Biology subjects

Goblirsch, M.

Publications and source records attributed to Goblirsch, M..

4 recordsLinked to original sources

bioRxiv2025

Clodronate liposomes untangle the role of hemocytes in Apis mellifera response to temperature variation and microbial infection

IntroductionThe cellular immune response, mediated by hemocytes, is a fundamental component of honey bee (Apis mellifera) health. However, the specific contribution of hemocyte subtypes to resilience against combined stressors remains poorly characterized. This study investigated the effect of temperature and bacterial challenge on hemocyte abundance. We employed clodronate liposomes (CLD) for the first time in honey bees as a novel tool for the selective depletion of hemocytes to investigate this gap. MethodsFive-day-old (nurses) and fifteen-day-old (foragers) honey bees were treated with CLD, control liposomes, PBS, or left untreated, then exposed at either 32{degrees}C or 22{degrees}C and challenged with the gram-negative bacterium, Escherichia coli, or the gram-positive bacterium, Staphylococcus aureus. Survival, hemolymph volume, total hemocyte counts, and differential hemocyte counts were monitored over seven days from the start of exposure. ResultsThe CLD application demonstrated significant reductions in granulocyte and prohemocyte populations, indicating the highest vulnerability. A temperature drop to 22{degrees}C buffered the negative impact on survival of CLD-induced immunosuppression. While bacterial challenges universally reduced hemocyte counts, we found an age-dependent difference where nurses maintained significantly higher baseline total hemocyte counts than foragers. Furthermore, temperature did not affect overall total hemocyte counts in 5-day-old nurse bees, but in 15-day-old foragers, it significantly modulated the hemocyte response to bacterial infection. ConclusionOur findings show that hemocyte function is non-uniform, with specific subtypes being essential for overall resilience. The results highlight a previously underappreciated role for temperature as a key modulator of immune capacity, particularly in immunocompromised bees. The age-related differences in hemocyte abundance suggest a life-history trade-off that may prompt the increased vulnerability of honey bees as they age. This work establishes CLD as a powerful tool for insect immunology and sets a precedent for using precise immune manipulation to study host-pathogen-environment interactions.

zoology↗

bioRxiv2025

Acute bee paralysis virus regulation of microRNA expression dynamics in the insect host (Apis mellifera) cell line, AmE-711

BackgroundHoney bees (Apis mellifera) are essential pollinators that support global food production and economic stability. Their health and survival are threatened by diminishing floral resources, pesticide exposure, parasitic mites, and microbial and viral diseases. Among these stressors, viral infections are particularly challenging, often exacerbated by the parasitic mite, Varroa destructor, a competent vector of multiple honey bee viruses. Understanding the mechanisms underlying honey bee-virus interactions is critical for mitigating the negative impact of infections on colony health. One understudied aspect is the role of microRNAs (miRNAs) in viral pathogenesis and antiviral defense. miRNAs are short, non-coding RNAs produced by both hosts and pathogens that act as post-transcriptional regulators of gene expression and can influence host-pathogen dynamics during infection. In this study, we used a honey bee-derived cell line to test the hypothesis that viral infection perturbs honey bee- and viral-encoded miRNA expression. MethodsSmall RNA libraries from honey bee AmE-711 cells subjected to one of four treatments: media change only (uninfected), heat-killed Acute bee paralysis virus (ABPV), the viral mimic Poly(I:C), or infectious ABPV, were prepared using an Illumina Truseq kit. Sequencing data were analyzed using miRDeep2 and sRNAtoolbox to identify differentially expressed (DE) miRNAs, which were subsequently validated by RT-qPCR assay. ResultsSequencing yielded > 3.6 x 108 raw reads that were assigned to 12 small RNA libraries, from which, 481 unique miRNAs were identified. Moreover, 15 miRNAs were DE in ABPV-infected cells compared to uninfected cells: miR-2b-5p, miR-33-5p, miR-133-3p, miR-6001-3p, miR-996-3p, miR-965-3p, miR-125-5p, miR-13b-3p, miR-79-3p, miR-971-3p, miR-277-3p, miR-92c-5p, miR-6065-3p, miR-965-5p, and miR-3786-5p. We highlight some of the DE miRNAs identified in ABPV-infected cells that show regulatory effects in other systems in response to infection. ConclusionThis study identified miRNAs differentially expressed in ABPV-infected cells, suggesting roles in either antiviral defense or in promoting viral pathogenesis through suppression of host immune responses. These results provide a foundation for functional studies using honey bee cell lines to clarify the cellular mechanisms governing honey bee-virus interactions.

