Location: Imported Fire Ant and Household Insects Research
Project Number: 6036-32000-051-027-S
Project Type: Non-Assistance Cooperative Agreement
Start Date: Sep 1, 2023
End Date: Dec 31, 2025
Objective:
The main objective of this project is to describe the ant microbial composition in Solenopsis invicta nests in Florida and other areas of the Southeast USA. To achieve this, there are 2 goals, Goal 1) Describe the bacterial community composition of fire ants in Florida and Southeast USA using 16S high through put amplicon sequencing. Goal 2) Isolate bacterial strains found in fire ants and soil from their nests and characterize them using 16S sequencing.
Approach:
Goal 1) Describe the bacterial community composition of fire ants in Florida and Southeast USA using 16S high through put amplicon sequencing.
Methods: Sampling sites will be defined in selected geographic areas in Florida and the Southeast USA where Red Imported Fire Ants occurs. To accomplish the first goal, a minimum of 100 ant nest and soil from nest will be included in this study. Ants will be collected directly on the nests by aspirators or forceps and preserved in Ethanol 95%. Soil from each nest will be collected using a sterile disposable spatula and kept frozen for preservation. Ants will be sorted and taxonomically identified by species. Ant DNA extractions will be conducted using the Puregene DNA isolation kit, while soil DNA will be obtained by using the DNeasy PowerSoil Pro Kit. DNA extractions will be subjected to sequencing using bacteria-specific taxonomic barcodes: 16S (V4 region) using high-through put sequencing. We will use bioinformatics pipeline to analyze sequences to define the fire ant core microbiome. Additional analysis will include the assessing of microbial differences among different geographic sites as well as correlation with climatic variables.
Goal 2) Isolate bacterial strains found in fire ants and soil from their nests and characterize them using 16S sequencing.
Methods: To accomplish goal 2, a set of 10 fire ant nests will be used for microbial culturing. A set of agarose plates will be inoculated with a serial dilution of ground ant material in 1XPBS, and another set of plates will be inoculated with a serial dilution of soil in sterile molecular grade water. A minimum of 5 bacterial colonies per plate will be used to obtain individual bacterial isolates that will be cultured in liquid media, reaching a total of 100 isolates. An aliquot from each isolate will be used to conduct DNA extraction followed by 16S sequencing to confirm taxonomy. Analysis will include taxonomic identification and phylogenetic analysis. These cultures will be used in future studies to assess functional role in ants or in soils from nests.
