Fig 1.
Mean Staphylococcus aureus (A) and Salmonella spp. (B) log CFU/g (mean ± SEM) during 8 days conversion in pig manure with 8-d-old black soldier fly larvae (control) and stored at 27°C, 60%-70%RH, and a photoperiod of 16:8 (L:D) h in a growth chamber.
Fig 2.
Mean Staphylococcus aureus (A) and Salmonella spp. (B). log CFU/g± SD during 8 days conversion in Duox-TLR3 RNAi larvae group with (control) 8-d-old black soldier fly larvae and stored at 27°C, 60–70%RH, and a photoperiod of 16:8 (L:D) h in a growth chamber.
Fig 3.
The response of the BsfDuox gene and BsfTLR3 gene in the gut during oral infection.
(A) Expression levels of BsfDuox at different time points in whole guts (without Malpighian tubules) after oral infection with S. aureus and Salmonella spp. (B) Expression levels of BsfTLR3 at different time points in whole guts (without Malpighian tubules) after oral infection with S. aureus and Salmonella spp. (C) The total intestinal ROS levels were quantified with flies at different time points after oral infection. Data are representative of three independent experiments (mean ± SEM). Statistical comparison was based on Student’s t–test (*p< 0.05,**p< 0.01, ***p< 0.001). Different letters indicate a significant difference in BsfDuox expression and BsfTLR3 expression and ROS levels among the oral infection with Salmonella spp. or S. aureus.
Fig 4.
(A) The expression level of Bsf dorsalgene at different time points after oral infection of S. aureus (B) The expression level of Bsf dif gene at different time points after oral infection of S. aureus.Values are the mean ± SEM of three independent experiments. Statistical comparison was based on Student’s t–test(*p< 0.05,**p< 0.01, ***p< 0.001 with Student’S t–test).
Fig 5.
Interference effects of RNAi of the BsfDuox-TLR3 gene.
(A) The expression level of the BsfDuox gene at different time points after injecting ds-Duox-TLR3 and ds-egfp. (B) The expression level of the BsfTLR3 gene at different time points after injecting ds-Duox-TLR3 and ds-egfp. (C) The total intestinal ROS levels at different time points. (D) The total gut bacterial load at different time points. Data are representative of three independent experiments (mean ± SEM). Statistical comparison was based on Student’s t–test (*p<0.05, **p<0.01,***p< 0.001).
Fig 6.
The expression level of antimicrobial peptide genes (A) Bsf cecropin, (B) Bsf ubiquitin, (C) Bsf stomoxynZH1, in black soldier fly larvae following the Duox and TLR3 RNA interference. Data are representative of three independent experiments (mean ± SEM). Statistical comparison was based on Student’s t–test (*p<0.05, **p<0.01, ***p< 0.001).
Fig 7.
(A) The beneficial symbiont in Duox-TLR3 RNAi larvae and egfp RNAi larvae at 4,8,12 days post dsRNA interfence (B) The pathogenic bacteria in Duox-TLR3 RNAi and egfp RNAi larvae at 4, 8, 12 days post dsRNA interfence. Values are the mean ± SEM of three independent experiments.
Fig 8.
BsfDuox-TLR3gene regulates the composition and structure of gut bacterial community.
(A) Taxonomic breakdown at the genus level grouped by ds-egfp and ds-Duox-TLR3 treatments. (B-G) Relative abundance of different bacterial taxa after injecting ds-Duox-TLR3and ds-egfp at 4, 8 and 12 Day. Data are representative of three independent experiments (mean+s.e.m.). Statistical comparison was based on Student’s t–test (*p<0.05). ND, not detected.
Fig 9.
All of the predicted KEGG metabolic pathways are shown at the second hierarchical level and grouped by major functional categories.
(A) KEGG_level2: Genetic information processing. (B) KEGG_level2: Matabolism.
Fig 10.
Effects of RNAi of the BsfDuox-TLR3gene on diversity metrics.
Richness measured as observed operational taxonomic units (OTUs; Sobs), Chao, ACE and Shannon indices of gut bacterial communities from different treatment at three timepoints. (A) 4 DPR. (B) 8 DPR. (C) 12 DPR. Data are representative of three independent experiments (mean ± SEM). Statistical comparison was based on Student’s t–test (p).