Efficacy of entomopathogenic nematodes against soil-dwelling life stages of western flower thrips, Frankliniella occidentalis (Thysanoptera: Thripidae).

The efficacy of six entomopathogenic nematode (EPN) strains was tested in a laboratory study against soil-dwelling life stages of western flower thrips (WFT), Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae). The EPN strain collections screened included two Heterorhabditis bacteriophora species, i.e., H. bacteriophora HK3 (H.b H) and H. bacteriophora HB Brecan (H.b B), three Steinernema feltiae species, i.e., S. feltiae Sylt (S.f S), S. feltiae OBSIII (S.f O), and S. feltiae strain CR (S.f C), and the S. carpocapsae strain DD136 (S.c D). All soil-dwelling life stages of WFT were susceptible to the tested EPN strains. The most virulent strains were S.f S, S.c D, and H.b H. The S.f O strain was highly virulent against late second instar larvae and prepupae of WFT under high soil moisture conditions, but less effective against pupae under comparatively drier soil conditions. Results from dose rate experiments indicate that a comparatively high concentration of 400 infective juveniles (IJs) per cm(2) was needed to obtain high mortality in all soil-dwelling life stages of WFT. However, dose rates of 100-200 IJs/cm(2) already caused 30-50% mortality in WFT. The chances for combining EPNs with other biological control agents of WFT are discussed.     

Evaluation of entomopathogenic nematodes for control of the beetle, Luperomorpha suturalis Chen (Col., Chrysomelidae)

Abstract: The strains Steinernema feltiae Otio and A54, Steinernema ceratophorum D43 and Steinernema carpocapsae BJ were tested for their infectivity to the larvae and pupae of beetle ( Luperomorpha suturalis Chen) at 25 ± 0.5°C and 15 ± 0.5°C in laboratory conditions. The results, based on comparison of the insect mortalities and nematode penetration rates among four nematode strains, showed that S. feltiae Otio was a potential biocontrol agent of the larvae and pupae of L. suturalis . The mortalities of the larvae and pupae exposed to S. feltiae Otio strain were 95.8 and 97.1% at 25 ± 0.5°C and 78.0 and 83.0% at 15 ± 0.5°C, respectively. The nematode penetration rates of S. feltiae Otio of the larvae and pupae were 15.6 and 19.0% at 25 ± 0.5°C, 2.6 and 6.3% at 15 ± 0.5°C, respectively. Field efficacy of S. feltiae Otio strain was examined against beetle larvae in Hebei province, northern China. The population reduction of insect larvae was 77.8 and 13.9% at doses of 30 and 15 infective juveniles (IJs)/cm² of S. feltiae Otio after 38 days of treatment and 90.2 and 92.4% after 100 days of treatment. However, the population of the insect larvae was reduced only to 15.5 and 15.7% when treated with pesticide after 38 and 100 days, respectively. The efficiency between the two nematode doses was not significantly different but it was remarkably higher than that of the pesticide after 100 days of application. The results suggest that S. feltiae Otio strain could be an alternative to pesticide for beetle control.     

Acclimation of entomopathogenic nematodes to novel temperatures: trehalose accumulation and the acquisition of thermotolerance

The effect of thermal acclimation on trehalose accumulation and the acquisition of thermotolerance was studied in three species of entomopathogenic nematodes adapted to either cold or warm temperatures. All three Steinernema species accumulated trehalose when acclimated at either 5 or 35 degrees C, but the amount of trehalose accumulation differed by species and temperature. The trehalose content of the cold adapted Steinernema feltiae increased by 350 and 182%, of intermediate Steinernema carpocapsae by 146 and 122% and of warm adapted Steinernema riobrave by 30 and 87% over the initial level (18.25, 27.24 and 23.97 microg trehalose/mg dry weight, respectively) during acclimation at 5 and 35 degrees C, respectively. Warm and cold acclimation enhanced heat (40 degrees C for 8h) and freezing (-20 degrees C for 4h) tolerance of S. carpocapsae and the enhanced tolerance was positively correlated with the increased trehalose levels. Warm and cold acclimation also enhanced heat but not freezing tolerance of S. feltiae and the enhanced heat tolerance was positively correlated with the increased trehalose levels. In contrast, warm and cold acclimation enhanced the freezing but not heat tolerance of S. riobrave, and increased freezing tolerance of only warm acclimated S. riobrave was positively correlated with the increased trehalose levels. The effect of acclimation on maintenance of original virulence by either heat or freeze stressed nematodes against the wax moth Galleria mellonella larvae was temperature dependent and differed among species. During freezing stress, both cold and warm acclimated S. carpocapsae (84%) and during heat stress, only warm acclimated S. carpocapsae (95%) maintained significantly higher original virulence than the non-acclimated (36 and 47%, respectively) nematodes. Both cold and warm acclimated S. feltiae maintained significantly higher original virulence (69%) than the non-acclimated S. feltiae (0%) during heat but not freezing stress. In contrast, both warm and cold acclimated S. riobrave maintained significantly higher virulence (41%) than the non-acclimated (14%) nematodes during freezing, but not during heat stress. Our data indicate that trehalose accumulation is not only a cold associated phenomenon but is a general response of nematodes to thermal stress. However, the extent of enhanced thermal stress tolerance conferred by the accumulated trehalose differs with nematode species.     

