Virulent and necrotrophic strategies of Bacillus thuringiensis in susceptible and resistant insects, Galleria mellonella.

Bacillus thuringiensis (Bt) is one of the most common entomopathogenic bacteria used as a biopesticide, and source of endotoxin genes for generating insect-resistant transgenic plants. The mechanisms underpinning an insect's susceptibility or resistance to B. thuringiensis are diverse. The bacterial lifecycle does not end with the death of a host, they continue to exploit the cadaver to reproduce and sporulate. Herein, we studied the progression of B. thuringiensis subsp. galleriae infection in two populations of wax moth larvae (Galleria mellonella) to gain further insight into the "arms race" between B. thuringiensis virulence and insect defences. Two doses of B. thuringiensis subsp. galleriae (spore and crystalline toxin mixtures) were administered orally to compare the responses of susceptible (S) and resistant (R) populations at ∼30% mortality each. To investigate B. thuringiensis-insect antibiosis, we used a combination of in vivo infection trials, bacterial microbiome analysis, and RNAi targeting the antibacterial peptide gloverin. Within 48 h post-inoculation, B. thuringiensis-resistant insects purged the midgut of bacteria, i.e., colony forming unit numbers fell below detectable levels. Second, B. thuringiensis rapidly modulated gene expression to initiate sporulation (linked to quorum sensing) when exposed to resistant insects in contrast to susceptible G. mellonella. We reinforce earlier findings that elevated levels of antimicrobial peptides, specifically gloverin, are found in the midgut of resistant insects, which is an evolutionary strategy to combat B. thuringiensis infection via its main portal of entry. A sub-population of highly virulent B. thuringiensis can survive the enhanced immune defences of resistant G. mellonella by disrupting the midgut microbiome and switching rapidly to a necrotrophic strategy, prior to sporulation in the cadaver.

Autoclaved Diet with Inactivated Spores of Bacillus spp. Decreased Reproductive Performance of Muc2-/- and Muc2+/- Mice.

Within barrier facilities, autoclaved diet and bedding are used for husbandry of laboratory rodents. Bacillus spp. are ubiquitous in nature and some of them are known as probiotics. Inactivation of the Bacillus spores and reduction of the diet nutritional value due to autoclavation could be especially critical for immunodeficient mice. We studied the effect of the autoclaved and non-autoclaved diets on the reproductive performance and the age of prolapse manifestation in Muc2-/- mice with impaired gut barrier function and, therefore, sensitive to change of microbiota. We found that the non-autoclaved diet led to enhancement of the fertility index of Muc2-/- and Muc2+/- female mice. The non-autoclaved diet affected the prolapse of Muc2-/- mice that occurred later in comparison with females eating the autoclaved diet. We showed that Bacillus spp. was present in the non-autoclaved diet and feces of mice on the non-autoclaved diet. Bacterial strains of the non-autoclaved diet and feces belonged to B. amyloliquefaciens, B. thuringiensis, B. subtilis, Lysinibacillus macrolides, B. cereus, and other representatives of Bacillus spp. Moreover, autoclavation of the diet affected on the percent of the blood and spleen immune cells, the bacterial composition of the intestine, and increased the level of methionine in the thigh muscle of mice. Enhanced reproductive performance and delayed prolapse manifestation in Muc2-/- mice could be due to improved digestion, as Bacillus spp. from diet and feces had enzymatic activity.