DOI: 10.5061/dryad.5tb2rbpdc
title: Data from: Wolbachia enhances the survival of Drosophila infected with fungal pathogens

description: Wolbachia bacteria of arthropods are at the forefront of basic and
 translational research on multipartite host-symbiont-pathogen
 interactions. These vertically transmitted microbes are the most
 widespread endosymbionts on the planet due to factors including host
 reproductive manipulation and fitness benefits. Importantly, some strains
 of Wolbachia can inhibit viral pathogenesis within and between arthropod
 hosts. Mosquitoes carrying the wMel Wolbachia strain of Drosophila
 melanogaster have a greatly reduced capacity to spread viruses like dengue
 and Zika to humans. While significant research efforts have focused on
 viruses, relatively little attention has been given to Wolbachia-fungal
 interactions despite the ubiquity of fungal entomopathogens in nature.
 Here, we demonstrate that Wolbachia increase the longevity of their
 Drosophila melanogaster hosts when challenged with a spectrum of yeast and
 filamentous fungal pathogens. We find that this pattern can vary based on
 host genotype, sex, and fungal species. Further, Wolbachia correlates with
 higher fertility and reduced pathogen titers during initial fungal
 infection, indicating a significant fitness benefit. Finally, RNAseq
 results show altered expression of many immune and stress response genes
 in the context of Wolbachia and fungal infection, suggesting host immunity
 may be involved in the mechanism. This study demonstrates Wolbachia’s role
 in diverse fungal pathogen interactions and determines that the phenotype
 is broad, but with several variables that influence both the presence and
 strength of the phenotype. It also is a critical step forward to
 understanding how symbionts can protect their hosts from a variety of
 pathogens.
description: All data in this dataset was collected in 2022-2023 in a
 laboratory setting from manual recordings of surviving flies and eggs laid
 or hatched, or qPCR Ct values. Minimal processing was applied to all data,
 including calculating proportions, delta Ct values, and visualization and
 statistical analysis in R.
description: # Data from: Wolbachia enhances the survival of *Drosophila* infected with
 fungal pathogens
 [https://doi.org/10.1186/s12915-025-02130-0](https://doi.org/10.1186/s12915-025-02130-0) ## Description of the data and file structure This version of the README file was generated on 03-07-25 by Jessamyn Perlmutter. ### General Information 1. Title of Dataset: *Wolbachia* enhances the survival of *Drosophila* infected with fungal pathogens 2. Author Information: 1. Principal investigator contact information Name: Robert Unckless Institution: University of Kansas Address: Lawrence, KS, USA Email: [unckless@ku.edu](mailto:unckless@ku.edu) 2\. First author contact information Name: Jessamyn Perlmutter Institution: University of Kansas Address: Lawrence, KS, USA Email: [jessie.perlmutter@gmail.com](mailto:jessie.perlmutter@gmail.com) 3. Dates of data collection: 2022-2023 4. Location of data collection: University of Kansas, Lawrence 5. Funding sources that supported the collection of the data: NIH R01 AI139154, P30 GM110761, K-INBRE P20 GM103418, K99 AI180425, NSF DBI 2109772, and NSF DEB 2330095. 6. This dataset contains the supplemental information associated with the article on BMC Biology ([https://bmcbiol.biomedcentral.com/articles/10.1186/s12915-025-02130-0](https://bmcbiol.biomedcentral.com/articles/10.1186/s12915-025-02130-0)). The contents include raw data underlying graphs and R code used to analyze results. All R code is accompanied by corresponding .csv files with the data. ### Sharing/Access information 1. Licenses/restrictions placed on this data: CC0 1.0 Universal (CC0 1.0) Public Domain 2. Links to publications that cite or use the data: Perlmutter, J. I., Atadurdyyeva, A., Schedl, M. E., & Unckless, R. L. (2025). *Wolbachia* enhances the survival of *Drosophila* infected with fungal pathogens. *BMC Biology*, *23 *(1), 42. [https://bmcbiol.biomedcentral.com/articles/10.1186/s12915-025-02130-0](https://bmcbiol.biomedcentral.com/articles/10.1186/s12915-025-02130-0). 