Med je při hojeni ran použivan již od starověku. Studie prokazuji jeho antimikrobialni působeni v... more Med je při hojeni ran použivan již od starověku. Studie prokazuji jeho antimikrobialni působeni vůci sirokemu spektru bakterii i hub a objasňuji mechanizmy jeho ucinku. Nizký výskyt alergii a dobre klinicke výsledky při hojeni ran za použiti medu davaji možnost jeho sirsiho využiti.
The winter beehive debris containing bodies of honeybee parasitic mite Varroa destructor is used ... more The winter beehive debris containing bodies of honeybee parasitic mite Varroa destructor is used for veterinary diagnostics. The Varroa sucking honeybee haemolymph serves as a reservoir of pathogens including bacteria. Worker bees can pick up pathogens from the debris during cleaning activities and spread the infection to healthy bees within the colony. The aim of this study was to detect entomopathogenic bacteria in the Varroa collected from the winter beehive debris.
Zařizeni slouži k filtraci roztaveneho vceliho vosku přes vrstvu polymeru, který navaže některe t... more Zařizeni slouži k filtraci roztaveneho vceliho vosku přes vrstvu polymeru, který navaže některe typy nežadoucich molekul. Zařizeni je kovový valec s děrovaným dnem a vikem, vklada se do vyhřivaneho plastě a umožňuje řizený přetlak. Filtrat je jiman do spodni casti plastě a da se vypoustět kohoutem.
Entomopathogenic nematodes (EPNs) are obligate insect parasites that are symbiotically associated... more Entomopathogenic nematodes (EPNs) are obligate insect parasites that are symbiotically associated with specific entomopathogenic bacteria. These nematobacterial complexes are highly pathogenic to insect hosts and therefore used in biological control of insect pests. As was shown previously, EPNs can be used also as a natural infection model and a powerful tool to study insect immunity. In this study we show that also honeybee larvae are suitable hosts for nematobacterial complex that offer an excellent environment for successful development of both nematodes and their symbiotic bacteria. Here we used EPNs for evaluation of the overall immune resistance of honeybee larvae treated with potentially immuno-modulating substances (plant alcaloid sanguinarin and probiotics). Honeybee larvae were infected with 10 IJ of Heterorhabditis bacteriophora or Steinernema feltiae per larva, both carrying their symbiotic bacteria, and then scored for mortality caused by the nematobacterial infection. In comparison to untreated honeybee larvae we observed the decrease in mortality of approx. 35 % in case of H. bacteriophora and approx. 10% upon S. feltiae infection in larvae treated by selected substances. Both sanguinarin and probiotic treatment showed similar protective effect that was less significant in highly virulent S. feltiae. This is the first record that the nematobacterial infection was used for evaluation of immune status of beneficial insect. We suggest and propose this method as a valuable tool for immunity testing in honeybees as well as in other insects. Our research is supported by grant from the Ministry of Agriculture of Czech Republic (project No. QJ1210047) and by the program CZ.1.07/2.3.00/30.0009 co-financed from European Social Fund and the state budget of the Czech Republic.
Recent colony losses have led to increased interest in the research of honey bee gut microbiota. ... more Recent colony losses have led to increased interest in the research of honey bee gut microbiota. Social bees host in their digestive tract characteristic microbiota and according to recent hypotheses, this plays a significant role in their health. Within our study, two lactobacilli of the Firm-5 cluster and one Gilliamella apicola strain isolates were administered in triplicates to colonies by spraying over the brood frames, either as single strains or in combinations. Data about gut microbial diversity of young bees and larvae (using denaturating gel electrophoresis of 16S DNA V3 region), plasma antioxidant activity (TRAP) and net hive colony weight were recorded weekly for two weeks prior the experiment and six weeks after. Administration of single strains or the mixture resulted in a slightly better performance (not significant). Whereas the net weight of the control dropped by 18.7% due to lack of forage, this drop was lower in hives treated with any bacterial suspension, averaging 8.5% for all treatments. Relatively best performance was seen for the mixture. The administration, however, had no effect on antioxidative capacity of the hemolymph, which tended to decrease after the administration (with the exception of Gilliamella sp., not significant). Expectedly, administration changed the profile of lactic acid bacteria in the gut, which persisted throughout the experiment. Due to limited experiment size, we did not see any significant effect. The slight increase in performance suggests positive role, which is probably not expressed into the tested biochemical parameters (Supported by NAZV QJ1530148 and NAZV QJ1210047)
Bees are used by mankind for several thousand years, but their immune system is still far from be... more Bees are used by mankind for several thousand years, but their immune system is still far from being fully understood and moreover we still don’t have clear idea about all immune mechanisms which mediate bees’ response to their pathogens. These pathogens negatively influence life of the bees and very often even their viability causing direct impact on agriculture and industry. Detailed knowledge of bee immunity is crucial for successful fight against bee diseases; that’s why we are assembling repertoire of immune methods applicable on bees that would enable to evaluate current state of their immunity and help us in proper disease control. General ability of bees to deal with bacterial infection is well characterized by antibacterial activity of their hemolymph. Total antibacterial activity against Gram negative bacteria is detectable by real time measurement of bioluminescence produced by bacteria Escherichia coli K12; the bioluminescence signal is equal to bacterial viability which is decreased by antibacterial factors in hemolymph to at least 55 % in healthy bees. Many effectors acting against Gram positive bacteria were already identified in hemolymph of bees; as marker of their activity we selected the activity of lysozyme, a well characterized enzyme damaging peptidoglycan in cell walls. Commonly used radial zone diffusion assay turned out to be ineffective to determine the level of lysozyme, because of its low sensitivity threshold and the need for large sample volumes. Alternatively, using ELISA kit for lysozyme we determined its level in hemolymph in range from 0.1 to 15 ng/ml. Infection agents activate also stress response leading to changes in antioxidant capacity of hemolymph and activity of particular antioxidant enzymes. TRAP was used to describe total antioxidant capacity in bees and could be complemented by colorimetric measurement of superoxide dismutase and glutathione S-transferase activity.
