Varroa destructor mite is a major threat to honeybee (Apis mellifera) populations, contributing to colony losses through parasitism and pathogen transmission. While extensive research has focused on Varroa biology and its role as a virus vector, its microbiome remains poorly understood, particularly regarding geographic variation. Here, we investigated the microbial diversity, composition, and functional potential of Varroa mite microbiota collected from two neighboring countries, Czechia and Slovakia. Using high-throughput sequencing and network analysis, we assessed alpha and beta diversity metrics, microbial co-occurrence patterns, and predicted metabolic functions. Our results revealed significant differences in microbial diversity between the two regions, with some bacterial taxa appearing more prevalent in specific populations. Network analysis suggested potential variation in the structural stability of microbial communities in Varroa mites, raising the possibility that geographic factors may influence microbial interactions. Functional profiling indicated region-associated differences in predicted metabolic pathways, possibly linked to certain bacterial taxa. While these findings provide new insights into the Varroa microbiome and its potential ecological role, the interpretation of geographic influence remains a subject of ongoing investigation to better understand its scope and underlying mechanisms. A deeper understanding of these microbial dynamics may contribute to the development of novel strategies for Varroa mite management and the conservation of honeybee health.

• Klíčová slova
• Varroa destructor, Geographical variation, Microbial co-occurrence networks, Microbiome, Predicted functional pathways,
• Publikační typ
• časopisecké články MeSH

The Western honey bee (Apis mellifera) is a vital agricultural pollinator whose populations are threatened by the parasitic mite Varroa destructor and associated pathogens. While the impact of Paenibacillus species on honey bees, particularly Paenibacillus larvae causing American foulbrood, is documented, their effect on the microbiota of Varroa mites remains unclear. This study aimed to investigate the influence of Paenibacillus sp. on the bacterial communities of Varroa mites and adult honey bees. We hypothesized that Paenibacillus sp. would significantly alter the microbiota of Varroa mites but have minimal effect on that of adult honey bees. Utilizing 16S rRNA sequencing data from a previous study, we reanalyzed samples categorized into four groups based on Paenibacillus sp. infection load: highly infected and lowly infected honey bees (A. mellifera) and mites (V. destructor). Infection status was determined by Paenibacillus sp. read counts, with more than three reads indicating high infection. Microbial diversity was assessed using alpha and beta diversity metrics. Co-occurrence networks were constructed to visualize bacterial community assemblies, and network robustness was evaluated through node addition and removal tests. Keystone taxa were identified based on eigenvector centrality and relative abundance. Highly infected Varroa mites exhibited a significant reduction in alpha diversity and a markedly different bacterial community composition compared to lowly infected mites (p < 0.05). Their bacterial co-occurrence networks showed decreased connectivity and robustness, indicating a disruptive effect of Paenibacillus sp. In contrast, adult honey bees displayed no significant differences in alpha diversity or network structure between highly and lowly infected groups (p > 0.05), suggesting a resilient microbiota. Keystone taxa analysis revealed fewer central species in highly infected Varroa mites, potentially impacting network stability. High Paenibacillus sp. infection is associated with significant alterations in the microbiota of Varroa mites, disrupting bacterial communities and potentially affecting mite physiology. The microbiota of adult honey bees appears more robust against Paenibacillus sp. influence. These findings enhance our understanding of the complex interactions within the "honey bee-mite-microorganism" system and may inform future strategies for managing Varroa mite infestations and associated pathogens.

