Next-Generation Sequencing reveals relationship between the larval microbiome and food substrate in the polyphagous Queensland fruit fly

Abstract

Insects typically host substantial microbial communities (the ‘microbiome’) that can serve as a vital source of nutrients and also acts as a modulator of immune function. While recent studies have shown that diet is an important influence on the gut microbiome, very little is known about the dynamics underpinning microbial acquisition from natural food sources. Here, we addressed this gap by comparing the microbiome of larvae of the polyphagous fruit fly Bactrocera tryoni (‘Queensland fruit fly’) that were collected from five different fruit types (sapodilla [from two different localities], hog plum, pomegranate, green apple, and quince) from North-east to South-east Australia. Using Next-Generation Sequencing on the Illumina MiSeq platform, we addressed two questions: (1) what bacterial communities are available to B. tryoni larvae from different host fruit; and (2) how does the microbiome vary between B. tryoni larvae and its host fruit? The abundant bacterial taxa were similar for B. tryoni larvae from different fruit despite significant differences in the overall microbial community compositions. Our study suggests that the bacterial community structure of B. tryoni larvae is related less to the host fruit (diet) microbiome and more to vertical transfer of the microbiome during egg laying. Our findings also suggest that geographic location may play a quite limited role in structuring of larval microbiomes. This is the first study to use Next-Generation Sequencing to analyze the microbiome of B. tryoni larvae together with the host fruit, an approach that has enabled greatly increased resolution of relationships between the insect’s microbiome and that of the surrounding host tissues.

Artificial Larval Diet Mediates the Microbiome of Queensland Fruit Fly

“…Artificial larval diet strongly modulated the microbial community structure (beta diversity) across all developmental stages, but did not affect total biodiversity (as assessed by alpha diversity metrics: species richness and Shannon's diversity index). Previously, the bacterial phyla Proteobacteria and Firmicutes have commonly been reported in Qfly larvae and adults in domesticated colonies reared on a carrot diet (Deutscher et al, 2018;Majumder et al, 2019;Woruba et al, 2019), and in other fruit flies including Bactrocera neohumeralis, Bactrocera jarvisi, Bactrocera cacuminata, Bactrocera oleae, Zeugodacus tau, A. ludens, Anastrepha obliqua, Anastrepha serpentina, Anastrepha striata, and C. capitata (Morrow et al, 2015;Malacrinò et al, 2018;Ventura et al, 2018;Koskinioti et al, 2020;Noman et al, 2020). Our results are consistent with these previous findings.…”

“…For sample processing, Qfly larvae, pupae and adult flies (male and female separately) from the G5 colonies were surface sterilized using 0.5% Tween 80 (Sigma-Aldrich, St. incubated at 32 and 35 • C for 24 to 48 hr (Majumder et al, 2019(Majumder et al, , 2020. Immediately after the sterilization process, the guts of adult flies were dissected using a stereomicroscope (Leica MZ6 stereo-microscope, Leica R , Wetzlar, Germany).…”

“…PCR amplification and sequencing were performed by the Australian Genome Research Facility, University of Adelaide, Plant Genomics Centre, Hartley Grove, URRBRAE, SA 5064, AU. For bacterial identification, the V1-V3 16S rRNA region was amplified using primers 27F (5 AGAGTTTGATCMTGGCTCAG-3 ) and 519R (3 GWATTACCGCGGCKGCTG-5 ) (Lane, 1991) as used previously in Majumder et al (2019Majumder et al ( , 2020 . The resulting amplicons were measured using a fluorimeter (Thermo Fisher Scientific, North Ryde, NSW, Australia) and normalized (Fouts et al, 2012).…”

“…Some microbiota may be acquired through insect diet, and diet is hence a major exogenous factor that can directly influence the composition of the insect gut microbial community and its metabolic capabilities (Chandler et al, 2011;Broderick and Lemaitre, 2012;Mason et al, 2014;Yun et al, 2014;Majumder et al, 2019). Additionally, variation in the diet nutritional composition (protein, carbohydrate and lipids) can influence both the gut microbiome biodiversity and community structure (Broderick et al, 2004;Ravenscraft et al, 2019;Woruba et al, 2019).…”

“…In Qfly, the bacterial microbiome is largely transmitted vertically from the mother to the offspring when eggs are laid (Deutscher et al, 2018;Majumder et al, 2019). Descriptions are available characterizing the bacteria associated with wild and domesticated Qfly larvae (Deutscher et al, 2018;Majumder et al, 2019), pupae (Fitt and O'Brien, 1985) and adult flies (Thaochan et al, 2010;Morrow et al, 2015;Woruba et al, 2019). However, the effects of larval diet on changes in the Qfly microbiome through the early stages of domestication are not known.…”