The infrared (ATR-FTIR) and Raman spectroscopy was used for the structural characterization of ho... more The infrared (ATR-FTIR) and Raman spectroscopy was used for the structural characterization of honeybee wings. Protein, chitin, and lipid characteristic spectral features were detected using both methods. The protein secondary structure was predominantly composed of the β-sheet molecular conformation with β-turns or coil contributions. The vibration modes of the side-chain aromatic amino acid residues (tyrosine, phenylalanine, tryptophan) occurred in the wing spectra. The results of discriminant analysis showed that the infrared spectroscopy of the wing in combination with a multivariate analysis seemed promising for a resolution of the chemical structure of the wings based on lipid, proteins, and chitin content.
Instrumental insemination of Apis mellifera L. queens is a widely employed technique used in hone... more Instrumental insemination of Apis mellifera L. queens is a widely employed technique used in honeybee breeding that enables the effective control of mating. However, drone semen represents a potential source of honeybee viruses. In this study, 43 semen doses collected from apparently healthy drones, and consequently used in instrumental insemination, were analysed using PCR or RT-PCR to detect the presence of viral genome of 11 honeybee viruses. In 91% of samples, viral infection was detected. The survey revealed genomes of five viruses, namely Deformed wing virus (DWV), Acute bee paralysis virus (ABPV), Black queen cell virus (BQCV), Sacbrood virus (SBV), and A. mellifera filamentous virus (AmFV) in 84%, 19%, 14%, 2%, and 67% of samples, respectively. Single infection (30% of samples) as well as multiple infection (61% of samples) of two, three or four pathogens were also evaluated. To the best of our knowledge, this is the first study describing the presence of the BQCV and SBV ge...
Honey is composed mainly of carbohydrates which are represented by mono-, di-, tri-, tetra-, pent... more Honey is composed mainly of carbohydrates which are represented by mono-, di-, tri-, tetra-, pentasaccharides, and oligosaccharides. The content and proportions of individual carbohydrates reveal information about the origin and technological properties of honey. A total of 5 987 samples of natural honeys originating from the Czech Republic and harvested during a period of nine consecutive years were analysed to research their carbohydrate content, sum of fructose and glucose (Fru + Glc), fructose and glucose ratio (Fru/Glc), and electrical conductivity (Ec). Monosaccharides, melezitose (Mel), and Ec varied according to the source of nectar or honeydew. Sucrose (Suc) content was low 0.87 ± 1.26 g (100 g)–1 and did not exceed 15 g (100 g)–1.
In this study we measured in vitro honey bee enzyme activities in presence of the synergist piper... more In this study we measured in vitro honey bee enzyme activities in presence of the synergist piperonyl butoxide (PBO) and some new benzodioxole and dihydrobenzofuran derivatives through absorbance and fluorescence spectrophotometric assays, to determine if and to what extent detoxification systems and acetylcholinesterases are affected by these compounds. Both Italian (Italy) and Carniolan (Czech Republic) honey bees were tested. In the case of Italian honey bees the esterase activity was partially but significantly inhibited by almost all the tested products. In Carniolan honey bees only the dihydrobenzofuran derivative EN16-41 significantly inhibited esterases. Other enzymatic systems, such as cytochrome P450 monooxygenases (P450s) and glutathione S-transferases (GSTs), often involved in xenobiotics detoxification, as well as acetylcholinesterases (AchEs) seem not to be targeted by the studied synergists. These data seem to suggest that some of the investigated PBO analogues could ...