Impact of Entomopathogenic Nematodes on Different Soil-dwelling Stages of Western Flower Thrips, Frankliniella occidentalis (Thysanoptera: Thripidae), in the Laboratory and Under Semi-field Conditions

The impact of entomopathogenic nematodes (EPN) on mortality of soil-dwelling stages of western flower thrips (WFT), Frankliniella occidentalis (Thysanoptera: Thripidae) with different insect stage combinations was studied in the laboratory and under semi-field conditions. In laboratory experiments, the efficacy of Steinernema feltiae strain Sylt (Rhabditida: Steinernematidae) at a concentration of 400 infective juveniles (IJs) cm -2 was tested against different proportions of soil-dwelling stages of WFT, i.e. late second instar larvae (L2), prepupae and pupae. Soil was used as the testing medium. S. feltiae significantly affected the mortality of all soil-dwelling life stages of WFT at all tested insect stage combinations. The proportion of late L2 in the population negatively correlated to EPN-induced mortality. WFT prepupa and pupa were similarly susceptible to S. feltiae and their proportion in the population did not affect the EPN-induced mortality under laboratory conditions. The highest mortality (80%) was recorded when the population consisted only of prepupae and/or pupae. In the semi-field study, the impact of S. feltiae , S. carpocapsae strain DD136 and Heterorhabditis bacteriophora strain HK3 (Rhabditida: Heterorhabditidae) ( H. bacteriophora ) at concentrations of 400 and 1000 IJs cm -2 was evaluated against WFT reared on green beans, Phaseolus vulgaris L., as host plant in pot experiments in a controlled climate chamber. All tested EPN strains at both dose rates significantly reduced the WFT populations. Up to 70% reduction of the WFT population was obtained at the higher EPN concentration.     

Virulence of entomopathogenic nematodes to pecan weevil larvae, Curculio caryae (Coleoptera: Curculionidae), in the laboratory

The pecan weevil, Curculio caryae (Horn), is a key pest of pecans in the Southeast. Entomopathogenic nematodes have been shown to be pathogenic toward the larval stage of this pest. Before this research, only three species of nematodes had been tested against pecan weevil larvae. In this study, the virulence of the following nine species and 15 strains of nematodes toward fourth-instar pecan weevil was tested: Heterorhabditis bacteriophora Poinar (Baine, HP88, Oswego, NJ1, and Tf strains), H. indica Poinar, Karunakar & David (original and Homl strains), H. marelatus Liu & Berry (IN and Point Reyes strains), H. megidis Poinar, Jackson & Klein (UK211 strain), H. zealandica Poinar (NZH3 strain), Steinernema riobrave Cabanillas, Poinar & Raulston (355 strain), S. carpocapsae (Weiser) (All strain), S. feltiae (Filipjev) (SN strain), and S. glaseri (Steiner) (NJ43 strain). No significant difference in virulence was detected among nematode species or strains. Nematode-induced mortality was not significantly greater than control mortality (in any of the experiments conducted) for the following nematodes: H. bacteriophora (Baine), H. zealandica (NZH3), S. carpocapsae (All), S. feltiae (SN), S. glaseri (NJ43), and S. riobrave (355). All other nematodes caused greater mortality than the control in at least one experiment. Heterorhabditis megidis (UK211) but not H. indica (original) displayed a positive linear relationship between nematode concentration and larval mortality. Results suggested that, as pecan weevil larvae age, they may have become more resistant to infection with entomopathogenic nematodes.     

