Field trials against Hoplia philanthus (Coleoptera: Scarabaeidae) with a combination of an entomopathogenic nematode and the fungus Metarhizium anisopliae CLO 53

The white grub, Hoplia philanthus Füessly (Coleoptera: Scarabaeidae), is a major pest of turf and ornamental plants in Belgium. Previously, the combination of lethal concentration of the entomopathogenic nematodes Heterorhabditis megidis or Steinernema glaseri with the entomopathogenic fungus Metarhizium anisopliae (strain CLO 53) caused additive or synergistic mortality to third-instar H. philanthus in the laboratory and greenhouse. In this present study, we examined this interaction under field conditions and compared a combination of a commercial formulation of Heterorhabditis bacteriophora (Nema-green^®) and M. anisopliae. Controls were M. anisopliae, chlorpyrifos (Dursban 5 Granules) and H. bacteriophora. Field applications (surface or subsurface) were made against a mixed population of second/third-instar H. philanthus at a sport field and lawn infested in the province of West-Flanders. In both trials, the combination of M. anisopliae with H. bacteriophora at 5 × 10¹² conidia/ha +2.5 × 10⁹ infective juveniles/ha resulted in additive or synergistic effects, causing more than 95% grub mortality when the nematodes was applied 4 weeks after the application of fungus. However, application of nematode, chlorpyrifos or fungus alone provided 39–66%, 42–60% (surface) and 33–76%, 82–100% or 37–65%, (subsurface) control of H. philanthus. We concluded that the pathogen combinations we tested are compatible elements of integrated pest management and are likely to improve control of H. philanthus larvae and perhaps other insect pests beyond what is expected from single application of the pathogen.     

Effects of Culture Fluids and Preinduction of Chitinase Production on Biocontrol of Bipolaris Leaf Spot by Stenotrophomonas maltophilia C3

Control of leaf spot, caused by Bipolaris sorokiniana, on tall fescue and perennial ryegrass by Stenotrophomonas maltophilia C3 was enhanced in growth chamber and field experiments by application of bacterial cells in culture fluids as compared to phosphate buffer. C3 population levels on leaves were up to 0.8 log units higher when applied with culture fluid than with phosphate buffer. Although fluids alone were inhibitory to conidial germination and leaf spot development, there was a synergistic effect when combined with C3 cells. Fluids from broth cultures with chitin as the carbon source were more inhibitory than those containing glucose, and the suppressiveness of a culture fluid was related to the age of the culture from which it was collected. Both of these effects were associated with the production of high levels of chitinase (EC 3.2.1.14), protease (EC 3.4.21-24), β-1,3-glucanase (EC 3.2.1.58), and lipase (EC 3.1.1.3) in the fluid. Culture fluids had a durable protective effect, inhibiting disease development even when applied 9 days before inoculation, and had a therapeutic effect if applied within 3 days after inoculation. When chitin was applied with C3 cells preinduced or noninduced for chitinase production (i.e., grown on chitin-containing or chitin-lacking media, respectively) biocontrol efficacy was significantly increased over either cell type without chitin, but the addition of chitin did not always increase colonization by C3. In general, preinduced cells were more effective than noninduced cells. The most effective field treatment was the combination of chitin with induced cells in culture fluid.     

