Genetic markers for the resistance of honey bee to Varroa destructor.

In the mid-20th century, the first case of infection of European bees Apis mellifera L. with the ectoparasite mite Varroa destructor was recorded. The original host of this mite is the Asian bee Apis cerana. The mite V. destructor was widespread throughout Europe, North and South America, and Australia remained the only continent free from this parasite. Without acaricide treatment any honeybee colony dies within 1-4 years. The use of synthetic acaricides has not justified itself - they make beekeeping products unsuitable and mites develop resistance to them, which forces the use of even greater concentrations that can be toxic to the bees. Therefore, the only safe measure to combat the mite is the use of biological control methods. One of these methods is the selection of bee colonies with natural mite resistance. In this article we summarize publications devoted to the search for genetic markers associated with resistance to V. destructor. The first part discusses the basic mechanisms of bee resistance (Varroa sensitive hygienic behavior and grooming) and methods for their assessment. The second part focuses on research aimed at searching for loci and candidate genes associated with resistance to varroosis by mapping quantitative traits loci and genome-wide association studies. The third part summarizes studies of the transcriptome profile of Varroa resistant bees. The last part discusses the most likely candidate genes - potential markers for breeding Varroa resistant bees. Resistance to the mite is manifested in a variety of phenotypes and is under polygenic control. The establishing of gene pathways involved in resistance to Varroa will help create a methodological basis for the selection of Varroa resistant honeybee colonies.

[New Approach to the Mitotype Classification in Black Honeybee Apis mellifera mellifera and Iberian Honeybee Apis mellifera iberiensis].

The black honeybee Apis mellifera mellifera L. is today the only subspecies of honeybee which is suitable for commercial breeding in the climatic conditions of Northern Europe with long cold winters. The main problem of the black honeybee in Russia and European countries is the preservation of the indigenous gene pool purity, which is lost as a result of hybridization with subspecies, A. m. caucasica, A. m. carnica, A. m. carpatica, and A. m. armeniaca, introduced from southern regions. Genetic identification of the subspecies will reduce the extent of hybridization and provide the gene pool conservation of the black honeybee. Modern classification of the honeybee mitotypes is mainly based on the combined use ofthe DraI restriction endonuclease recognition site polymorphism and sequence polymorphism of the mtDNA COI-COII region. We performed a comparative analysis of the mtDNA COI-COII region sequence polymorphism in the honeybees ofthe evolutionary lineage M from Ural and West European populations of black honeybee A. m. mellifera and Spanish bee A. m. iberiensis. A new approach to the classification of the honeybee M mitotypes was suggested. Using this approach and on the basis of the seven most informative SNPs of the mtDNA COI-COII region, eight honeybee mitotype groups were identified. In addition, it is suggested that this approach will simplify the previously proposed complicated mitotype classification and will make it possible to assess the level of the mitotype diversity and to identify the mitotypes that are the most valuable for the honeybee breeding and rearing.

[The effect of high-molecular weight chitosans on the antioxidant and immune systems of the honeybee].

High-molecular weight chitosan (200 kDa, 75% deacetylated) and N-succinoyl chitosan (300 kDa, 75% deacetylated) were shown to have a preadaptive effect and increase the lifespan of honeybees due to the induction of protective antioxidant and immune mechanisms. Chitosan with a molecular weight of 200 kDa had a fungistatic effect on a pathogenic fungus that causes ascospherosis, a disease of bee larvae and pupae.

[Molecular genetic analysis of five extant reserves of black honeybee Apis melifera melifera in the Urals and the Volga region].