病原线虫对桔小实蝇种群的控制作用

通过室内和田间实验研究了昆虫病原线虫对桔小实蝇Bactrocera (Bactrocera) dorsalis (Hendel)的控制作用。室内实验结果表明,供试的3种线虫的4个品系（小卷蛾斯氏线虫Steinernema carpocapsae All品系与A24品系,夜蛾斯氏线虫Steinernema feltiae SN品系和嗜菌异小杆线虫Heterorhabditis bacteriophora H06品系）,以小卷蛾斯氏线虫All品系对桔小实蝇的侵染力最强,其3天的LD₅₀和LD₉₅分别为35.0和257.1条/cm²土壤。按300条/cm²土壤的量施用,小卷蛾斯氏线虫All品系对当代桔小实蝇的控制效果为86.3%。用以虫期作用因子组建的生命表方法评价了小卷蛾斯氏线虫All品系对田间桔小实蝇下代种群的控制作用,结果表明,按300条/cm²土壤的量施用线虫,对照杨桃园的桔小实蝇种群趋势指数为105.9,而处理杨桃园的桔小实蝇种群趋势指数下降为15.5;小卷蛾斯氏线虫All品系对桔小实蝇的干扰控制指数为0.146,即线虫处理果园的下代种群密度仅为对照果园的14.6%。     

Effectiveness of Entomopathogenic Nematodes in an Alginate Gel Formulation Against Lepidopterous Pests

An improved calcium alginate gel formulation was developed and tested as a carrier for entomopathogenic nematodes against Spodoptera littoralis and Helicoverpa armigera larvae. Mortality of 100% was caused in 4th instar larvae of the two insects by feeding them on 1000 infective juveniles (IJ) g -1 of Steinernema carpocapsae (ALL strain) in the gel for 24 h. Exposing 2nd to 5th instars of H. armigera and 3rd to 6th of S. littoralis to 500 IJ g -1 of S. carpocapsae (ALL strain) resulted in 70-100% larval mortality. Mature larvae were less susceptible to the nematodes. Mortality of larvae exposed to 500 IJg -1 of S. carpocapsae (ALL strain) ranged from about 45-55% at 4 h to 90-95% at 48 h. Fourth instar larvae fed for 24 h with 250 IJ g -1 of nematode strains in gel showed in S. littoralis ranges of susceptibility in the following descending order: S. feltiae (IS -7 strain) = S. carpocapsae (DT strain) = S. feltiae (IS-6 strain) > S. carpocapsae (Mexican strain) = S. carpocapsae (ALL strain) = Heterorhabditis bacteriophora (HP-88 strain) = H sp. (IS-5 strain) > S. riobravae (Texas strain); in H. armigera the rating was: S. feltiae (IS-7 strain) = H. bacteriophora (HP88 strain) > S. carpocapsae (ALL strain) = S. feltiae (IS-6 strain ) = Heterorhabditis sp. (IS5 strain) > S. carpocapsae (Mexican strain) > S. riobravae (Texas strain) . The number of nematodes per larval cadaver increased with mortality rates. In greenhouse tests at 28 &#45 2°C and 90% relative humidity, gel discs containing 500 IJ g -1 of nematodes were pinned to leaves of potted plants of cotton ( Gossypium hirsutum ) (Acala SJ2) and the plants were offered to S. littoralis larvae. Larval mortality of 89 &#45 12.7% was caused by S. feltiae (IS-7 strain) and most of the plant leaves were protected against the larvae by the nematodes. In the control, larval mortality was 3.3 &#45 0.05% and the plants were almost completely defoliated. Possibilities of using the gel-nematode formulation to protect sheltered crops against insect pests are discussed     

Virulence of entomopathogenic nematodes to Diaprepes abbreviatus (Coleoptera: Curculionidae) in the laboratory