3\. Links to other publicly accessible locations of the data: None 4\. Was data derived from another source? No A. If yes, list source(s): N/A 5\. Recommended citation for this dataset: Perlmutter, J. I., Atadurdyyeva, A., Schedl, M. E., & Unckless, R. L. (2025). Data from: *Wolbachia* enhances the survival of *Drosophila* infected with fungal pathogens. Dryad Digital Repository. ### Data & File Overview 1\. File List: A) Fig1-3&1S-3SInfectionSurvivalCurves.R B) Fig1&2AInfectionswk.csv C) Fig1S&S2AInfectionsw1118.csv D) Fig2InfectionsMetarhiziumwk.csv E) Fig2InfectionsTrichodermaClonostachyswk.csv F) Fig2SInfectionsMetarhiziumw1118.csv G) Fig2SInfectionsTrichodermaClonostachysw1118.csv H) Fig3InfectionsYeastwk.csv I) Fig3SInfectionsYeastw1118.csv J) Fig4.S4.Fertility.R K) Fig4Fertilitywk.csv L) Fig4SFertilityw1118.csv M) Fig5.5S.Titers.R N) Fig5BbasTiterswk.csv O) Fig5SBbasTitersw1118.csv P) Fig5SWolbTitersw1118.csv Q) Fig5WolbTiterswk.csv R) all.longest.counts.txt S) id_validation_table_93194.txt T) RNAseq.R 2\. Relationship between files, if important: A) Data underlying all figures is included in .csv files (Files B-I, K-L, & N-Q, above). B) R code used to analyze all of the data .csv files is included (Files A, J, & M above). C) List of corresponding data and analysis files: 1. Figs 1-3, S1-S3 (Infection survival curves): Files A-I 2. Fig 4, S4 (Fertility): Files J-L 3. Fig 5, S5 (Titers): Files M-Q 4. Fig 6, S6, 7 (RNAseq): Files R-T 3\. Additional related data collected that was not included in the current data package: None 4\. Are there multiple versions of the dataset? No ### Files and variables ### Data-Specific Information Software and packages used: Data analysis and figure generation were performed in R v4.2.2, using several packages: coxme (v2.2.18.1), ggplot2 (v3.4.0), cowplot (v1.1.1), car (v3.1.1), SurvMiner (v0.4.9), SurvMisc (v0.5.6), DESeq2 (v1.44.0), tidyverse (v2.0.0), rtracklayer (v1.64.0), RColorBrewer (v1.1-3), edgeR (v4.2.1), EnhancedVolcano (v1.22.0), ape (v5.8), apeglm (v1.26.1), and reshape2 (v1.4.4). Exact packages used for each figure may be found in the corresponding R file. Refer to the [BMC Biology article](https://bmcbiol.biomedcentral.com/articles/10.1186/s12915-025-02130-0) for citations of these packages. ##### Information for Files A, J, & M: Data analysis code in .R files 1. Description: All .R files used to analyze data from .csv files, including creation of figures and statistical analysis. 2. Variables, data codes, formats, & abbreviations: 1. For data corresponding to each .R file, see "Data and File Overview" section, part 2. 2. Variables are either created and defined in the code or details can be read below in the Information sections below for corresponding .csv data files. ##### Information for File B: Fig1&2AInfectionswk.csv 1. Description: Infection survival data underlying Figures 1 and 2A for the wk host genetic background with B. bassiana, A. fumigatus, F. oxysporum, F. graminaerum, and A. flavus infection. Note that while some samples were counted past day 21, only survival through day 21 was analyzed for this manuscript. 2. Variables, data codes, formats, & abbreviations: 1. Vial: Number with each representing a unique biological sample. 2. Gt: Genotype of the host (wk or w1118) 3. Wolbachia: Indicator of presence (I, infected) or absence (U, uninfected) of Wolbachia symbiont 4. Line: Particular host isoline (all the same in this file, left blank) 5. Treatment: Infection or control treatment used for a given sample 1. 