The dynamic gut microbiota of zoophilic members of the Anopheles gambiae complex (Diptera: Culicidae)

Abstract

The gut microbiota of mosquitoes plays a critical role in the life history of the animal. There is a growing body of research characterising the gut microbiota of a range of mosquito species, but there is still a paucity of information on some members of the Anopheles gambiae complex. In this study, the gut microbiota of four laboratory strains were characterised. SENN (Anopheles arabiensis-insecticide susceptible major vector), SENN DDT (Anopheles arabiensis-insecticide resistant major vector), MAFUS (Anopheles merus-minor vector) and SANGWE (Anopheles quadriannulatus-non-vector) were used in this study. The microbiota of fourth instar larvae, 3-day old, 15-day old non-blood fed and 15-day old blood fed females were characterised by MALDI-TOF mass spectroscopy and 16 s rRNA gene sequencing by next generation sequencing. The four strains differed in species richness but not diversity. The major vectors differ in β-diversity from that of the minor and non-vectors. There was no difference in α- or β-diversity in 15 non-blood fed females and 15-day old females that had 3 blood meals before day 15. These differences may be related to a mixture of the effect of insecticide resistance phenotype as well as a potential relationship to vector competence to a limited extent. Bacterial diversity is affected by species and age. There is also a potential relationship between the differences in gut microbiota and capacity to transmit parasites. This genetic background of the mosquitoes, however, play a major role, and must be considered in this relationship.

Figure 1

Comparison of alpha diversity between the zoophilic members of the An. gambiae complex. (A) Chao1 index. (B) Abundance-based Coverage Estimator (ACE) index. (C) Shannon diversity index. (D) Simpson diversity index.

Figure 2

Rarefaction curves and beta diversity plots of the An. gambiae complex. (A) rarefaction curves of MAFUS (An. merus), SANGWE (An. quadriannulatus), SENN (An. arabiensis-insecticide susceptible) and SENN-DDT (An. arabiensis-insecticide resistant). (B) Principle Coordinates Analysis plot of beta-diversity highlighting the differences between strains. (C) Principal Component Analysis plot of beta-diversity highlighting differences between life stages.

Figure 3

Overlapping midgut bacteria identified between the different mosquito strains. The blue dots represent the number of overlaps per strain, with single dots indicating the number of unique families/genera/species per strain. (A) Comparison of common families. There are 17 common families between all four strains. (B) Comparison of common genera. There are 33 common genera between all four strains. (C) Comparison of common species. There are 34 common species between all strains.

Figure 4

Comparison of differential abundance of genera between strains. (A) Differential abundance of genera in SENN vs SANGWE at an 99% Confidence interval. (B) Differential abundance of genera in SENN-DDT vs SANGWE at a 99% Confidence interval. (C) Differential abundance of genera in MAFUS vs SANGWE at a 99% Confidence interval. (D) Differential abundance of genera in SENN vs MAFUS at a 99% Confidence interval. (E) Differential abundance of genera in SENN vs SANGWE at an 99% Confidence interval. (F) Differential abundance of genera in 15-day bloodfed SENN vs 15 -day bloodfed SENN-DDT at a 95% confidence interval. No genus was found to be significantly abundantly expressed between these two strains overall at the 99% confidence interval. The dotted line at 0 represents the change from over representation to underrepresentation of bacterial genera. The species name on either side of the dotted line where the relevant bacterial species are abundant.. Each dot represents a single OTU.