Expression in Escherichia coli and Simple Purification of Human Fhit Protein

The fragile histidine triad (Fhit) protein is a homodimeric protein with diadenosine 5′,5-P¹,P³-triphosphate (Ap₃A) asymmetrical hydrolase activity. We have cloned the human cDNA Fhit in the pPROEX-1 vector and expressed with high yield in Escherichia coli with the sequence Met-Gly-His₆-Asp-Tyr-Asp-Ile-Pro-Thr-Thr followed by a rTEV protease cleavage site, denoted as “H6TV,” fused to the N-terminus of Fhit. Expression of H6TV–Fhit in BL21(DE3) cells for 3 h at 37°C produced 30 mg of H6TV–Fhit from 1 L of cell culture (4 g of cells). The H6TV–Fhit protein was purified to homogeneity in a single step, with a yield of 80%, using nickel-nitrilotriacetate resin and imidazole buffer as eluting agent. Incubation of H6TV–Fhit with rTEV protease at 4°C for 24 h resulted in complete cleavage of the H6TV peptide. There were no unspecific cleavage products. The purified Fhit protein could be stored for 3 weeks at 4°C without loss of activity. The pure protein was stable at −20°C for at least 18 months when stored in buffer containing 25% glycerol. Purified Fhit was highly active, with a K_m value for Ap₃A of 0.9 μM and a k_cat(monomer) value of 7.2 ± 1.6 s⁻¹ (n = 5). The catalytic properties of unconjugated Fhit protein and the H6TV–Fhit fusion protein were essentially identical. This indicates that the 24-amino-acid peptide containing the six histidines fused to the N-terminus of Fhit does not interfere in forming the active homodimers or in the binding of Ap₃A.     

粘菌素对耐碳青霉烯类抗生素鲍曼不动杆菌的抗菌作用

感染耐碳青霉烯的鲍曼不动杆菌(CR-Ab)常与高发病率和死亡率相关联,而可供选择的治疗方案有限,大多基于与粘菌素联用。长期用药导致CR-Ab对粘菌素也产生一定抗性。为了评估含有或不含有粘菌素的不同抗菌组合对从CR-Ab感染患者收集的CR-Ab临床分离株的体外抗菌活性,本研究从本院就诊的患者中收集CR-Ab菌株,通过常量肉汤稀释法(MBD)测定最低抑菌浓度(MICs),通过定性(棋盘法)和定量(即杀菌测试)方法评估各组药物协同活性。结果发现所有菌株均是碳青霉烯类抗性的,且其中两株菌对粘菌素有抗性。棋盘法结果表明含粘菌素的组合在不同处理时间下具有完全协同作用,粘菌素+万古霉素和粘菌素+利福平表现出最高的协同增效作用;不含粘菌素的组合则在35.7%的菌株中观察到完全协同作用。杀菌测试表明粘菌素+美罗培南、粘菌素+替加环素和美罗培南+替加环素组合对粘菌素敏感和低粘菌素抗性的菌株具有杀菌和协同作用,而只有粘菌素+万古霉素和粘菌素+利福平组合表现出持久的杀菌活性。     

In vitro evaluation of combination of Trichoderma harzianum and chitosan for the control of sapstain fungi

In vitro assays were undertaken to evaluate the control of two sapstain fungi, Leptographium procerum and Sphaeropsis sapinea by a combination of chitosan or chitosan oligomer and an albino strain of Trichoderma harzianum. Spore germination and hyphal growth of the test fungi were assessed on media amended with chitosan or chitosan oligomer with and without T. harzianum using either simultaneous inoculation with test fungus or inoculation 1, 2, or 3 days after pre-infection with test fungus.There was no mycelial growth of the test fungi regardless of chitosan concentrations used when either L. procerum or S. sapinea was simultaneously inoculated with T. harzianum. However, the dose–response of chitosan or chitosan oligomer on the test fungi was apparent when T. harzianum was not simultaneously inoculated with test fungus but introduced later. There was a greater growth reduction at higher concentrations (0.075–0.1% v/v) of chitosan, and overall chitosan oligomer was more effective than chitosan aqueous solution.Chitosan alone was able to restrict or delay the germination of spores but the combination of chitosan and T. harzianum inhibited spore germination and hence colony formation of test fungi regardless of time delay.     