zoology↗

bioRxiv2024

Successful delivery of CRISPR-Cas9 with a baculovirus vector for insect brain targets

CRISPR-Cas9 (clustered, regularly interspaced, short palindromic repeats with CRISPR-associated protein 9) is a powerful, versatile, and cost-effective molecular tool that can be used for genetic engineering purposes and beyond1 and is especially suited for non-model organisms2. Effective delivery of this system, however, remains a challenge for in vivo genetic manipulation of specific tissues3, particularly the brain4, and in adult indivuduals5,6. We designed a new CRISPR-Cas9 plasmid that was inserted into a baculovirus vector to knockdown the octopamine beta subtype 2 receptor (AmOct{beta}2R), a transmembrane protein found in the mushroom body neurons of the honey bee (Apis mellifera) brain, to determine if octopamine plays a role in appetite regulation. We first confirmed that gene editing of AmOct{beta}2R is possible with Sanger sequencing. We then demonstrated expression of the CRISPR-Cas9 system with the baculovirus vector in vitro using live cell imaging, flow cytometry analysis, and in vivo using confocal imaging, showing widespread expression in the cells and throughout the honey bee brain, three days post treatment. There was also in vitro and in vivo knockdown of AmOct{beta}2R three days post-infection, that corresponded with appetite suppression in starved forager bees. Our findings suggest that we successfully delivered the CRISPR-Cas9 system and knocked down AmOctB2R in neuronal cells of the honey bee brain that were previously inaccessible due to the blood brain barrier and lack of infectivity of lentivirus vectors7. The newly characterized AmOct{beta}2R8 can now be assigned a functional role and other targets for gene editing are now possible using this CRISPR-Cas9 system. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=140 SRC="FIGDIR/small/624635v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@9fba0eorg.highwire.dtl.DTLVardef@7025dforg.highwire.dtl.DTLVardef@c6706borg.highwire.dtl.DTLVardef@1fea727_HPS_FORMAT_FIGEXP M_FIG C_FIG

genetics↗

bioRxiv2023

Honey bee Apis mellifera L. Responses to Oxidative Stress Induced by Pharmacological and Pesticide Compounds

The western honey bee, Apis mellifera L., is a eusocial insect that plays major roles in ecosystem balances and pollination of plants and food crops. Honey bees face multiple biotic and abiotic stressors, such as pathogens, diseases, chemical pesticides, and climate change, which all contribute to honey bee colony loss. This study investigated the impacts of multiple pharmacological and pesticide molecules on honey bee survival and gene regulation responses. In an 11-day cage experiment, sublethal doses of tunicamycin, thapsigargin, metformin, paraquat, hydrogen peroxide, and imidacloprid were administered to newly emerged sister bees. Daily treatment consumption and mortality were recorded, as well as the transcription expression of twelve major genes (AChE-2, Apisimin, Apidaecin, mrjp1, Sodq, cp450, SelT, SelK, Ire1, Xbp1, Derl-1, Hsc70), some of which are markers of oxidative and endoplasmic reticulum (ER) stresses in honey bees. At day 9 of the treatments, protein damage was quantified in caged bees. Kaplan-Meier model indicated significant (p < 0.001) toxicological effects of paraquat, H2O2 and tunicamycin on bee survivorship compared to controls with better survivals for other molecules. Post-ingestive aversion responses were recorded only in the case of tunicamycin, hydrogen peroxide and imidacloprid. Nonetheless, significantly higher protein damage on day 9 was only identified in bees exposed to paraquat and imidacloprid. Some antioxidant genes significantly regulated vis-a-vis specific treatments. Our results reveal age-related regulation of other major genes with significant inter-gene positive correlations.

zoology↗