20% Glycerol= 20% Glycerol negative infection control for filamentous fungi 2. B. bassiana= Beauveria bassiana filamentous fungus 3. A. fumigatus= Apergillus fumigatus filamentous fungus 4. F. graminaerum= Fusarium graminaerum filamentous fungus 5. F. oxysporum= Fusarium oxysporum filamentous fungus 6. A. flavus= Aspergillus flavus filamentous fungus 6. Replicate: Replicate number within each sample set of a given Gt x Treatment x Day x Wolbachia combination 7. Sex: Sex of flies from a given sample (F for female, M for male) 8. DayCollected: Date flies were collected and set aside for experiment 9. DayInfected: Date flies were initially infected (Day 0) 10. Day# Columns: Surviving flies on Day# post-infection. Day0 indicates day 0, the day of infection. All vials start with 10 flies at Day0. ##### Information for File C: Fig1S\&S2AInfectionsw1118.csv 1. Description: Infection survival data underlying Figures S1 and S2A for the w1118 host genetic background with B. bassiana, A. fumigatus, F. oxysporum, F. graminaerum, and A. flavus infection. Note that while some samples were counted past day 21, only survival through day 21 was analyzed for this manuscript. 2. Variables, data codes, formats, & abbreviations: 1. All variables and abbreviations are the same as listed for File B, except for the genotype (Gt), which is w1118 here. ##### Information for File D: Fig2InfectionsMetarhiziumwk.csv 1. Description: Infection survival data underlying Figure 2B for the wk host genetic background with M. anisopliae infection. 2. Variables, data codes, formats, & abbreviations: 1. All variables and abbreviations are the same as listed for File B, with the exception of the "Treatment" variable: 1. Treatment: Infection or control treatment used for a given sample 1. 20% Glycerol= 20% Glycerol negative infection control for filamentous fungi 2. M. anisopliae= Metarhizium anisopliae filamentous fungus ##### Information for File E: Fig2InfectionsTrichodermaClonostachyswk.csv 1. Description: Infection survival data underlying Figures 2C & 2D for the wk host genetic background with T. atroviride or C. rosea infection. 2. Variables, data codes, formats, & abbreviations: 1. All variables and abbreviations are the same as listed for File B, with the exception of the "Treatment" variable: 2. Treatment: Infection or control treatment used for a given sample 1. 20% Glycerol= 20% Glycerol negative infection control for filamentous fungi 2. C. rosea 1:100= Clonostacys rosea filamentous fungus diluted 1:100 from its concentrated stock 3. T. atroviride 1:20: Trichoderma atroviride filamentous fungus diluted 1:20 from its concentrated stock ##### Information for File F: Fig2SInfectionsMetarhiziumw1118.csv 1. Description: Infection survival data underlying Figure S2B for the w1118 host genetic background with M. anisopliae infection. 2. Variables, data codes, formats, & abbreviations: 1. All variables and abbreviations are the same as listed for File D, except for the genotype (Gt), which is w1118 here. ##### Information for File G: Fig2SInfectionsTrichodermaClonostachysw1118.csv 1. Description: Infection survival data underlying Figures 2C & 2D for the w1118 host genetic background with T. atroviride or C. rosea infection. 2. Variables, data codes, formats, & abbreviations: 1. All variables and abbreviations are the same as listed for File E, except for the genotype (Gt), which is w1118 here. ##### Information for File H: Fig3InfectionsYeastwk.csv 1. Description: Infection survival data underlying Figure 3 for the wk host genetic background with C. glabrata, C. auris, or G. psuedocandidus infection. 2. Variables, data codes, formats, & abbreviations: 1. Vial: Number with each representing a unique biological sample. 