Comparison of Surface Sterilization Methods for the Analysis of insect gut microbiota: Solenopsis invicta (Formicidae) as an example

Abstract

The gut microbiota of vertebrates and invertebrates has become the focus of recent research. However, current sterilizing practices need more systematic quantitative methods due to limitations caused by their minute body size. Thus, an effective sterilization process incorporating organic and inorganic methods to obtain invertebrate microbiota, particularly when evaluating smaller insects, has yet to be elucidated. This study investigated if the whole abdomen should be utilized as the material to sterilize and examined whether physical and chemical surface sterilization methods could be combined to facilitate the acquisition of gut microbiota from theimported red fire ant (Solenopsis invicta Buren, 1972). Eight methods were designed by incorporating three chemical reagents (sterile water, 2.0 mg/L NaClO, and 75% ethanol) and one physical treatment (250 nm UV). The length range of the amplified fragment in the red imported fire ant is 401-450 bp. According to the results of the GLM regression model and interaction effect model, none of these factors (sterile water, 2.0 mg/L NaClO, and 75% ethanol, 250 nm UV) were significant for statistical regression of the Chao index, and these factors did not significantly interact with each other. Based on Alpha and Beta diversity analysis, none of the methods significantly affected the diversity of insects’ gut microbiome. Finally, we suggested that it is feasible for different species of small insects to select appropriate methods according to the current situation. Still, it is best to achieve unity in the same group.