The Diaprepes root weevil, Diaprepes abbreviatus (L.) is the most severe weevil pest in Florida citrus. Entomopathogenic nematodes have effectively suppressed larval populations of D. abbreviatus. Our objective was to conduct a broad laboratory comparison of entomopathogenic nematodes for virulence toward larvae of D. abbreviatus. The study was conducted at three temperatures (20, 24, and 29 degrees C) and included nine entomopathogenic species and 17 strains: Heterorhabditis bacteriophora Poinar (Baine, NJl, Hb, Hbl, HP88, and Lewiston strains), H. indica Poinar, Karunakar & David (original and Homl strains), H. marelatus Liu & Berry (IN and Point Reyes strains), H. megidis Poinar, Jackson & Klein (UK21l strain), H. zealandica Poinar (NZH3 strain), Steinernema riobrave Cabanillas, Poinar & Raulston (355 strain), S. carpocapsae (Weiser) (All strain), S. feltiae (Filipjev) (SN and UK76 strains), and S. glaseri (Steiner) (NJ43 strain). At 20 degrees C, the greatest mortality was caused by S. riobrave although it was not significantly greater than H. bacteriophora (Baine), H. bacteriophora (Hb), H. bacteriophora (Hbl), and H. indica (original). At 24 and 29 degrees C, S. riobrave caused greater larval mortality than other nematodes tested. Two strains of H. indica, H. bacteriophora (Baine), and S. glaseri were next in terms of virulence at 29 degrees C. Our results suggest that S. riobrave has the greatest potential for control of D. abbreviatus.     

Foliar Application of the Entomopathogenic Nematode Steinernema feltiae Against Leafminers on Vegetables

Data from a comparative study of the efficacy of Steinernema feltiae for the control of three species of leafminer formed the basis of an application schedule which successfully suppressed an outbreak of the statutory leafminer pest, Liriomyza huidobrensis . All three instars of Liriomyza bryoniae and L. huidobrensis were similarly susceptible to S. feltiae at 20 o C and > 90% relative humidity (RH). Although all larval instars of Chromatomyia syngenesiae were susceptible to S. feltiae , mortality was lower than for L. bryoniae . Repeat applications of S. feltiae to L. bryoniae and C. syngenesiae indicated that a nematode treatment to the second/early third instar larvae was more effective than applying higher rates of nematodes when humidities were less than 90% RH. In a trial on lettuce at a commercial glasshouse a mean L. huidobrensis mortality of 82 +/- 5% was recorded after an S. feltiae application, significantly higher than the chemical treatment, heptenophos.     

Reclassification of subspecies of Acidovorax avenae as A. Avenae (Manns 1905) emend., A. cattleyae (Pavarino, 1911) comb. nov., A. citrulli Schaad et al., 1978) comb. nov., and proposal of A. oryzae sp. nov

The bacterium Acidovorax avenae causes disease in a wide range of economically important monocotyledonous and dicotyledonous plants, including corn, rice, watermelon, anthurium, and orchids. Genotypic and phenotypic relatedness among strains of phytopathogenic A. avenae subsp. avenae, A. avenae subsp. citrulli, A. avenae subsp. cattleyae and A. konjaci, as well as all other Acidovorax species, including A. facilis, the type strain of Acidovorax, was determined. The 16s rDNA sequencing confirmed previous studies showing the environmental species to be very distant from the phytopathogenic species. DNA/DNA reassociation assays on the different strains of A. avenae revealed four (A, B, C, and D) distinct genotypes. Taxon A included six A. avenae subsp. avenae strains from corn that had a mean reciprocal similarity of 81%; taxon B included six A. avenae subsp. avenae strains from rice that had a mean reciprocal similarity of 97%; taxon C contained 11 A. avenae subsp. citrulli strains from cucurbits (cantaloupe, watermelon, and pumpkin) that had a mean reciprocal similarity of 88%, and taxon D contained four A. avenae subsp. cattleyae strains from orchids that had a mean similarity of 98%. The mean reciprocal relatedness between taxa A, B, C, and D was less than 70%. Sequence analysis of 16S rDNA and the 16S-23S rDNA internally transcribed spacer region, as well as AFLP analysis, revealed the same four taxa. All four were easily differentiated phenotypically from each other and from all other recognized Acidovorax species. Strains of A. avenae did not contain 3-hydroxyoctanoic acid, which was found in all other species. On the basis of these and previous genetic and phenotypic results, we propose an emendation of the species A. avenae. A. avenae subsp. citrulli (C strains) and A. avenae subsp. cattleyae (D strains) should be elevated to species rank as A. citrulli and A. cattleyae, respectively. We further propose a new taxon for the B strains, A. oryzae sp. nov. with FC-143T = ICPB 30003T = ICMP 3960T = ATCC 19882T as the type strain.     

