Suppression of cytoplasmic incompatibility in the leaf-mining fly Liriomyza sativae with a nuclear Wolbachia insert
Data files
Apr 28, 2025 version files 25.23 KB
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crossing_experiment_results.csv
4.48 KB
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Liriomyza_amino_acid_sequences.fasta
4.37 KB
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Liriomyza_DNA_sequences.fasta
10.60 KB
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README.md
5.78 KB
Abstract
Cytoplasmic incompatibility (CI) drives maternally transmitted endosymbionts such as Wolbachia through insect populations by inducing embryonic mortality when infected males fertilize uninfected females. CI is controlled by Wolbachia cif operons that are categorized into multiple phylogenetic Types. CI strength is further shaped by poorly understood host factors, including development and genetic background. To study the strength of CI across different host species, we genotyped a Japanese field population of Liriomyza sativae. By uncovering paternal transmission of Wolbachia genic elements, we collected strong evidence of horizontal genome transfer, including Type I and Type V cif operons, from Wolbachia into the nuclear genome of L. sativae. We established a transinfection of wLtri in L. sativae, a Wolbachia variant that induces strong CI in Liriomyza trifolii. No CI was observed in both intraspecific and interspecific reciprocal crosses with L. trifolii, suggesting that both uninfected females and infected males of L. sativae completely suppress wLtri-mediated CI. Our results raise the appealing hypothesis that host suppression of Wolbachia-induced CI might evolve due to horizontal transfer of cif operons into the host nuclear genome.
https://doi.org/10.5061/dryad.41ns1rnrg
Description of the data and file structure
This dataset contains reproductive and Wolbachia transmission data from crossing experiments between infected and uninfected strains of Liriomyza trifolii and Liriomyza sativae. It includes egg counts, hatching rates, emergence rates, sex ratios, and Wolbachia infection rates.
Additionally, the dataset comprises nucleotide sequences of fbpA, cifA, cifB, clpA, and pyrD, obtained through PCR amplification and Sanger sequencing. These data were collected to investigate the genetic composition and functional characteristics of Wolbachia genes, with a focus on horizontal gene transfer and cytoplasmic incompatibility mechanisms. This dataset provides insights into Wolbachia -host interactions and their evolutionary implications.
Files and variables
File: crossing_experiment_results.csv
Description:
Variables
- Cross_ID: Unique identifier for each cross. Lt means L. trifolii, Ls means L. sativae. "+" and "-" indicate the Wolbachia infection status of Host ("+": Infected, "-": Uninfected).
- Egg_Count: Number of eggs laid
- Hatch_Count: Number of larvae hatched
- Pupae_Count: Number of pupae formed
- Adult_Count: Number of emerged adults
- Female_Count: Number of female adults
- Male_Count: Number of male adults
- Wolbachia_Confirmed_F1: Number of Wolbachia infected F1 generation.
File: Liriomyza_amino_acid_sequences.fasta
Description: Translated amino acid sequences of the genes.
File: Liriomyza_DNA_sequences.fasta
Description: Nucleotide sequences of the Wolbachia genes from Liriomyza sativae (uninfected strain) and Wolbachia endosymbiont of Liriomyza trifolii.
This dataset includes the following sequence files in FASTA format, each containing nucleotide sequences of Wolbachia genes from different host strains and experimental conditions.
Multi Fasta File Structure
Each sequence follows the standard FASTA format, where:
The header line starts with ">" and contains:
- The species or strain name
- The gene name
- The GenBank accession number
Variables and Abbreviations
The dataset uses the following definitions:
Wolbachia endosymbiont of Liriomyza trifolii: The Wolbachia strain from a Liriomyza trifolii
Units of measurement:
Nucleotide sequences are represented in 5' to 3' direction using standard IUPAC nucleotide codes.
Amino acid sequences follow one-letter amino acid codes.
Code/software
File: crossing_experiment_results.csv
Methods for Data Collection
- Crossing experiments were conducted using unmated individuals of the Liriomyza species.
- One female and two males were placed in test tubes for 24 hours, after which egg counts were recorded.
- The eggs were monitored for hatching, and emerging adults were sexed and tested for Wolbachia infection using PCR.
Data Processing
- The dataset represents raw data collected from the experiments.
- Statistical analyses were conducted using R (version 4.2.1).
- Wolbachia infection was confirmed using the wsp gene PCR.
File: Liriomyza_amino_acid_sequences.fasta
File: Liriomyza_DNA_sequences.fasta
Software Required to View and Analyze Data: The dataset consists of FASTA files, which can be viewed and analyzed using widely available free or open-source software.
1. Viewing and Editing FASTA Files
FASTA format sequences can be opened and analyzed using:
- BioEdit (Version 7.2.5 or later) – A free sequence alignment editor for Windows.
- MEGA (Version 11 or later) – Used for sequence alignment and phylogenetic analysis.
- AliView (Version 1.28 or later) – A lightweight alignment viewer.
- Any standard text editor (e.g., Notepad++, VS Code, or nano).
Sequence Analysis and Alignment
- ClustalW (Version 2.1) – Used for multiple sequence alignment.