酸枣果提取物在体内外对抗生素的增效作用及机制研究

作者研究团队先前从酸枣果的氯仿提取物中精制得到其低极性范围的活性组合物Fr.2a,发现Fr.2a与多种抗生素联用显示出广泛的协同抗菌作用。该研究在Fr.2a的基础上利用硅胶柱层析对酸枣果氯仿提取物中其他极性范围内的活性成分进行了分离纯化,得到精制物Fr.B,并对精制物Fr.B进行GC MS、核磁共振氢谱、红外光谱分析,以确定Fr.B的组成成分;通过抗菌谱分析和细胞通透性分析,以明确Fr.B的抗菌增效谱和抗菌增效机制;采用熔和法将精制物Fr.B制备成软膏,通过小鼠伤口感染模型评价该软膏对抗生素的增效效果。结果表明：(1)由酸枣果氯仿提取物进一步精制得到的Fr.B组分,主要包含反油酸、油酸、顺 10 十六碳烯醇、棕榈酸等脂肪酸类化合物。(2)Fr.B分别与庆大霉素、妥布霉素、氨苄青霉素、氯霉素、红霉素、夫西地酸、制霉菌素、酮康唑和两性霉素B等多种抗生素联用时显示出广泛的协同抗菌作用。(3)Fr.B可破坏细胞膜和细胞壁的完整性而增强细菌细胞的通透性。(4)在体内和体外Fr.B均能显著增强红霉素对耐甲氧西林金黄色葡萄球菌(MRSA)的杀菌作用,从而提高红霉素对MRSA菌株引起的伤口感染的治疗效果。研究表明,本研究所得到的Fr.B具有广谱的抗菌增效活性,能够增强红霉素对伤口耐药菌感染的治疗效果。该研究结果为克服微生物对抗生素的耐药性提供了新的思路和解决方案。     

High-level expression of recombinant phospholipase C from Bacillus cereus in Pichia pastoris and its characterization

The phospholipase c (plc) gene from Bacillus cereus was cloned into the pPICZC vector and integrated into the genome of Pichia pastoris. The phospholipase C (PLC) when expressed in P. pastoris was fused to the -factor secretion signal peptide of Saccharomyces cerevisiae and secreted into a culture medium. Recombinant P. pastoris X-33 had a clear PLC band at 28.5 kDa and produced an extracellular PLC with an activity of 678 U mg^–1 protein which was more than a recombinant P. pastoris GS115 (552 U mg^–1 protein) or KM71H (539 U mg^–1 protein). The PLCs were purified using a HiTrap affinity column with a specific activity of 1335 U mg^–1 protein by P. pastoris GS115, 1176 U mg^–1 protein by P. pastoris KM71H and 1522 U mg^–1 protein by P. pastoris X-33. The three recombinant PLCs had high PLC activity in the low pH range of 4-5 and higher thermal stability (e.g. stable at 75 °C) than the wild-type PLC from B. cereus. Some organic solvents, surfactants and metal ions, e.g. methanol, acetone, Co²⁺ and Mn²⁺ etc., also influenced the activity of the recombinant PLCs.     

Synergistic hepatoprotective effect of Schisandrae lignans with Astragalus polysaccharides on chronic liver injury in rats

The aim of this study was to investigate the synergistic hepatoprotective effect of lignans from Fructus Schisandrae chinensis (LFS) with Astragalus polysaccharides (APS) on chronic liver injury in male Sprague-Dawley rats. Subcutaneous injection of 10% CCl₄ twice a week for 3 months resulted in significantly (p<0.001) elevated serum alanine aminotransferase (ALT), asparate aminotransferase (AST), alkaline phosphatase (ALP) activities compared to controls. In the liver, significantly elevated levels (p<0.001) of malondialdehyde (MDA), lowered levels of reduced glutathione (GSH) (p<0.05) and catalase (CAT) (p<0.001), superoxide dismutase (SOD) (p<0.01)were observed following CCl₄ administration. ‘LFS+ASP’ treatment of rats at doses of ‘LFS (45 mg/kg)+APS (150 mg/kg)’ and ‘LFS (135 mg/kg)+APS (450 mg/kg)’ displayed hepatoprotective and antioxidative effects than the administration of either LFS or APS, as evident by lower (p<0.005 or 0.001) levels of serum ALT, AST, ALP and hepatic MDA (p<0.001) concentration, as well as higher SOD (p<0.05 or 0.005), CAT activities(p<0.01 or 0.005), GSH concentration (p<0.05 or 0.005) compared to the toxin treated group. Histopathological examinations revealed severe fatty degeneration in the toxin group, and mild damage in groups treated with ‘LFS+APS’ were observed. The coefficients drug interaction (CDI) between each individual drug and their combination (at the same dose of their single treatment) of these foregoing parameters were all less than 1, indicating that LFS and APS display hepatoprotective and antioxidant properties and act in a synergistic manner in CCl₄ induced liver injury in rats.     