Susceptibility of diamond back moth,Plutella xylostella (L) to entomopathogenic nematodes

Eight entomopathogenic nematode species / strains, Steinernema glaseri (steiner), S. carpocapsae (Weiser), S. feltiae (Filipjev), Steinernema sp. Ecomax strain, Heterorhabditis bacteriophora (Pioner), Heterorhabditis sp. Ecomax strain, two locally isolated strains called as JFC and TFC were tested against the final instar larvae of diamond back moth, Plutella xylostella (L.). All nematodes were found pathogenic. However, H. bacteriophora was adjudged the most pathogenic amongst the test nematodes on the basis of LD50 (9.16 IJS/larva), LT50 (43.26 hr), Lex T50 (3.24 hr) and the propagation potential (average of 271.42 IJS/mg) on the host body weight.     

Entomopathogenic Nematode Infectivity Assay: Comparison of Penetration Rate into Different Hosts

Penetration rate (the percentage of the initial infective juvenile inoculum that invades an insect host) was tested as an indicator of entomopathogenic nematode infectivity. Several host-parasite-substrate combinations were evaluated for penetration rate. Four steinernematids, Steinernema carpocapsae, S. glaseri, S. feltiae, S. riobravis and two strains of Heterorhabditis bacteriophora were tested in a contact bioassay against the wax moth, Galleria mellonella, the yellow meal worm, Tenebrio molitor, the beet armyworm, Spodoptera exigua, the black cutworm, Agrotis ipsilon, and the European corn borer, Ostrinia nubilalis. The insect larvae were confined individually in sand and filter paper arenas and exposed to 200 infective juveniles. After incubation, dead insects were dissected in order to count the nematodes penetrated. The data were analyzed for the effects of nematode strain and substrate on penetration rate. The bioassay substrate had a variable effect depending on the insect species. The nematode effect was highly significant for all insects tested. The penetration rate therefore allowed comparisons among nematode strains invading a host. Nematode ranking for infectivity differed according to the insect tested.     

利用26S rDNAD1/D2区序列和微卫星标记分析各种工业酿酒酵母的种内遗传差异

选取30株传统发酵工业中不同来源的酿酒酵母菌株和实验室常用酿酒酵母菌株,利用大亚基(26S)rDNA D1/D2区序列和微卫星标记分析酿酒酵母种内菌株间的遗传多样性和系统发育关系,以揭示酿酒酵母在长期的各种工业发酵环境下发生的遗传变异。结果表明:30株酿酒酵母的26S rDNA D1/D2区序列(591bp)比较保守,与测序菌株S288C相比序列相似度在99.8%–100%,说明种内菌株间存在一定的多态性,但序列差异并不十分明显,其变异情况表现在个别碱基的差异(大多由转换突变引起);通过扩增11个微卫星位点,每个菌株均有其独特的基因型,即30种基因型,共得到188个等位基因,观测杂合度平均值和期望杂合度平均值分别为0.576、0.886,多态信息含量平均值高达0.858,说明酿酒酵母种内菌株间具有较高的遗传多样性,而聚类分析表明30株酿酒酵母可以得到很好地区分,但是没有呈现与其工业来源相关的聚类。     

Laboratory Evaluation of Seven Pakistani Strains of Entomopathogenic Nematodes Against a Stored Grain Insect Pest, Pulse beetle Callosobruchus chinensis (L.)

Seven Pakistani strains of entomopathogenic nematodes belonging to the genera Steinernema and Heterorhabditis were tested against last instar and adult stages of the pulse beetle, Callosobruchus chinensis (L.). These nematodes included Steinernema pakistanense Shahina, Anis, Reid and Maqbool (Ham 10 strain); S. asiaticum Anis, Shahina, Reid and Rowe (211 strain); S. abbasi Elawad, Ahmad and Reid (507 strain); S. siamkayai Stock, Somsook and Reid (157 strain); S. feltiae Filipjev (A05 strains); Heterorhabditis bacteriophora Poinar (1743 strain); and H. indica Poinar, Karunakar and David (HAM-64 strain). Activity of all strains was determined at four different nematode densities in Petri dishes and in concrete containers. A significant nematode density effect was detected for all nematode species tested. Overall, Heterorhabditis bacteriophora, S. siamkayai, and S. pakistanense were among those that showed the highest virulence to pulse beetle larvae and adults. For all nematode species, the last larval stage of the pulse beetle seems to be more susceptible than the adult. LC(50) values in Petri dish and concrete containers were 14-340 IJs/larvae and 41-441 IJs/larvae, respectively, and 59-1376 IJs/adult and 170-684/adult, respectively.     