• Klíčová slova
• Apis mellifera, Microbial networks, Microbiota analysis, Paenibacillus sp., Varroa destructor,
• Publikační typ
• časopisecké články MeSH

A novel Bartonella-like symbiont (BLS) of Tyrophagus putrescentiae was characterized. BLS formed a separate cluster from the Bartonella clade together with an ant symbiont. BLS was present in mite bodies (103 16S DNA copies/mite) and feces but was absent in eggs. This indicated the presence of the BLS in mite guts. The BLS showed a reduction in genome size (1.6 Mb) and indicates gene loss compared to Bartonella apis. The BLS can be interacted with its host by using host metabolic pathways (e.g., the histidine and arginine metabolic pathways) as well as by providing its own metabolic pathways (pantothenate and lipoic acid) to the host, suggesting the existence of a mutualistic association. Our experimental data further confirmed these potential mutualistic nutritional associations, as cultures of T. putrescentiae with low BLS abundance showed the strongest response after the addition of vitamins. Despite developing an arguably tight dependency on its host, the BLS has probably retained flagellar mobility, as evidenced by the 32 proteins enriched in KEGG pathways associated with flagellar assembly or chemotaxis (e.g., fliC, flgE, and flgK, as highly expressed genes). Some of these proteins probably also facilitate adhesion to host gut cells. The microcin C transporter was identified in the BLS, suggesting that microcin C may be used in competition with other gut bacteria. The 16S DNA sequence comparison indicated a mite clade of BLSs with a broad host range, including house dust and stored-product mites. Our phylogenomic analyses identified a unique lineage of arachnid specific BLSs in mites and scorpions.IMPORTANCEA Bartonella-like symbiont was found in an astigmatid mite of allergenic importance. We assembled the genome of the bacterium from metagenomes of different stored-product mite (T. putrescentiae) cultures. The bacterium provides pantothenate and lipoic acid to the mite host. The vitamin supply explains the changes in the relative abundance of BLSs in T. putrescentiae as the microbiome response to nutritional or pesticide stress, as observed previously. The phylogenomic analyses of available 16S DNA sequences originating from mite, scorpion, and insect samples identified a unique lineage of arachnid specific forming large Bartonella clade. BLSs associated with mites and a scorpion. The Bartonella clade included the previously described Ca. Tokpelaia symbionts of ants.

• Klíčová slova
• Bartonella, ants, house dust, mite, nutrition, stored-product, symbionts, vitamin,
• MeSH
• Acaridae * mikrobiologie   MeSH
• alergeny MeSH
• Bacteria MeSH
• Bartonella * genetika   MeSH
• kyselina lipoová * MeSH
• roztoči * genetika   MeSH
• symbióza MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• alergeny MeSH
• kyselina lipoová * MeSH

Interactions with microorganisms might enable house dust mites (HDMs) to derive nutrients from difficult-to-digest structural proteins and to flourish in human houses. We tested this hypothesis by investigating the effects of changes in the mite culture growth and population of two HDM species on HDM microbiome composition and fitness. Growing cultures of laboratory and industrial allergen-producing populations of Dermatophagoides farinae (DFL and DFT, respectively) and Dermatophagoides pteronyssinus (DPL and DPT, respectively) were sampled at four time points. The symbiotic microorganisms of the mites were characterized by DNA barcode sequencing and quantified by qPCR using universal/specific primers. The population growth of mites and nutrient contents of mite bodies were measured and correlated with the changes in bacteria in the HDM microbiome. The results showed that both the population and culture age significantly influenced the microbiome profiles. Cardinium formed 93% and 32% of the total sequences of the DFL and DFT bacterial microbiomes, respectively, but this bacterial species was less abundant in the DPL and DPT microbiomes. Staphylococcus abundance was positively correlated with increased glycogen contents in the bodies of mites, and increased abundances of Aspergillus, Candida, and Kocuria were correlated with increased lipid contents in the bodies of mites. The xerophilic fungus Wallemia accounted for 39% of the fungal sequences in the DPL microbiome, but its abundance was low in the DPT, DFL, and DFT microbiomes. With respect to the mite culture age, we made three important observations: the mite population growth from young cultures was 5-8-fold higher than that from old cultures; specimens from old cultures had greater abundances of fungi and bacteria in their bodies; and yeasts predominated in the gut contents of specimens from young cultures, whereas filamentous mycelium prevailed in specimens from old cultures. Our results are consistent with the hypothesis that mites derive nutrients through associations with microorganisms.