Noninvasive assessment of cardiac contractility by using (dP/dt)/P of carotid artery pulses during exercise

In earlier studies we have shown that both the pressure (P) of the carotid artery pulse (CAP) and its first derivative (CAP dP/dt) could be recorded during moderate exercise. To establish that the CAP (dP/dt)/P is a noninvasive substitute for the left ventricular (LV) value, LV (dP/dt)/P, an index of cardiac contractility, we studied CAP (dP/dt)/P under various states of activity in the autonomic nervous system in 12 healthy male subjects. Increased sympathetic nerve activities yielded by passive tilting, emotional load, or cold stress increased CAP (dP/dt)/P significantly (P< 0.05). Increased parasympathetic nerve activity by ocular compression, however, did not significantly affect the value. Moderate exercise at a heart rate of approximately 150 beats·min^–1 increased it significantly from 16.7 to 25.2·s^–1 in a supine position (P<0.001) and from 16.6 to 24.8·s^–1 in an upright position (P<0.001). It increased monotonically as heart rate increased, but the slope was steeper when the heart rate was greater than approximately 100 beats·min^–1 than it was when the rate was less than 100 beats·min^–1. In conclusion, the present study indicated that CAP (dP/dt)/P can be used as a noninvasive index of cardiac contractility even in moderate exercise.     

Comparative toxicity of Pfiesteria spp., prolonging toxicity of P. piscicida in culture and evaluation of toxin(s) stability

Toxicity of Pfiesteria piscicida (strain CAAE #2200) in the presence of fish (juvenile hybrid tilapia, Oreochromis sp., total length 3–6 cm) has been maintained in the laboratory for 19 months by serial transfer of toxic cells using a modified maintenance protocol. Toxicity was re-induced when toxin-producing P. piscicida cells were separated from fish and cultured on algal prey for 50 days and then re-introduced to new tanks containing fish. We confirmed toxicity in a strain of P. shumwayae (strain CAAE #101272). Toxicity to fish was demonstrated in culture filtrates (0.2 μm) derived from cultures of both Pfiesteria spp., however, it was markedly reduced in comparison to unfiltered water. Filtrates retained toxic activity when stored at −20 °C for up to 6 months. Toxicity to fish was retained when filtrates were held at room temperature for 48 h, at 70 °C for 30 min or at 88–92 °C for 2 h. P. piscicida killed all finfish species tested. Grass shrimp (Paleomonetes pugio; adult 2–3 cm), blue crab (Callinectes sapidus; juvenile 4–7 cm) and brine shrimp (Artemia sp.; 18–24 h post-hatch) were unaffected by concentrations of toxin(s) that killed juvenile tilapia in 4–24 h. Ichthyotoxic activity of filtrates from fish-killing cultures and stability of the toxic activity were similar among P. piscicida and P. shumwayae. These results confirm previously reported observations on toxicity of P. piscicidaand P. shumwayae to finfish. We have maintained toxicity in the laboratory for longer periods than have previously been routinely achieved, and we have demonstrated that the toxic activity is heat stable. In contrast to previous studies with other toxic P. piscicida strains, we did not observe toxic activity to blue crabs or other crustaceans.     