Med je při hojeni ran použivan již od starověku. Studie prokazuji jeho antimikrobialni působeni v... more Med je při hojeni ran použivan již od starověku. Studie prokazuji jeho antimikrobialni působeni vůci sirokemu spektru bakterii i hub a objasňuji mechanizmy jeho ucinku. Nizký výskyt alergii a dobre klinicke výsledky při hojeni ran za použiti medu davaji možnost jeho sirsiho využiti.
The winter beehive debris containing bodies of honeybee parasitic mite Varroa destructor is used ... more The winter beehive debris containing bodies of honeybee parasitic mite Varroa destructor is used for veterinary diagnostics. The Varroa sucking honeybee haemolymph serves as a reservoir of pathogens including bacteria. Worker bees can pick up pathogens from the debris during cleaning activities and spread the infection to healthy bees within the colony. The aim of this study was to detect entomopathogenic bacteria in the Varroa collected from the winter beehive debris.
Zařizeni slouži k filtraci roztaveneho vceliho vosku přes vrstvu polymeru, který navaže některe t... more Zařizeni slouži k filtraci roztaveneho vceliho vosku přes vrstvu polymeru, který navaže některe typy nežadoucich molekul. Zařizeni je kovový valec s děrovaným dnem a vikem, vklada se do vyhřivaneho plastě a umožňuje řizený přetlak. Filtrat je jiman do spodni casti plastě a da se vypoustět kohoutem.
Entomopathogenic nematodes (EPNs) are obligate insect parasites that are symbiotically associated... more Entomopathogenic nematodes (EPNs) are obligate insect parasites that are symbiotically associated with specific entomopathogenic bacteria. These nematobacterial complexes are highly pathogenic to insect hosts and therefore used in biological control of insect pests. As was shown previously, EPNs can be used also as a natural infection model and a powerful tool to study insect immunity. In this study we show that also honeybee larvae are suitable hosts for nematobacterial complex that offer an excellent environment for successful development of both nematodes and their symbiotic bacteria. Here we used EPNs for evaluation of the overall immune resistance of honeybee larvae treated with potentially immuno-modulating substances (plant alcaloid sanguinarin and probiotics). Honeybee larvae were infected with 10 IJ of Heterorhabditis bacteriophora or Steinernema feltiae per larva, both carrying their symbiotic bacteria, and then scored for mortality caused by the nematobacterial infection. In comparison to untreated honeybee larvae we observed the decrease in mortality of approx. 35 % in case of H. bacteriophora and approx. 10% upon S. feltiae infection in larvae treated by selected substances. Both sanguinarin and probiotic treatment showed similar protective effect that was less significant in highly virulent S. feltiae. This is the first record that the nematobacterial infection was used for evaluation of immune status of beneficial insect. We suggest and propose this method as a valuable tool for immunity testing in honeybees as well as in other insects. Our research is supported by grant from the Ministry of Agriculture of Czech Republic (project No. QJ1210047) and by the program CZ.1.07/2.3.00/30.0009 co-financed from European Social Fund and the state budget of the Czech Republic.
Recent colony losses have led to increased interest in the research of honey bee gut microbiota. ... more Recent colony losses have led to increased interest in the research of honey bee gut microbiota. Social bees host in their digestive tract characteristic microbiota and according to recent hypotheses, this plays a significant role in their health. Within our study, two lactobacilli of the Firm-5 cluster and one Gilliamella apicola strain isolates were administered in triplicates to colonies by spraying over the brood frames, either as single strains or in combinations. Data about gut microbial diversity of young bees and larvae (using denaturating gel electrophoresis of 16S DNA V3 region), plasma antioxidant activity (TRAP) and net hive colony weight were recorded weekly for two weeks prior the experiment and six weeks after. Administration of single strains or the mixture resulted in a slightly better performance (not significant). Whereas the net weight of the control dropped by 18.7% due to lack of forage, this drop was lower in hives treated with any bacterial suspension, averaging 8.5% for all treatments. Relatively best performance was seen for the mixture. The administration, however, had no effect on antioxidative capacity of the hemolymph, which tended to decrease after the administration (with the exception of Gilliamella sp., not significant). Expectedly, administration changed the profile of lactic acid bacteria in the gut, which persisted throughout the experiment. Due to limited experiment size, we did not see any significant effect. The slight increase in performance suggests positive role, which is probably not expressed into the tested biochemical parameters (Supported by NAZV QJ1530148 and NAZV QJ1210047)
Bees are used by mankind for several thousand years, but their immune system is still far from be... more Bees are used by mankind for several thousand years, but their immune system is still far from being fully understood and moreover we still don’t have clear idea about all immune mechanisms which mediate bees’ response to their pathogens. These pathogens negatively influence life of the bees and very often even their viability causing direct impact on agriculture and industry. Detailed knowledge of bee immunity is crucial for successful fight against bee diseases; that’s why we are assembling repertoire of immune methods applicable on bees that would enable to evaluate current state of their immunity and help us in proper disease control. General ability of bees to deal with bacterial infection is well characterized by antibacterial activity of their hemolymph. Total antibacterial activity against Gram negative bacteria is detectable by real time measurement of bioluminescence produced by bacteria Escherichia coli K12; the bioluminescence signal is equal to bacterial viability which is decreased by antibacterial factors in hemolymph to at least 55 % in healthy bees. Many effectors acting against Gram positive bacteria were already identified in hemolymph of bees; as marker of their activity we selected the activity of lysozyme, a well characterized enzyme damaging peptidoglycan in cell walls. Commonly used radial zone diffusion assay turned out to be ineffective to determine the level of lysozyme, because of its low sensitivity threshold and the need for large sample volumes. Alternatively, using ELISA kit for lysozyme we determined its level in hemolymph in range from 0.1 to 15 ng/ml. Infection agents activate also stress response leading to changes in antioxidant capacity of hemolymph and activity of particular antioxidant enzymes. TRAP was used to describe total antioxidant capacity in bees and could be complemented by colorimetric measurement of superoxide dismutase and glutathione S-transferase activity.