• Klíčová slova
• Bacteria, Dermatophagoides farinae, Dermatophagoides pteronyssinus, Diet, Fungi, Gut, Nutrition, Symbiosis, Yeasts,
• MeSH
• Bacteria * klasifikace   MeSH
• bakteriální RNA analýza   MeSH
• druhová specificita MeSH
• fungální RNA analýza   MeSH
• houby * klasifikace   MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• mikrobiota * MeSH
• populační dynamika MeSH
• Pyroglyphidae mikrobiologie   fyziologie   MeSH
• RNA ribozomální 16S analýza   MeSH
• RNA ribozomální 18S analýza   MeSH
• taxonomické DNA čárové kódování MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bakteriální RNA MeSH
• fungální RNA MeSH
• RNA ribozomální 16S MeSH
• RNA ribozomální 18S MeSH

Varroa destructor is the major cause of honey bee (Apis mellifera) colony losses. Mite control is limited to several miticides. The overuse of tau-fluvalinate has resulted in resistance via a knockdown resistance (kdr) mutation in the sodium channel gene NaVChs (L925V/I/M). In this study, we used the discriminating concentration of tau-fluvalinate (0.25 µg/mL) to detect the resistance of mites in a bioassay. Further, we verified the presence of the kdr mutation in mites from the bioassay via PCR amplification of a fragment of the voltage-gated sodium channel gene (NaVCh), restriction fragment length polymorphisms (RFLPs), and densitometry analyses in pools of surviving or dead mites. Resistance values corresponding to the densitometry of the resistant allele were related to mite survival. In the vial test, the survival of the control group was significantly higher (70.4%) than that of the tau-fluvalinate-treated group (34.3%). Mite survival in the vial test was significantly correlated with the mean proportion of resistance values. Individuals that died after tau-fluvalinate application exhibited an average resistance value of 0.0783, whereas individuals that survived exhibited an average resistance of 0.400. The concentration of tau-fluvalinate in the vials was checked using high performance liquid chromatography under different temperatures and exposure times, and indicates that the stability of tau-fluvalinate stored in the refrigerator (4 ± 1 °C) is at least 14 days. PCR-RFLP of the NaVCh gene fragment verified that the vial test is a suitable, rapid, and cost-effective method for the identification of tau-fluvalinate resistance based on kdr mutation in V. destructor in apiaries.

• Klíčová slova
• Acaricide, Apiary, Honey bee, Restriction, Varroa destructor,
• MeSH
• akaricidy farmakologie   MeSH
• biotest metody   MeSH
• denzitometrie metody   MeSH
• léková rezistence genetika   MeSH
• nitrily farmakologie   MeSH
• polymerázová řetězová reakce metody   MeSH
• polymorfismus délky restrikčních fragmentů MeSH
• pyrethriny farmakologie   MeSH
• Varroidae účinky léků   genetika   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH
• Názvy látek
• akaricidy MeSH
• fluvalinate MeSH Prohlížeč
• nitrily MeSH
• pyrethriny MeSH

Blood feeding red poultry mites (RPM) serve as vectors of pathogenic bacteria and viruses among vertebrate hosts including wild birds, poultry hens, mammals, and humans. The microbiome of RPM has not yet been studied by high-throughput sequencing. RPM eggs, larvae, and engorged adult/nymph samples obtained in four poultry houses in Czechia were used for microbiome analyses by Illumina amplicon sequencing of the 16S ribosomal RNA (rRNA) gene V4 region. A laboratory RPM population was used as positive control for transcriptome analysis by pyrosequencing with identification of sequences originating from bacteria. The samples of engorged adult/nymph stages had 100-fold more copies of 16S rRNA gene copies than the samples of eggs and larvae. The microbiome composition showed differences among the four poultry houses and among observed developmental stadia. In the adults' microbiome 10 OTUs comprised 90 to 99% of all sequences. Bartonella-like bacteria covered between 30 and 70% of sequences in RPM microbiome and 25% bacterial sequences in transcriptome. The phylogenetic analyses of 16S rRNA gene sequences revealed two distinct groups of Bartonella-like bacteria forming sister groups: (i) symbionts of ants; (ii) Bartonella genus. Cardinium, Wolbachia, and Rickettsiella sp. were found in the microbiomes of all tested stadia, while Spiroplasma eriocheiris and Wolbachia were identified in the laboratory RPM transcriptome. The microbiomes from eggs, larvae, and engorged adults/nymphs differed. Bartonella-like symbionts were found in all stadia and sampling sites. Bartonella-like bacteria was the most diversified group within the RPM microbiome. The presence of identified putative pathogenic bacteria is relevant with respect to human and animal health issues while the identification of symbiontic bacteria can lead to new control methods targeting them to destabilize the arthropod host.