Rainbow trout (Oncorhynchus mykiss) recombinant IL-1β and derived peptides induce migration of head-kidney leucocytes in vitro

The present work provides the first information concerning the chemoattractant activity of trout recombinant IL-1β and its derived peptides, referred to as P1, P2 and P3. The predicted rainbow trout mature interleukin-1β peptide was produced as a recombinant protein in Escherichia coli. The first peptide, P1, corresponded to fragment 146–157 (YVTPVPIETEAR) of the trout sequence and had an MW of 1·37 kDa. It was equivalent to a region known to be part of the receptor binding domain from the mammalian crystal structure of IL-1β complexed to its receptor. P2 was used as control peptide, consisting of the same 12 amino acids as P1, but arranged in a random sequence (VVEEYIRAPPTT). P3 was synthesised to complex with an adjacent region of the IL-1 receptor, and corresponded to fragment 207–216 (YRRNTGVDIS) of the trout sequence, with an MW of 1·18 kDa. Migration was stimulated when leucocytes were exposed to concentrations of ≥10 ng ml⁻¹rIL-1β. Peptide P3 also induced leucocyte migration, with an optimal dose of 0·25 mm being recorded. While P1 had no effect on cell migration when used alone, synergism was evident as a consequence of combining P1 with a suboptimal dose (0·01 mm) of P3. No synergism occurred when cells were exposed to a combination of P3 and the control peptide P2.     

Chaetosphaeria tortuosa, the newly discovered teleomorph of Menispora tortuosa, with a key to known Menispora species

Chaetosphaeria tortuosa is described as the newly discovered teleomorph of Menispora tortuosa, based on specimens from Canada and the Czech Republic, and single spore isolations from both morphs. The fungus produces superficial, more or less globose, papillate, dark brown to black smooth perithecia (200–)220–250 × (220–)230–260 μm. The asci are unitunicate, 8-spored, cylindrical-fusiform, (110–)120–133(–145) × 12–14 with a distinct apical, nonamyloid annulus 1–1.5 μm high, 3.5–4 μm wide. The ascospores are fusiform, 19–24 × 5–6 μm, hyaline, 3-septate, smooth, and 2-seriate in the ascus. The morphology of the teleomorph and anamorph are similar to that of C. ovoidea (anamorph: M. glauca), differing in dimensions of asci and ascospores, and in the disposition and morphology of the phialides of the anamorphs. The generic concept and phylogeny of Menispora is briefly discussed, and a key to the 11 species currently accepted in the genus is provided.     

A multi-component upstream activation sequence of the Saccharomyces cerevisiae glyceraldehyde-3-phosphate dehydrogenase gene promoter

Summary The majority of the activation potential of the Saccharomyces cerevisiae TDH3 gene promoter is contained within nucleotides –676 to –381 (relative to the translation initiation codon). An upstream activation sequence (UAS) in this region has been characterized by in vitro and in vivo assays and demonstrated to be composed of two small, adjacent DNA sequence elements. The essential determinant of this upstream UAS is a general regulatory factor 1 (GRF1) binding site at nucleotides –513 to –501. A synthetic DNA element comprising this sequence, or an analogue in which two of the degenerate nucleotides of the GRF1 site consensus sequence were altered, activated 5 deleted TDH3 and CYC1 promoters. The second DNA element of the UAS is a 7 by sequence which is conserved in the promoters of several yeast genes encoding glycolytic enzymes and occurs at positions –486 to –480 of the TDH3 promoter. This DNA sequence represents a novel promoter element: it contains no UAS activity itself, yet potentiates the activity of a GRF1 UAS. The potentiation of the GRFl UAS by this element occurs when placed upstream from the TATA box of either the TDH3 or CYC1 promoters. The characteristics of this element (termed GPE for GRF1 site potentiator element) indicate that it represents a binding site for a different yeast protein which increases the promoter activation mediated by the GRF1 protein. Site-specific deletion and promoter reconstruction experiments suggest that the entire activation potential of the –676 to –381 region of the TDH3 gene promoter may be accounted for by a combination of the GRF1 site and the GPE.     

