Reference Gene Selection for Insect Expression Studies Using Quantitative Real-Time PCR: The Head of the Honeybee, Apis mellifera, After a Bacterial Challenge

In this study an important and often neglected aspect of gene expression studies in insects, the validation of appropriate reference genes with stable expression levels between sample groups, is addressed. Although in this paper the reference gene selection for the honeybee, Apis mellifera L. (Hymenoptera: Apidae) head was tested in the context of bacterial challenge with Escherichia coli, this work can serve as a resource to help select and screen insect reference genes for gene expression studies in any tissue and under any experimental manipulation. Since it is recommended to use multiple reference genes for accurate normalization, we analyzed the expression of eleven candidate reference genes in the honeybee head, for their potential use in the analysis of differential gene expression following bacterial challenge. Three software programs, BestKeeper, Normfinder and geNorm, were used to assess candidate reference genes. GeNorm recommended the use of four reference genes. Both geNorm and Normfinder identified the genes GAPDH, RPS18, actin and RPL13a as the most stable ones, only differing in their ranking order. BestKeeper identified RPS18 as being the reference gene with the least overall variation, but also actin and GAPDH were found to be the second and third most stable expressed gene. By a combination of three software programs the genes actin, RPS18 and GAPDH were found suitable reference genes in the honeybee head in the context of bacterial infection

Index de gravité de la fonction ventilatoire. A propos de 380 cas de traumatismes thoraciques graves

peer reviewe

Measurement of endocardial viability ratio (E.V.R.) during anesthesia for cardiac surgery

peer reviewedSince sub-endocardial ischemia is the consequence of a discrepancy between the blood demand and supply of oxygen at this level, the study of the myocardial performance by the measurement of the endocardial viability ratio (E.V.R.) is both useful and possible during anesthesia. E.V.R. is the ratio between the oxygen supply and demand of the myocardium. It is equal to the diastolic pressure time index (D.P.T.I.) over the tension time index (T.T.I.). Measurements are made at different times, by means of the arterial pressure and the left atrial pressure, as well as with the Datascope-E.V.R. Computer. During gradual morphine administration (0.5-1-1.5 mg/kg) and if no major surgical stress occurs, E.V.R. remains excellent and stable (1.46 - 1.48 - 1.43). It deteriorates more or less (1.29 - 1.09) during tachycardia or hypertension. Within the hour following the end of extracorporeal circulation, E.V.R. significantly improves (1.04 - 1.06 - 1.09 - 1.23). Although E.V.R. measurement is easy during cardiac surgery, it is impossible to carry out in case of arrhythmia. While morphine anesthesia induces no variation in E.V.R., tachycardia or hypertension require the addition of therapeutic drug. Within one hour following the end of extra-corporeal circulation, E.V.R. measurement shows improved endocardial viability, although the hemodynamic parameters undergo no significant change

Effet de drogues utilisées en anesthésie sur l'index de viabilité de l'endocarde (EVR ou "endocardial viability ratio") en chirurgie cardiaque

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Characterization of Native Honey Bee Subspecies in Republic of Benin Using Morphometric and Genetic Tools

Morphometric characteristics combined with genetic markers are powerful tools used for determining honey bee subspecies. Bees samples collected from 94 established apiaries distributed throughout all of the Republic of Benin were morphometricaly characterized using seven parameters and the COI-COII regions of mitochondrial DNA were sequenced. Based on the morphometric data the native honey bees could be divided into three distinct ecotypes - the Benino-dry-tropical-ecotype in the north, the Benino-Sudanian-ecotype in the central part and the Benino-Sudano-Guinean-ecotype in the south. The DNA COI-COII regions sequence analyses confirmed that the honey bee population of the Republic of Benin belongs to different mitotypes but do not correspond with the determined ecotypes. We could determine three new haplotypes which missed the P0 segment but the Q region was duplicated or triplicated. Phylogenetic analyses clustered them together in the A evolutionary lineage. In conclusion, morphometric and genetic analysis of the native West African honey bees indicated that each of the different mitotypes was able to adapt to the different ecological conditions in the country by morphometric adjustments

Nationwide Screening for Bee Viruses and Parasites in Belgian Honey Bees.

The health of honey bees is threatened by multiple factors, including viruses and parasites. We screened 557 honey bee (Apis mellifera) colonies from 155 beekeepers distributed all over Belgium to determine the prevalence of seven widespread viruses and two parasites (Varroa sp. and Nosema sp.). Deformed wing virus B (DWV-B), black queen cell virus (BQCV), and sacbrood virus (SBV) were highly prevalent and detected by real-time RT-PCR in more than 95% of the colonies. Acute bee paralysis virus (ABPV), chronic bee paralysis virus (CBPV) and deformed wing virus A (DWV-A) were prevalent to a lower extent (between 18 and 29%). Most viruses were only present at low or moderate viral loads. Nevertheless, about 50% of the colonies harbored at least one virus at high viral load (>10(7) genome copies/bee). Varroa mites and Nosema sp. were found in 81.5% and 59.7% of the honey bee colonies, respectively, and all Nosema were identified as Nosema ceranae by real time PCR. Interestingly, we found a significant correlation between the number of Varroa mites and DWV-B viral load. To determine the combined effect of these and other factors on honey bee health in Belgium, a follow up of colonies over multiple years is necessary

Molecular cloning and expression of icarapin, a novel IgE-binding bee venom protein

AbstractThe 1045bp full-length cDNA sequence of a new bee venom component was obtained by rapid amplification of cDNA ends. The 672bp coding sequence corresponds to a protein with a signal peptide and multiple carbohydrate binding sites, and it was named icarapin. It has the new consensus sequence N-[TS]-T-S-[TV]-x-K-[VI](2)-[DN]-G-H-x-V-x-I-N-[ED]-T-x-Y-x-[DHK]-x(2,6)- [STA]-[VLFI]-x-[KR]-V-R-[VLI]-[IV]-[DN]-V-x-P. At least two transcript variants were found. Recombinant icarapin was tested for recognition by IgE antibodies and gave a positive dot blot with sera from 4 out of 5 bee venom allergic patients, all beekeepers. Indirect immunofluorescent staining localized the protein in the cuticular lining of the venom duct