• Klíčová slova
• Bartonella, Blood sucking, Mite, Poultry, Ricketsiella, Transmission, Tsukamurella, Wolbachia,
• MeSH
• Bacteria klasifikace   genetika   izolace a purifikace   MeSH
• bakteriální RNA genetika   MeSH
• Bartonella klasifikace   genetika   izolace a purifikace   MeSH
• druhová specificita MeSH
• mikrobiota * MeSH
• RNA ribozomální 16S genetika   MeSH
• roztoči růst a vývoj   mikrobiologie   MeSH
• taxonomické DNA čárové kódování MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• bakteriální RNA MeSH
• RNA ribozomální 16S MeSH

The honey bee, Apis mellifera, is a globally important species that suffers from a variety of pathogens and parasites. These parasites and pathogens may have sublethal effects on their bee hosts via an array of mechanisms, including through a change in symbiotic bacterial taxa. Our aim was to assess the influence of four globally widespread parasites and pathogens on the honey bee bacteriome. We examined the effects of the ectoparasitic mite Varroa destructor, the fungal pathogens Nosema apis and Nosema ceranae, and the trypanosome Lotmaria passim. Varroa was detected by acaricidal treatment, Nosema and L. passim by PCR, and the bacteriome using MiSeq 16S rRNA gene sequencing. Overall, the 1,858,850 obtained sequences formed 86 operational taxonomic units (OTUs) at 3 % dissimilarity. Location, time of year, and degree of infestation by Varroa had significant effects on the composition of the bacteriome of honey bee workers. Based on statistical correlations, we found varroosis more important factor than N. ceranae, N. apis, and L. passim infestation influencing the honey bee bacteriome and contributing to the changes in the composition of the bacterial community in adult bees. At the population level, Varroa appeared to modify 20 OTUs. In the colonies with high Varroa infestation levels (varroosis), the relative abundance of the bacteria Bartonella apis and Lactobacillus apis decreased. In contrast, an increase in relative abundance was observed for several taxa including Lactobacillus helsingborgensis, Lactobacillus mellis, Commensalibacter intestini, and Snodgrassella alvi. The results showed that the "normal" bacterial community is altered by eukaryotic parasites as well as displaying temporal changes and changes associated with the geographical origin of the beehive.

• Klíčová slova
• Bacteria, Lotmaria passim, Nosema apis, Nosema ceranae, Sequencing, Varroa destructor,
• MeSH
• Bartonella klasifikace   genetika   izolace a purifikace   MeSH
• infestace roztoči patologie   MeSH
• Kinetoplastida patogenita   MeSH
• Lactobacillus klasifikace   genetika   izolace a purifikace   MeSH
• mikrobiota genetika   MeSH
• Nosema patogenita   MeSH
• RNA ribozomální 16S genetika   MeSH
• symbióza MeSH
• Varroidae patogenita   MeSH
• včely mikrobiologie   parazitologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• RNA ribozomální 16S MeSH