Critical factors required by the nematode Steinernema feltiae for the control of the leafminers Liriomyza huidobrensis, Liriomyza bryoniae and Chromatomyia syngenesiae

The critical factors required by entomopathogenic nematodes for the control of dipteran leafminers were investigated and compared. The foliar application of Steinernema feltiae and Heterorhabditis sp. (strain UK 211) caused a significant reduction in the numbers of dipteran leafminer larvae. There was no significant difference in efficacy between these nematode species against Liriomyza huidobrensis at 20°C and 85–90% r.h. Application of S. feltiae to all three larval instars of L. huidobrensis significantly reduced larval survival, with application to the second instar being the most effective at 20 ± 2°C and 80 ± 10% r.h. Humidities of >90% significantly enhanced the efficacy of S. feltiae. When S. feltiae was applied to second instar larvae, it was equally effective throughout the temperature range 10–30°C. At 20°C and >90% r.h., sufficient nematodes were able to enter the leaf tissues within the first 12 h after application to reduce larval survival to c. 15%. A comparison between L. huidobrensis, L. bryoniue and Chromatomyia syngenesiue indicated that the efficacy of S. feltiae was affected by the same critical factors for all three species.     

Interaction of microbial populations in Steinernema (Steinernematidae,Nematoda) infected Galleria mellonella larvae

Infection of Galleria mellonella larvae with the entomopathogenic nematodes Steinernema feltiae (A21 and R strains) and Steinernema glaseri (Dongrae) resulted in several species of bacteria, including the respective bacterial symbiont, Xenorhabdus spp., growing in the infected insect cadavers. These other bacteria were Enterococcus in all three nematode infections studied and Acinetobacter in the S. feltiae infections. The respective populations of these bacteria changed with time. Following infection of G. mellonella larvae with any one of the Steinernema sp., only Enterococcus bacteria were detected initially in the dead larvae. Between 30 and 50h post-infection Xenorhabdus bacteria were detected and concurrent with this Enterococcus population declined to zero. This was probably due to secondary metabolites with antibacterial properties that were produced by Xenorhabdus. In the S. feltiae (both R and A21 strains) infections a third bacterium, Acinetobacter, appeared at about 130h (in S. feltiae A21 infections) or 100h (in S. feltiae R infections) and increased in population size to approximately that of Xenorhabdus. It was demonstrated that Enterococcus, orginating from the G. mellonella digestive tract, was sensitive to the organically soluble antimicrobials produced by Xenorhabdus but Acinetobacter, which was carried by the nematode, was not.     

Selective breeding of Steinernema feltiae (Filipjev) (Nematoda: Steinernematidae) for improved efficacy in control of a mushroom fly,Lycoriella solani Winnertz (Diptera: Sciaridae)

A method of selecting a Steinernema feltiae strain that is effective against a mushroom fly, Lycoriella solani, is described in detail. The pest control efficacy of the selected nematode strain was evaluated and compared with the efficacy of two unselected strains. The selection procedure was designed to give preference to nematode individuals with the greatest ability (1) to search effectively for the target insect larvae in their natural habitat, (2) to infect them shortly after application and (3) to reproduce in their haemocoel. Thirty‐four rounds of selection achieved a 4‐fold improvement in nematode ability to find and parasitize third‐ and fourth‐instar larvae of the pest in the mushroom substrate. In 24‐h laboratory experiments, mortality of the insect caused by nematode juveniles rose from 22.5%, recorded for the original unselected isolate, to 92.5% for the selected strain. In a 51‐day experiment conducted on a mixed age mushroom house population of L. solani, the enhanced pest control ability of the selected strain was detected shortly after nematode application and remained high throughout the experimental period. During the first 4 weeks of the trial the selected nematode strain was significantly better than both unselected strains and caused 91.1–92.7% reduction of the fly emergence from the mushroom substrate. No difference was observed between the efficacy of the selected nematodes applied at 1 × 10⁶ and 3 ×10⁶ infective juveniles per m², while the unselected strains performed significantly better at the higher concentration. All the nematodes examined showed good persistence in the mushroom casing apparently due to recycling in the insect host.     