- ExPASy Translate Tool (ExPASy Server) – Used for translating nucleotide sequences into amino acid sequences.
Statistical and Bioinformatics Analysis
- R (Version 4.2.1 or later) – Used for data analysis, visualization, and statistical tests.
- Relevant R packages: tidyverse (data handling), ggplot2 (visualization)
2. Primer Design
- Primer3 Plus (Online tool) – Used to design primers for target genes.
3. PCR and Sequencing
- Sequences were obtained using Sanger sequencing at Fasmac, Inc.
- Raw sequence data were checked and trimmed.
- ExPASy Translate Tool was used for protein sequence prediction.
Access information
File: crossing_experiment_results.csv
Other publicly accessible locations of the data:
- This dataset is exclusively available on Dryad and has not been deposited in any other public repository.
Data was derived from the following sources:
- All data in this dataset were collected and generated as part of this study.
Other publicly accessible locations of the data:
- GenBank: The accession number for accessing each sequence is listed at the end of the header line for each sequence.
Data was derived from the following sources:
- This dataset was generated as part of this study. However, the Wolbachia genome sequences used for primer design were obtained from: Pramono, A. K., Hidayanti, A. K., Tagami, Y., & Ando, H. (2024). Bacterial community and genome analysis of cytoplasmic incompatibility-inducing Wolbachia in American serpentine leafminer, Liriomyza trifolii. Frontiers in Microbiology, 15, 1304401. DOI: https://doi.org/10.3389/fmicb.2024.1304401
crossing_experiment_results.csv:
The dataset was collected through controlled crossing experiments involving four different strains of Liriomyza trifolii and Liriomyza sativae with varying Wolbachia infection statuses. Below is a breakdown of the data collection and processing methods:
1. Experimental Design & Data Collection
Strains Used:
L. trifolii (Wolbachia-infected and antibiotic-treated uninfected strains)
L. sativae (Wolbachia-transinfected and uninfected strains)
Crossing Design:
A full factorial crossing experiment was performed, yielding 16 different combinations.
Individual Selection & Handling:
Only unmated individuals were used, ensuring standardization of reproductive conditions.
L. trifolii uninfected individuals were used from the third generation after antibiotic treatment to avoid direct antibiotic effects.
L. sativae infected individuals were used from the third generation after Wolbachia transinfection via microinjection.
Mating Conditions:
1 female + 2 males per test tube (7.5 cm × 1.0 cm; Thermo Fisher Scientific).
Individuals were placed in tubes for 24 hours with 50% honey in water as food.
Egg Collection & Rearing:
Quail bean leaves were used as oviposition substrates.
Eggs were counted after 24 hours and then transferred to a conical flask with water.
After hatching, larvae were moved to square acrylic cases (9.5 cm × 6.5 cm × 6.5 cm) with mesh for further development.
Counts recorded: Number of eggs, hatched larvae, pupae, and emerging adults.
Adults were sexed and counted upon emergence.
2. Wolbachia Infection Confirmation
Diagnostic PCR using wsp primers was performed on:
Parental generation (P) individuals before crossing.
Offspring (F1) to confirm vertical transmission of Wolbachia.
3. Environmental Conditions
Controlled in a 16L:8D photoperiod at 25°C to standardize environmental effects.
4. Data Processing & Statistical Analysis
Measured Parameters:
Number of eggs laid per female.
Hatching rate (larvae/eggs).
Emergence rate (adults/larvae).
Sex ratio (female/male proportion in F1).
Wolbachia infection rate in offspring.
Liriomyza_DNA_sequences.csv, Liriomyza_amino_acid_sequences.csv:
This dataset contains nucleotide sequence data related to Wolbachia genes, including fbpA, cifA, cifB, clpA, and pyrD, from Liriomyza trifolii, Liriomyza sativae. The dataset also includes predicted amino acid sequences. These data were generated as part of a study investigating Wolbachia horizontal gene transfer and cytoplasmic incompatibility mechanisms in leaf miner flies. The sequence data were obtained from PCR amplification of the target genes followed by Sanger sequencing.
Methods: How was this dataset collected?
Primer Design & PCR Amplification
Primers for fbpA, cifA, cifB, clpA, and pyrD were designed using Primer3 Plus, based on a previously published fragmented wLtri genome assembly.
PCR reaction mixtures (50 µL) contained:
KOD One® PCR Master Mix -Blue- (Takara) (25 µL)
Distilled water (21 µL)
Primers (1.5 µL each, forward & reverse)
DNA extract (1.0 µL)
PCR conditions:
Initial denaturation: 98°C for 2 min
30 cycles of:
98°C for 10 sec
53°C for 5 sec
68°C for 3 sec
Final extension: 68°C for 10 min
PCR products were verified via 1% agarose gel electrophoresis.
Sequencing & Processing
PCR products were submitted to Fasmac, Inc. (Kanagawa, Japan) for Sanger sequencing.
Raw sequence data were quality-checked and trimmed. The ExPASy Translate Tool was used to predict amino acid sequences from nucleotide sequences.