2. Gt: Genotype of the host (wk or w1118) 3. Wolbachia: Indicator of presence (I, infected) or absence (U, uninfected) of Wolbachia symbiont 4. Line: Placeholder column which also records the optical density (OD) of each sample that was used 5. Treatment: Infection or control treatment used for a given sample 1. PDB= Potato dextrose broth, negative control for yeast 2. C. auris= Candida auris yeast 3. C. glabrata= Candida glabrata yeast 4. G. pseudocandidus= Galactomyces pseudocandidus yeast 6. Replicate: Replicate number within each sample set of a given Gt x Treatment x Day x Wolbachia combination 7. Sex: Sex of flies from a given sample (F for female, M for male) 8. DayCollected: Date flies were collected and set aside for experiment 9. DayInfected: Date flies were initially infected (Day 0) 10. Day# Columns: Surviving flies on Day# post-infection. Day0 indicates day 0, the day of infection. All vials start with 10 flies at Day0. ##### Information for File I: Fig3SInfectionsYeastw1118.csv 1. Description: Infection survival data underlying Figure 3 for the w1118 host genetic background with C. glabrata, C. auris, or G. psuedocandidus infection. 2. Variables, data codes, formats, & abbreviations: 1. All variables and abbreviations are the same as listed for File H, except for the genotype (Gt), which is w1118 here. ##### Information for File K: Fig4Fertilitywk.csv 1. Description: Fertility data underlying Figure 4 for the wk host genetic background, including egg lay and egg hatch rates over different time periods. 2. Variables, data codes, formats, & abbreviations: 1. Vial: Number with each representing a unique biological sample. 2. Background: Genotype of the host (wk or w1118) 3. Wolbachia: Indicator of presence (I, infected) or absence (U, uninfected) of Wolbachia symbiont 4. Treatment: Infection or control treatment used for a given sample 1. Glycerol: 20% Glycerol negative infection control for filamentous fungi 2. Bbas: Beauveria bassiana filamentous fungus 5. Replicate: Replicate number within each sample set of a given Background x Treatment x Day x Wolbachia combination 6. Eggs0.3: Eggs laid by a single female until day 3 post treatment 7. Eggs 4.6: Eggs laid by a single female from days 4-6 post treatment (not used for analysis in this study) 8. Block: Experimental replicates (A or B), with each replicate performed on different days 9. Hatch0.3: Number of eggs from a single female laid until day 3 post treatment which then hatched into larvae 10. Hatch4.6: Intended to be equivalent to Hatch0.3 for days 4-6, but data was not recorded or subsequently analyzed 11. HR0.3: The percent of eggs that hatch from a single female over days 0-3 post treatment (calculated as: Hatch0.3/Eggs0.3*100) 12. HR4.6: Intended to be equivalent to HR4.6 for days 4-6, but data was not recorded or subsequently analyzed ##### Information for File L: Fig4SFertilityw1118.csv 1. Description: Fertility data underlying Figure S4 for the w1118 host genetic background, including egg lay and egg hatch rates over different time periods. 2. Variables, data codes, formats, & abbreviations: 1. All variables and abbreviations are the same as listed for File K, except for the Background, which is w1118 here. ##### Information for File N: Fig5BbasTiterswk.csv 1. Description: Beauveria bassiana titer data underlying Figure 5 for the wk host genetic background. 2. Variables, data codes, formats, & abbreviations: 1. Vial: Number with each representing a unique biological sample. 2. Background: Genotype of the host (wk or w1118) 3. Wolbachia: Indicator of presence (I, infected) or absence (U, uninfected) of Wolbachia symbiont 4. Treatment: Hours post Beauveria bassiana infection 1. 0hr: Biological replicate of qPCR data from fly DNA extracted immediately after infection 2. 