The infrared (ATR-FTIR) and Raman spectroscopy was used for the structural characterization of ho... more The infrared (ATR-FTIR) and Raman spectroscopy was used for the structural characterization of honeybee wings. Protein, chitin, and lipid characteristic spectral features were detected using both methods. The protein secondary structure was predominantly composed of the β-sheet molecular conformation with β-turns or coil contributions. The vibration modes of the side-chain aromatic amino acid residues (tyrosine, phenylalanine, tryptophan) occurred in the wing spectra. The results of discriminant analysis showed that the infrared spectroscopy of the wing in combination with a multivariate analysis seemed promising for a resolution of the chemical structure of the wings based on lipid, proteins, and chitin content.
Instrumental insemination of Apis mellifera L. queens is a widely employed technique used in hone... more Instrumental insemination of Apis mellifera L. queens is a widely employed technique used in honeybee breeding that enables the effective control of mating. However, drone semen represents a potential source of honeybee viruses. In this study, 43 semen doses collected from apparently healthy drones, and consequently used in instrumental insemination, were analysed using PCR or RT-PCR to detect the presence of viral genome of 11 honeybee viruses. In 91% of samples, viral infection was detected. The survey revealed genomes of five viruses, namely Deformed wing virus (DWV), Acute bee paralysis virus (ABPV), Black queen cell virus (BQCV), Sacbrood virus (SBV), and A. mellifera filamentous virus (AmFV) in 84%, 19%, 14%, 2%, and 67% of samples, respectively. Single infection (30% of samples) as well as multiple infection (61% of samples) of two, three or four pathogens were also evaluated. To the best of our knowledge, this is the first study describing the presence of the BQCV and SBV ge...
Honey is composed mainly of carbohydrates which are represented by mono-, di-, tri-, tetra-, pent... more Honey is composed mainly of carbohydrates which are represented by mono-, di-, tri-, tetra-, pentasaccharides, and oligosaccharides. The content and proportions of individual carbohydrates reveal information about the origin and technological properties of honey. A total of 5 987 samples of natural honeys originating from the Czech Republic and harvested during a period of nine consecutive years were analysed to research their carbohydrate content, sum of fructose and glucose (Fru + Glc), fructose and glucose ratio (Fru/Glc), and electrical conductivity (Ec). Monosaccharides, melezitose (Mel), and Ec varied according to the source of nectar or honeydew. Sucrose (Suc) content was low 0.87 ± 1.26 g (100 g)–1 and did not exceed 15 g (100 g)–1.
In this study we measured in vitro honey bee enzyme activities in presence of the synergist piper... more In this study we measured in vitro honey bee enzyme activities in presence of the synergist piperonyl butoxide (PBO) and some new benzodioxole and dihydrobenzofuran derivatives through absorbance and fluorescence spectrophotometric assays, to determine if and to what extent detoxification systems and acetylcholinesterases are affected by these compounds. Both Italian (Italy) and Carniolan (Czech Republic) honey bees were tested. In the case of Italian honey bees the esterase activity was partially but significantly inhibited by almost all the tested products. In Carniolan honey bees only the dihydrobenzofuran derivative EN16-41 significantly inhibited esterases. Other enzymatic systems, such as cytochrome P450 monooxygenases (P450s) and glutathione S-transferases (GSTs), often involved in xenobiotics detoxification, as well as acetylcholinesterases (AchEs) seem not to be targeted by the studied synergists. These data seem to suggest that some of the investigated PBO analogues could ...
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