一株嗜麦芽窄食单胞菌裂解性噬菌体的分离及生物学特性

【背景】嗜麦芽窄食单胞菌是一种广泛存在于医院和自然环境中的条件致病菌,其分离率与耐药率逐年增加。噬菌体是一类能特异性感染并杀灭细菌的病毒。【目的】分离一株新型嗜麦芽窄食单胞菌噬菌体,为临床嗜麦芽窄食单胞菌感染及防控提供补充手段。【方法】以临床分离的嗜麦芽窄食单胞菌为宿主菌,用点板法从医院污水中分离鉴定噬菌体;用双层平板法测定噬菌体效价及一步生长曲线等生物学特性;用透射电镜观察噬菌体形态;提取噬菌体基因组DNA进行全基因组测序,拼接噬菌体基因组并进行注释。【结果】分离到一株嗜麦芽窄食单胞菌裂解性噬菌体,命名为v B＿Sma S＿P11。该噬菌体感染宿主菌的潜伏期小于5 min,快速增殖60 min后达到平稳期,暴发量为100 PFU/cell。透射电镜观察该噬菌体为长尾噬菌体,具有典型的二十面体头和不可收缩的尾部。基因组测序结果表明,该噬菌体基因组全长44 600 bp,GC含量为63.7%,无抗生素耐受基因、毒力基因和t RNA,与NCBI数据库中所有已知嗜麦芽窄食单胞菌噬菌体相比同源性很低。基因组注释显示该噬菌体含有66个开放阅读框(open reading frame,ORF),其...     

Partial In Vitro Digestion of Active Gliadin-Related Peptides in Celiac Disease

Peptides corresponding to residues 75–86 (RPQQPYPQPQPQ) and 75–85 of the A-gliadin structure, which were shown to be active in an animal model of celiac disease, were digested in vitro with small intestinal mucosa from children with celiac disease in remission and with mucosa from normal children. The products of digestion were separated into two fractions by gel permeation chromatography. Undigested residues (M _r > 400 fraction) from both peptides contained mainly glutamine, proline, and tyrosine, while the digested materials (M _r < 400 fraction) contained mainly proline, glutamine and arginine. Much larger amounts of undigested peptides were obtained from digestion with celiac mucosa than from normal mucosa. The results with peptide 75–86 indicated that the octapeptide 77–84 (QQPYPQPQ) was the main residual component and this peptide was shown to be active in the assay. Peptide 77–84 was also obtained as a residue from digestion of peptide 75–85, together with heptapeptide 77–83. The results lend further support for a primary mucosal defect in celiac disease and indicate that residual peptides in the small intestine of patients with the disease still retain appreciable toxicity.     

Antagonism of Aeromonas hydrophila by propolis and its effect on the performance of Nile tilapia, Oreochromis niloticus