Local populations of the black honeybee Apis mellifera mellifera from the Urals and the Volga region were examined in comparison with local populations of southern honeybee subspecies A. m. caucasica and A. m. carpatica from the Caucasus and the Carpathians. Genetic analysis was performed on the basis of the polymorphism of nine microsatellite loci of nuclear DNA and the mtDNA COI­COII locus. On the territory of the Urals and the Volga region, five extant populations (reserves) of the black honeybee A. m. mellifera were identified, including the Burzyanskaya, Tatyshlinskaya, Yuzhno-Prikamskaya, Visherskaya, and Kambarskaya populations. These five populations are the basis of the modern gene pool of the black honeybee A. m. mellifera from the Urals and the Volga region. The greatest proportion of the remaining indigenous gene pool of A. m. mellifera (the core of the gene pool of the population of A. m. mellifera) is distributed over the entire territory of Perm krai and the north of the Republic of Bashkortostan. For the population of A. m. mellifera from the Urals and the Volga region, the genetic standards were calculated, which will be useful for future population studies of honeybees.

[Seven genes of mitochondrial genome enabling differentiation of honeybee subspecies Apis mellifera].

On the basis of comparative sequence analysis of 12 honeybee mitochondrial genes, seven genes enabling us to differentiate honeybees subspecies of the A, M, C, O evolutionary lineages were found. Applying comparative sequence analysis of ND2 gene mtDNA as an example on a statistically valid sample size, we showed a high level of differentiating ability of this gene and assumed that each of these seven genes probably can be used for differentiation of the subspecies within four evolutionary lineages.

[Genetic Differentiation of Local Populations of the Dark European Bee Apis mellifera mellifera L. in the Urals].

For the last two centuries, beekeepers in Russia and Europe have been introducing bees from the southern regions to the northern ones, subjecting the genetic pool of the dark European bee Apis mellifera mellifera L. subspecies to extensive hybridization. In order to reconfirm on the genetic level the previously published morphological data on the native bee population in the Urals, the Bashkortostan Republic, and the Perm Krai, we analyzed the polymorphism of the mitochondrial (mtDNA COI-COII intergenic locus) and nuclear (two microsatellite loci, ap243 and 4a110) DNA markers. Four local populations of the dark European bee A. m. mellifera surviving in the Urals have been identified, and their principal genetic characteristics have been determined. Data on the genetic structure and geographical localization of the areals of the dark European bee local populations in the Urals may be of use in restoring the damaged genetic pool of A. m. mellifera in Russia and other northern countries.

[New SNP markers of the honeybee vitellogenin gene (Vg) used for identification of subspecies Apis mellifera mellifera L].

Preservation of the gene pool of honeybee subspecies Apis mellifera mellifera is of vital importance for successful beekeeping development in the northern regions of Eurasia. An effective method of genotyping honeybee colonies used in modern science is the mapping of sites of single nucleotide polymorphism (SNP). The honeybee vitellogenin gene (Vg) encodes a protein that affects reproductive function, behavior, immunity, longevity, and social organization in the honeybee Apis mellifera and is therefore a topical research subject. The results of comparative analysis of honeybee Vg sequences show that there are 26 SNP sites that differentiate M and C evolutionary branches and can be used as markers in selective breeding, DNA-barcoding, and the creation of genetic passports for A. m. mellifera colonies.

[Analysis of the Genetic Structure of Honeybee (Apis mellifera L.) Populations].

The genetic structure of honeybee populations from the southern part of Bashkortostan was assessed based on an analysis of mtDNA (COI-COII locus) and five nuclear DNA microsatellite loci (Ap243, 4A110, A8, A113, and A28). The data indicate that the examined populations experience a deficit of heterozygotes despite intense interpedigree hybridization. It is suggested that there is a boundary between the population of Apis mellifera mellifera L. and the hybrid zone in the examined region.

[Effect of population density on enzymatic activity of antioxidative and phenol oxidase systems of imagoes and nymphs of the marble cockroach Nauphoeta cinerea].

The work deals with effect of density of population on functional activity of components pf protective system of adult individuals and nymphs of the marble cockroach. The resistance of individuals has been noted to decrease both at individual maintenance and under conditions of overpopulation. Changes in activities of enzymes of antioxidative and phenoloxidase systems are studied ion the insect hemolymph and intestine. Possible consequences of isolation and overpopulation are discussed both for stability and for individual development.

[Defensins in the honeybee antinfectious protection].