Steinernema cholashanense n. sp. (Rhabditida, Steinernematidae) a new species of entomopathogenic nematode from the province of Sichuan, Chola Shan Mountains, China

During a random survey of entomopathogenic nematodes in the provinces of Sichuan and Gansu (eastern Tibet) in 2004, soil samples from several sites were collected and tested for the incidence of entomopathogenic nematodes. A new species was collected in this survey and it is described herein as Steinernema cholashanense n. sp. Steinernema cholashanense n. sp. is characterized by morphology and morphometry of the IJ and male. For the IJ, the new species can be recognized by the average body length 843 microm, esophagus length 125 microm, H%=39% and E%=81%. The lateral field pattern is 2, 5, 7, 4, 2. The male of the first generation is characterized by spicule shape and length and especially with prominent velum and the presence of a mucron on both generations. The average body length of the IJ of S. cholashanense n. sp. (843 microm) is shorter than that of S. oregonense (980 microm), S. kraussei (951 microm) and S. litorale (909 microm), similar to that of S. feltiae (849 microm), but longer than that of S. weiseri (740 microm), S. jollietti (711 microm) and S. hebeiense (658 microm). Esophagus length of the new species (125 microm) is closer to that of S. jollieti (123 microm) but longer than that of S. weiseri (113 microm) and shorter than that of S. oregonense (132 microm), S. kraussei (134 microm) and S. feltiae (136 microm). E% of the new species (81) is similar to that of S. kraussei (80), but smaller than that of S. jollieti (88), S. weiseri (95), S. oregonense (100) and S. feltiae (119). Spicule head length of the new species is almost the same as its width, this character is similar to that of S. kraussei but it is different from this species by its prominent velum. The new species can be recognized further by characteristics of sequences of ITS and D2D3 regions and cross hybridization with closely related species, S. feltiae, S. kraussei and S. oregonense.     

两种色型黄粉虫抗冻蛋白cDNA克隆、序列分析与表达分析

产生抗冻蛋白(antifreeze proteins, AFPs)是大多数昆虫抵抗低温的一个重要策略。为给黄粉虫Tenebrio molitotr耐寒机理研究提供参考, 本研究采用RT-PCR、 5′ RACE和3′ RACE法克隆了黄、 黑两种色型黄粉虫幼虫的抗冻蛋白基因Tm-afp, 分析了其基因序列及所编码的氨基酸序列, 并检测了其在这两种色型间的mRNA水平差异。结果表明： 从黄、 黑两种色型黄粉虫幼虫克隆出的Tm-afp cDNA全长分别为579 bp和588 bp, 它们包含一个20 bp的5′端非翻译区、 一个402 bp的开放阅读框和一个变异较大的3′ 端非翻译区, 其碱基序列一致性为95%。由于这两个抗冻蛋白基因编码的蛋白成熟肽存在两个氨基酸差异： 第35位（D→E）和130位（T→S）, 因此将其判定为黄粉虫抗冻蛋白基因Tm-afp的两个异构体, 并分别命名为Tm-afp-1和Tm-afp-2。Tm-afp-1和Tm-afp-2编码的抗冻蛋白异构体（分别为Tm-afp-1和Tm-afp-2 ）信号肽之间有3个氨基酸差异： 第2位（G→A）、 9位（S→T）和27位（Y→N）; 其成熟肽的第一个氨基酸为谷氨酰胺, 含8个高度保守的12 aa短串联重复序列, 每个重复序列的第2位和第8位为半胱氨酸。此外, Tm-afp-1和Tm-afp-2均属富含苏氨酸和半胱氨酸的昆虫抗冻蛋白, 其二级结构均由大量的β-折叠和无规则卷曲组成, 三级结构为8个特殊的右手β-螺旋, 每一圈螺旋由12个氨基酸组成; 在β-螺旋的一个侧面规则排列着由保守XCT形成的β折叠片层。同时, 低温可以诱导黄、 黑两种色型黄粉虫Tm-afp的表达, 但长时间低温诱导下黑色型黄粉虫幼虫的Tm-afp表达量显著高于黄色型黄粉虫幼虫Tm-afp表达量。结果提示, 黄粉虫种内不同色型间, 抗冻蛋白基因Tm-afp可能存在多种异构体, 并且黑色型黄粉虫Tm-afp比黄色型黄粉虫Tm-afp对低温的应答更强烈。这一研究结果为进一步探索黄粉虫的耐寒性机理提供了有益参考。