The antimicrobial potential of Streptomyces from insect microbiomes

Abstract

Antimicrobial resistance is a global health crisis and few novel antimicrobials have been discovered in recent decades. Natural products, particularly from Streptomyces, are the source of most antimicrobials, yet discovery campaigns focusing on Streptomyces from the soil largely rediscover known compounds. Investigation of understudied and symbiotic sources has seen some success, yet no studies have systematically explored microbiomes for antimicrobials. Here we assess the distinct evolutionary lineages of Streptomyces from insect microbiomes as a source of new antimicrobials through large-scale isolations, bioactivity assays, genomics, metabolomics, and in vivo infection models. Insect-associated Streptomyces inhibit antimicrobial-resistant pathogens more than soil Streptomyces. Genomics and metabolomics reveal their diverse biosynthetic capabilities. Further, we describe cyphomycin, a new molecule active against multidrug resistant fungal pathogens. The evolutionary trajectories of Streptomyces from the insect microbiome influence their biosynthetic potential and ability to inhibit resistant pathogens, supporting the promise of this source in augmenting future antimicrobial discovery.

Fig. 1

Sampling strategy for Streptomyces from insect microbiomes. Streptomyces were isolated from a wide range of insects and geographies (1445 insects; 10,178 strains; dot size, insects sampled). Streptomyces production of the antifungal mycangimycin (1) in the Southern Pine Beetle system is shown at right. Cyphomycin (2) is a new antifungal described herein. Photo credits: southern pine beetle - Erich G. Vallery; fungus-growing ant – Alexander Wild

Fig. 2

Distinct lineages of Streptomyces associate with insect hosts. a A genomic phylogeny constructed from 93 single-copy core bacterial genes is shown to the left with the major clades, Clade I (C–I) and Clade II (C-II), labeled (B = Basal, S = Streptomyces). 16S sequences were mapped to the genomic phylogeny and the distribution of free-living (tan) and insect-associated (blue) strains is shown as both pie and bar charts to the right. Number of total strains is shown to left of pies. b–f A more detailed mapping of 16S sequences onto the genomic tree is shown for clades S01 (b), S06 (c), S07 (d), S08 (e), and S13 (f). g–j 16S phylogenies from sequences mapped to clade S06 (g), S07 (h), S08 (i), and S13 (j)

Fig. 3

Bioactivity of insect-associated Streptomyces. a Fungal and b Gram-negative pathogens are significantly more inhibited by insect-associated isolates compared to soil- and plant-sourced Streptomyces (n = 1162, 186, and 178 for insect, soil, and plant, respectively; ***p < 1e−3; **p < 1e−2; t-test, BY correction). c Strains vary in antimicrobial bioactivity by insect host orders (n = 87, 69, 327, 518, 94, and 39 for Blattodea, Coleoptera, Diptera, Hymenoptera, Lepidoptera, and Orthoptera, respectively). a–c: center, median; box, upper and lower quantiles; notches, 95% confidence; whiskers, 1.5× interquartile range; points, outliers. d Hit rate for insect, soil, and plant strains against individual pathogens (n = 1162, 186, and 178 for insect, soil, and plant, respectively)

Fig. 4

Ecology and phylogeny influence biosynthetic potential. a A core-genome phylogeny shows evolutionarily distinct lineages of Streptomyces associate with insects (subset shown, see Supplementary Figure 2). BGC similarity to known BGCs highlights the biosynthetic diversity of insect microbiome strains. b Source invariant (blue) and sub-clade invariant (red) BGC families suggest BGC presence is influenced by both source and phylogeny. LC/MS metabolomics revealed MFs that are unique to c source and d phylogeny. e PCA of the metabolomes identified an outlier strain and f MFs that contribute to its uniqueness, including cyphomycin

Fig. 5

Insect-associated Streptomyces are a source of active antimicrobials. a Fractionated extracts from insect microbiomes are active in multiple murine models of drug-resistant infection. Less infective burden is seen in intraperitoneally treated mice after 8 h of infection. Each dot represents a unique fraction in one mouse study. (n = 15, 11, and 8 for C. albicans, E. coli, and P. aeruginosa models, respectively; center, median; box, upper and lower quantiles; whiskers, 1.5× interquartile range. b Most fractions from insect microbiomes show no hemolysis in cell-based assays. Safe indicates no toxicity at >100× concentration associated with efficacy. c The antifungal cyphomycin is produced by Streptomyces isolated from d the fungus-growing ant Cyphomyrmex sp. Photo credit: Alexander Wild e Cyphomycin-containing fractions show potency against the ant pathogen Escovopsis sp. (top left, bottom). f Purified cyphomycin exhibits potency against resistant pathogens. g Mouse candidiasis (C. albicans) models showcase reduced infection and a dose-like response to cyphomycin. Dots indicate individual mice