Specific conditions of the honeybee life honeybee life require the presence of effective mechanisms of antiinfectious protection whose one of the most important components are defensins--the family of antimicrobial peptides. In the honeybee, defensins are present in the form of two different peptides--defensin 1 and 2 that are similar between each other only by 55.8 %. Defensin 1 synthesized in salivary glands plays an important role in social immunity, whereas defensin 2 synthesized by cells of lat body and lymph is an important factor in the system of the honeybee individual immunity. Defensins are inducible, are controlled by interaction of Toll and Imd signal pathways and have a large specter of antimicrobial action.

[Intraspecies differences in mechanisms of formation of protective processes in the honeybee Apis mellifera].

Presented in the work are data on interrace differences in mechanisms of formation of antioxidant and phenoloxidase complexes in honeybees of the Middle Russian (Apis mellifera L.) and Caucasian (Apis mellifera caucasica Gorb.) geographic races. Different degree has been revealed of activation of these complexes depending on their localization in the insect organism and providing different strategy of adaptation to temperature stress. A diverse role of ascorbic acid in regulation of the honeybee protective system has been shown.

[Two types of metabolic regulation of phenol oxidase activity in ontogenesis of house fly].

Changes of the tyrosinase activity in ontogenesis of the house fly Musca domestica were shown to be phase-specific and ontogenetic changes of tyrosinase and dihydroxyphenylalanine oxidase activities proved to be coordinated. Ascorbic acid stimulated some ontogenetic stages of the house fly and physiological indices, such as fertility, survival at different stages, and weight of puparia. Also, ascorbic acid modulated the tyrosinase activity.

[Polymorphism of locus COI-COII of mitochondrial DNA in the honeybee Apis mellifera L. from southern Ural region]. / Polimorfizm lokusa COI-COII mitokhondrial'noi DNK medonosnoi pchely Apis mellifera L. na Iuzhnom Urale.

In spite of high biodiversity within the honeybee species Apis mellifera L., only one geographical race, the dark-colored forest honeybee A. m. mellifera, is uniquely adapted to severe environmental conditions of Eurasian forest and forest-steppe zones. Within the vast range of this race, only single isolates remain, where the dark forest honeybee is purebred. The Bashkir population is supposed to be one of these isolates. Molecular-genetic assessment of the state of the gene pool of this population revealed that southern honeybee races were introduced into the Bashkortostan Republic with great intensity, which was above the assimilation capacity of the population. The main part of the former range of the Bashkir population represents a hybrid zone with approximately equal ratio between gene pools of local and introduced honeybees. Our studies provide the possibility to single out one extant reserve of A. m. mellifera, Burzyanskii raion, in which the proportion of local bees in the gene pool is 0.98.

[Use of a PCR method for controlling pure-breeding of honeybees Apis mellifera mellifera L. in the southern Urals]. / Ispol'zovanie metoda PTsR dlia kontrolia chistoporodnosti pchelosemei Apis mellifera mellifera L. v usloviiakh iuzhnogo Urala.

A key problem of honeybee (Apis mellifera mellifera) breeding in the Southern Urals is its cross-breeding with the Caucasian honeybee Apis mellifera caucasica. Mitochondrial DNA (mtDNA) in these subspecies differ in the length of a fragment localized between genes CO-I and CO-II, which can be used as a marker. A pair of 20-mer primers for PCR was chosen by means of computer design in order to determine the fragment size in both of the subspecies. The amplified fragment was shown to have a length of 350 bp in A. m. caucasica and 600 bp in A. m. mellifera. The difference in length results from the different ratio between two main elements P and Q, which comprise a major part of this sequence in these subspecies: a copy of P element and two copies of Q element in A. m. mellifera, and a copy of Q element only in A. m. caucasica. This sharply defined distinction allows us to use PCR for differentiating the subspecies, estimating the heterogeneity in the colonies, and rejecting queens in the selection process because of the maternal inheritance of the studied character. The nucleotide sequence of the amplified mtDNA fragment of A. m. mellifera was determined.