Screening and secretomic analysis of enthomopatogenic Beauveria bassiana isolates in response to cowpea weevil (Callosobruchus maculatus) exoskeleton

The production of cowpea (Vigna unguiculata), an important self-sustained crop in Latin America and Africa, is severely affected by damage by the cowpea weevil Callosobruchus maculatus. The presence of a single larva in stored seeds can lead to losses of almost 40%. Control of C. maculatus currently relies on the inefficient use of chemical insecticides and post-harvest treatments. The use of entomopathogenic fungus became a reliable alternative for coleopteran pest control and has been extensively investigated. Among them, Beauveria bassiana and Metarhizium anisopliae were widely evaluated in order to measure their virulence toward many insects. In this report, we evaluated the insecticidal activity of ten strains of B. bassiana and the most lethal fungi strains were analyzed for proteinaceous secretions by two dimensional electrophoresis and for enzyme activities, including chitinolytic, proteolytic and alpha-amylolytic activities. This study could, in the near future, help to establish novel biotechnological tools to use for cowpea weevil control.     

PER-TIM interactions with the photoreceptor cryptochrome mediate circadian temperature responses in Drosophila

Drosophila cryptochrome (CRY) is a key circadian photoreceptor that interacts with the period and timeless proteins (PER and TIM) in a light-dependent manner. We show here that a heat pulse also mediates this interaction, and heat-induced phase shifts are severely reduced in the cryptochrome loss-of-function mutant cry^b. The period mutant per^L manifests a comparable CRY dependence and dramatically enhanced temperature sensitivity of biochemical interactions and behavioral phase shifting. Remarkably, CRY is also critical for most of the abnormal temperature compensation of per^L flies, because a per^L; cry^b strain manifests nearly normal temperature compensation. Finally, light and temperature act together to affect rhythms in wild-type flies. The results indicate a role for CRY in circadian temperature as well as light regulation and suggest that these two features of the external 24-h cycle normally act together to dictate circadian phase.     

Horizontal transmission of begomoviruses between Bemisia tabaci biotypes

We have previously shown that the monopartite Tomato yellow leaf curl virus (TYLCV), a begomovirus (family Geminiviridae, genus Begomovirus) infecting tomato plants can be transmitted in a gender-dependent manner among its insect vector the whitefly Bemisia tabaci type B (Gennaduis) (Aleyrodidae: Hemiptera) during mating. Viruliferous females were able to transmit the virus to non-viruliferous males and vice versa, in the absence of any other virus source. The recipient insects were able to infect tomato plants. In this communication, we present evidence that two bipartite begomoviruses infecting cucurbits, Squash leaf curl virus (SLCV) and Watermelon chlorotic stunt virus (WmCSV) can be transmitted in a gender-dependent manner among whiteflies. In addition we show that TYLCV can be transmitted during mating among individuals from the same biotype (from B-males to B-females and vice versa; and from Q-males to Q-females and vice versa). However, viruliferous males of the B biotype are unable to transmit the virus to females of the Q biotype (and vice versa); similarly, viruliferous males of the Q biotype are unable to transmit the virus to females of the B biotype (and vice versa). These findings support the hypothesis that a pre-zygotic mating barrier between the Q and B biotypes is the cause for the absence of gene flow between the two biotypes, and that virus transmission can be used as a marker for inter-biotype mating. To be transmitted during mating, the virus needs to be present in the haemolymph of the donor insect. Abutilon mosaic virus (AbMV), a bipartite begomovirus that can be ingested but not transmitted by B. tabaci, is absent in the whitefly haemolymph, and cannot be transmitted during mating. Mating was a precondition for horizontal virus transfer from male to female, or female to male. Virus was not transmitted when viruliferous B. tabaci were caged with the non-vector non-viruliferous whitefly Trialeurodes vaporariorum (Westwood) (Aleyrodidae: Hemiptera) and vice versa.     

Ligand selectivity of soluble guanylyl cyclase: effect of the hydrogen-bonding tyrosine in the distal heme pocket on binding of oxygen, nitric oxide, and carbon monoxide

Although soluble guanylyl cyclase (sGC) functions in an environment in which O(2), NO, and CO are potential ligands for its heme moiety, the enzyme displays a high affinity for only its physiological ligand, NO, but has a limited affinity for CO and no affinity for O(2). Recent studies of a truncated version of the sGC beta(1)-subunit containing the heme-binding domain (Boon, E. M., Huang, S H., and Marletta, M. A. (2005) Nat. Chem. Biol., 1, 53-59) showed that introduction of the hydrogen-bonding tyrosine into the distal heme pocket changes the ligand specificity of the heme moiety and results in an oxygen-binding sGC. The hypothesis that the absence of hydrogen-bonding residues in the distal heme pocket is sufficient to provide oxygen discrimination by sGC was put forward. We tested this hypothesis in a context of a complete sGC heterodimer containing both the intact alpha(1)- and beta(1)-subunits. We found that the I145Y substitution in the full-length beta-subunit of the sGC heterodimer did not produce an oxygen-binding enzyme. However, this substitution impeded the association of NO and destabilized the NO.heme complex. The tyrosine in the distal heme pocket also impeded both the binding and dissociation of the CO ligand. We propose that the mechanism of oxygen exclusion by sGC not only involves the lack of hydrogen bonding in the distal heme pocket, but also depends on structural elements from other domains of sGC.     

Thermotolerance and gene expression following heat stress in the whitefly Bemisia tabaci B and Q biotypes

The whitefly Bemisia tabaci (Gennadius) causes tremendous losses to agriculture by direct feeding on plants and by vectoring several families of plant viruses. The B. tabaci species complex comprises over 10 genetic groups (biotypes) that are well defined by DNA markers and biological characteristics. B and Q are amongst the most dominant and damaging biotypes, differing considerably in fecundity, host range, insecticide resistance, virus vectoriality, and the symbiotic bacteria they harbor. We used a spotted B. tabaci cDNA microarray to compare the expression patterns of 6000 ESTs of B and Q biotypes under standard 25 °C regime and heat stress at 40 °C. Overall, the number of genes affected by increasing temperature in the two biotypes was similar. Gene expression under 25 °C normal rearing temperature showed clear differences between the two biotypes: B exhibited higher expression of mitochondrial genes, and lower cytoskeleton, heat-shock and stress-related genes, compared to Q. Exposing B biotype whiteflies to heat stress was accompanied by rapid alteration of gene expression. For the first time, the results here present differences in gene expression between very closely related and sympatric B. tabaci biotypes, and suggest that these clear-cut differences are due to better adaptation of one biotype over another and might eventually lead to changes in the local and global distribution of both biotypes.