24hr: Biological replicate of qPCR data from fly DNA extracted 24 hr after infection 5. Replicate: Replicate number within each sample set of a given Background x Treatment x Day x Wolbachia combination 6. Titer: The titer calculated as 2^-deltaCt, with qPCR Ct values from B. bassiana gamma tubulin copy numbers to host rp49 copy numbers ##### Information for File O: Fig5SBbasTitersw1118.csv 1. Description: Beauveria bassiana titer data underlying Figure S5 for the w1118 host genetic background. 2. Variables, data codes, formats, & abbreviations: 1. All variables and abbreviations are the same as listed for File N, except for the Background, which is w1118 here. ##### Information for File P: Fig5SWolbTitersw1118.csv 1. Description: Wolbachia titer data underlying Figure S5 for the wk host genetic background. 2. Variables, data codes, formats, & abbreviations: 1. All variables and abbreviations are the same as listed for File N, except for the Background, which is w1118 here, and the "Titer" variable: 1. Titer: The titer calculated as 2^-deltaCt, with qPCR Ct values from Wolbachia groEL copy numbers to host rp49 copy number ##### Information for File Q: Fig5WolbTiterswk.csv 1. Description: Wolbachia titer data underlying Figure 5 for the wk host genetic background. 2. Variables, data codes, formats, & abbreviations: 1. All variables and abbreviations are the same as listed for File N, except for the "Titer" variable: 1. Titer: The titer calculated as 2^-deltaCt, with qPCR Ct values from Wolbachia groEL copy numbers to host rp49 copy number ##### Information for File R: all.longest.counts.txt 1. Description: A list of counts per transcript from the RNAseq analysis, corresponding to RNAseq analysis in the manuscript, Figures 6, S6, and 7. 2. Variables, data codes, formats, & abbreviations: 1. target_id: FlyBase Gene ID for the given gene 2. est_counts: estimated number of counts for the given gene in each sample ##### Information for File S: id\_validation\_table\_93194.txt 1. Description: ID conversion table, used to convert the FlyBase Gene ID for a given gene to specific gene acronyms, corresponding to RNAseq analysis in the manuscript, Figures 6, S6, and 7. 2. Variables, data codes, formats, & abbreviations: 1. \#submitted_item: the FlyBase Gene ID that is being converted 2. validated_id: result of FlyBase output after checking the input "submitted_item". Matches are given if validated, and a fixed ID or blank is given if not 3. current_symbol: gene acronyms specific to a given gene ##### Information for File T: RNAseq.R 1. Description: annotated R code for DESeq2 analysis corresponding to RNAseq analysis in the manuscript, Figures 6, S6, and 7. 2. Variables, data codes, formats, & abbreviations: 1. For data corresponding to each .R file, see "Data and File Overview" section, part 2. 2. Variables are either created and defined in the code or details can be read above in the Information sections below for corresponding .txt data files. ## Code/software Software and packages used: Data analysis and figure generation were performed in R v4.2.2, using several packages: coxme (v2.2.18.1), ggplot2 (v3.4.0), cowplot (v1.1.1), car (v3.1.1), SurvMiner (v0.4.9), SurvMisc (v0.5.6), DESeq2 (v1.44.0), tidyverse (v2.0.0), rtracklayer (v1.64.0), RColorBrewer (v1.1-3), edgeR (v4.2.1), EnhancedVolcano (v1.22.0), ape (v5.8), apeglm (v1.26.1), and reshape2 (v1.4.4). Exact packages used for each figure may be found in the corresponding R file. Refer to [BMC Biology article](https://bmcbiol.biomedcentral.com/articles/10.1186/s12915-025-02130-0) or citations of these packages. ## Version Changes 1. 02.14.25: Original version published 2. 03.07.25: Added files R, S, and T and associated information above in README file relating to RNAseq from the manuscript (Figures 6, S6, 7), which was missing from the prior version. Files include: R) all.longest.counts.txt, S) id_validation_table_93194.txt, T) RNAseq.R