Propolis, a resinous substance collected by Apis mellifera bees from various plant sources and mixed with secreted beeswax, is a multifunctional material used by bees in the construction, maintenance, and protection of their hives. The collected propolis sample, from High Egypt, was dark-green with olive-odor. The minimal inhibition concentration (MIC) of propolis-ethanolic-extract, against Aeromonas hydrophila, was 80 μg Propolis-ethanolic-extract and crude propolis (1%) were added to artificial basal diet with (30% crude protein) to evaluate their efficacy on the fish growth-performance, immunostimulation and resistance to A. hydrophila. Two hundred and twenty-five Oreochromis niloticus (8 ± 0.45 g/fish) were divided into three equal treatments (T) of triplet replicates. The fish of T₁ were fed on basal diet (control). The fish of T₂ were given the basal diet, containing propolis-ethanolic-extract. The fish of T₃ were given the basal diet containing crude propolis for 28 day. The fish were intraperitoneally challenged by A. hydrophila (0.2 × 10⁷ cells ml⁻¹) at the end of the feeding period and kept for 15 more days.The best growth rate and feed conversion ratio were obtained with T_2. The increase in the average daily gain, specific growth rate and feed efficiency ratio were highly significances in T₂ followed by T₃ when compared with the control group. The HCT-level and monocyte-counts were increased (T₂). No significant change, in the large lymphocytic-count was found among the three treatments (28–27–28%), while the neutrophil-count was significantly decreased (7%) with T₂ and increased (13.11%) with the control. A significant increase in serum lysozyme and serum bactericidal activities was found with T₂. The RLP against A. hydrophila was high with T₂ and T₃.The propolis-ethanolic-extract enhanced the growth, immunity and resistance of O. niloticus against A. hydrophila more than the crude propolis.     

Determination of Homalodisca coagulata (Hemiptera: Cicadellidae) egg ages suitable for oviposition by Gonatocerus ashmeadi, Gonatocerus triguttatus, and Gonatocerus fasciatus (Hymenoptera: Mymaridae)

Homalodisca coagulata (Say) (Hemiptera: Cicadellidae) eggs 1–10 days of age were exposed to Gonatocerus ashmeadi Girault, Gonatocerus triguttatus Girault, and Gonatocerus fasciatus Girault (all Hymenoptera: Mymaridae) in no choice laboratory trials to investigate egg age utilization and to determine which egg ages are vulnerable to attack by these three parasitoids. The H. coagulata egg ages that were most suitable for oviposition by G. ashmeadi, G. triguttatus, and G. fasciatus were eggs 3, 4, and 2 days of age, respectively. Egg ages least suitable for parasitoid development were 6–10 days for G. ashmeadi (resulting in <50% parasitism), 1–2 and 7–10 days for G. triguttatus (resulting in <25% parasitism), and 3–10 days for G. fasciatus (resulting in <11% parasitism). Pooling parasitism data across all egg ages showed that parasitism by G. ashmeadi was 12.9 and 28.5% higher compared with G. triguttatus and G. fasciatus, respectively, and G. triguttatus resulted in 15.6% higher percentage parasitism compared with G. fasciatus. Egg age had a significant effect on the percentage of female G. ashmeadi offspring produced, but this was not significant for G. triguttatus, and low G. fasciatus parasitism prevented statistical analyses for comparisons. Results from tests where females were offered a choice for oviposition between eggs 1, 3, and 5 days of age demonstrated that G. ashmeadi and G. triguttatus showed no significant oviposition preference, while percentage parasitism by G. fasciatus was 29.4 and 7.4% higher when females were presented eggs 1 and 3 days of age, respectively, compared with eggs 5 days of age. Choice tests indicated that an overlap in egg age suitability for oviposition exists between G. ashmeadi, G. triguttatus, and G. fasciatus, and that interspecific competition for eggs 1, 2, and 3 days of age may occur in the field environment.     

Synergistic anti-proliferative effects of vitamin D derivatives and 9-cis retinoic acid in SH-SY5Y human neuroblastoma cells

