hr) than the recombinant (T. reesei host) wild-type TrEGI (t_1/2 = 74 hr at 60°C, Tr_TrEGI). Furthermore, G230A/D113S/D115T Tr_TrEGI showed 2-fold improved activity compared to Tr_TrEGI at 65°C on solid cellulosic substrates, and was as efficient in hydrolyzing cellulose at 60°C as Tr_TrEGI was at 50°C. The activities and stabilities of the recombinant TrEGI enzymes followed similar trends but differed significantly in magnitude depending on the expression host (Escherichia coli cell-free, Saccharomyces cerevisiae, Neurospora crassa, or T. reesei). Compared to N.crassa-expressed TrEGI, S. cerevisiae-expressed TrEGI showed inferior activity and stability, which was attributed to the lack of cyclization of the N-terminal glutamine in Sc_TrEGI and not to differences in glycosylation. N-terminal pyroglutamate formation in TrEGI expressed in S. cerevisiae was found to be essential in elevating its activity and stability to levels similar to the T. reesei or N. crassa-expressed enzyme, highlighting the importance of this ubiquitous modification in GH7 enzymes.

Conclusion

Structure-guided evolution of T. reesei EGI was used to engineer enzymes with increased thermal stability and activity on solid cellulosic substrates. Production of TrEGI enzymes in four hosts highlighted the impact of the expression host and the role of N-terminal pyroglutamate formation on the activity and stability of TrEGI enzymes.

Electronic supplementary material

The online version of this article (doi:10.1186/s12896-015-0118-z) contains supplementary material, which is available to authorized users.     

Improved Temperature-Gradient Incubator and the Maximal Growth Temperature and Heat Resistance of Salmonella

An improved all-metal temperature-gradient incubator produces its gradient by means of a bar permanently installed in a near-vertical position with its lower end in a cool constant-temperature water bath and with thermostatically controlled heaters near its top. Bolts hold the incubator in contact with the temperature-gradient bar, and polyurethane foam insulates the entire assemblage during use. Maximal growth temperatures of 34 representative strains of Salmonella were found to be between 43.2 and 46.2 C. In an agar medium with an initial level of 10⁶ cells per milliliter, no strain survived 50 C for 48 hr. S. senftenberg 775W showed no greater heat resistance at or near 48 C than did other species or other S. senftenberg strains. However, it was considerably more resistant than other strains at 55 C.