The ectoparasitic mite Varroa destructor is a major pest of the honeybee Apis mellifera. In a previous study, bacteria were found in the guts of mites collected from winter beehive debris and were identified using Sanger sequencing of their 16S rRNA genes. In this study, community comparison and diversity analyses were performed to examine the microbiota of honeybees and mites at the population level. The microbiota of the mites and honeybees in 26 colonies in seven apiaries in Czechia was studied. Between 10 and 50 Varroa females were collected from the bottom board, and 10 worker bees were removed from the peripheral comb of the same beehive. Both bees and mites were surface sterilized. Analysis of the 16S rRNA gene libraries revealed significant differences in the Varroa and honeybee microbiota. The Varroa microbiota was less diverse than was the honeybee microbiota, and the relative abundances of bacterial taxa in the mite and bee microbiota differed. The Varroa mites, but not the honeybees, were found to be inhabited by Diplorickettsia. The relative abundance of Arsenophonus, Morganella, Spiroplasma, Enterococcus, and Pseudomonas was higher in Varroa than in honeybees, and the Diplorickettsia symbiont detected in this study is specific to Varroa mites. The results demonstrated that there are shared bacteria between Varroa and honeybee populations but that these bacteria occur in different relative proportions in the honeybee and mite bacteriomes. These results support the suggestion of bacterial transfer via mites, although only some of the transferred bacteria may be harmful.

• Klíčová slova
• Apis mellifera, Arsenophonus, Diplorickettsia, Spiroplasma, Symbiosis, Varroa destructor,
• MeSH
• biodiverzita MeSH
• DNA bakterií genetika   MeSH
• mikrobiota * MeSH
• RNA ribozomální 16S genetika   MeSH
• roční období MeSH
• sekvenční analýza DNA MeSH
• Spiroplasma klasifikace   izolace a purifikace   MeSH
• symbióza MeSH
• Varroidae mikrobiologie   MeSH
• včely mikrobiologie   parazitologie   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• DNA bakterií MeSH
• RNA ribozomální 16S MeSH

The parasitic mite Varroa destructor is a major pest of the western honeybee, Apis mellifera. The development of acaricide resistance in Varroa populations is a global issue. Discriminating concentrations of acaricides are widely used to detect pest resistance. Two methods, using either glass vials or paraffin capsules, are used to screen for Varroa resistance to various acaricides. We found the glass vial method to be useless for testing Varroa resistance to acaridices, so we developed a polypropylene vial bioassay. This method was tested on tau-fluvalinate-, acrinathrin-, and amitraz-resistant mite populations from three apiaries in Czechia. Acetone was used as a control and technical grade acaricide compounds diluted in acetone were applied to the polypropylene vials. The solutions were spread on the vial surface by rolling the vial, and were then evaporated. Freshly collected Varroa females were placed in the vials and the mortality of the exposed mites was measured after 24 h. The Varroa populations differed in mortality between the apiaries and the tested compounds. Mites from the Kyvalka site were resistant to acrinathrin, tau-fluvalinate, and amitraz, while mites from the Postrizin site were susceptible to all three acaricides. In Prelovice apiary, the mites were susceptible to acrinathrin and amitraz, but not to tau-fluvalinate. The calculated discriminating concentrations for tau-fluvalinate, acrinathrin, and amitraz were 0.66, 0.26 and 0.19 µg/mL, respectively. These results indicate that polyproplyne vial tests can be used to determine discriminating concentrations for the early detection of acaricide resistant Varroa. Finally, multiple-resistance in Kyvalka may indicate metabolic resistance.

• Klíčová slova
• Acaricide, Apiculture, Discriminating concentrations, Multiple-resistance, Varroa,
• MeSH
• akaricidy * MeSH
• fixní kombinace léků MeSH
• kontrola klíšťat * MeSH
• nitrily * MeSH
• polylysin analogy a deriváty   MeSH
• pyrethriny * MeSH
• toluidiny * MeSH
• Varroidae * MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• acrinathrin MeSH Prohlížeč
• akaricidy * MeSH
• amitraz MeSH Prohlížeč
• fixní kombinace léků MeSH
• fluvalinate MeSH Prohlížeč
• gemals MeSH Prohlížeč
• nitrily * MeSH
• polylysin MeSH
• pyrethriny * MeSH
• toluidiny * MeSH