This study examines the effect of 1,25-dihydroxyvitamin D₃ [1,25(OH)₂D₃], 24,25-dihydroxyvitamin D₃ [24,25(OH)₂D₃], two vitamin D analogues (KH 1060 and EB 1089, which are 20-epi-22-oxa and 22,24-diene-analogues, respectively), 9-cis retinoic acid and all-trans retinoic acid on proliferation of SH-SY5Y human neuroblastoma cells, after treatment for 7 days. Cell number did not change when the cells were incubated with 1, 10 or 100 nM 1,25(OH)₂D₃ or its derivatives, but significantly decreased in the presence of the two retinoids (0.001–10 μM final concentration). A synergistic inhibition was observed, when SH-SY5Y cells were treated combining 0.1 μM 9-cis retinoic acid and 10 nM 1,25(OH)₂D₃ or 10 nM KH 1060, and 1 μM 9-cis retinoic acid and 10 nM 1,25(OH)₂D₃ or 10 nM EB 1089. Acetylcholinesterase activity showed a significant increase, in comparison with controls, after treatment of the cells for 7 days with 0.1 or 1 μM 9-cis retinoic acid, alone or combined with 10 nM 1,25(OH)₂D₃ or 10 nM KH 1060 or 10 nM EB 1089. This increase was synergistic, combining 1 μM 9-cis retinoic acid and 10 nM 1,25(OH)₂D₃ or EB 1089. The levels of the c-myc encoded protein remarkably decreased after treatment of SH-SY5Y cells for 1, 3, 7 days with 0.1 and 1 μM 9-cis retinoic acid, alone or combined with 10 nM 1,25(OH)₂D₃ or 10 nM KH 1060 or 10 nM EB 1089. In particular, the association of 1 μM 9-cis retinoic acid and 10 nM 1,25(OH)₂D₃ or 10 nM EB 1089 resulted in a synergistic c-myc inhibition, in comparison with that obtained in the presence of the retinoid alone. These findings may have therapeutic implications in human neuroblastoma.     

Evaluation of entomopathogenic nematodes against the olive fruit fly, Bactrocera oleae (Diptera: Tephritidae)

Infectivity of six entomopathogenic nematode (EPNs) species against Bactrocera oleae was compared. Similar infection levels were observed when third-instar larvae were exposed to infective juveniles (IJs) on a sand-potting soil substrate. When IJs were sprayed over naturally infested fallen olives, many larvae died within treated olives as well as in the soil; Steinernema feltiae caused the highest overall mortality of 67.9%. In addition, three laboratory experiments were conducted to optimize a time period for S. feltiae field application. (1) Abundance of fly larvae inside fallen olives was estimated over the 2006–2007 season with the highest number of susceptible larvae (3 mm and larger) per 100 olives being observed during December, 2006. (2) S. feltiae efficacy against fly larvae dropped to the soil post-IJ-application was determined. B. oleae added to the substrate before and after nematode application were infected at similar levels. (3) Effect of three temperature regimes (min–max: 10–27, 6–18, and 3–12 °C) corresponding to October through December in Davis, California on S. feltiae survival and infectivity was determined. After 8 weeks, the IJs at the 3–12 °C treatment showed the highest survival rate. However, the cold temperature significantly limited S. feltiae infectivity. Our results demonstrate that B. oleae mature larvae are susceptible to EPN infection both in the soil and within infested olives. Being the most effective species, S. feltiae may have the potential to suppress overwintering populations of B. oleae. We suggest that November is the optimal time for S. feltiae field application in Northern California.     

Successful oviposition and reproductive biology of Aprostocetus vaquitarum (Hymenoptera: Eulophidae): A predator of Diaprepes abbreviatus (Coleoptera: Curculionidae)

Aprostocetus vaquitarum (Wolcott) causes 78–91 percent mortality to eggs of Diaprepes abbreviatus (L.), under field conditions in southern Florida. In the laboratory, A. vaquitarum was reared on D. abbreviatus eggs at 25 °C, a photoperiod of 12:12 (L:D) and with abundant hosts, A. vaquitarum adult females lived around 15 days. Oviposition was significantly affected by the age of the host egg mass. Egg masses aged 0- to 3-day-old were accepted significantly better than those aged 4–6 days. The mean number of eggs deposited per female was around 53, with extreme values of 124 and 19 eggs per female. Using these data in combination with the sex ratio observed in the field (0.16) and the duration of the preimaginal stages, r_m (0.168–0142 day⁻¹), T (22.39–22.89 days), and R₀ (43.03–25.81 females per female) were calculated.