Burkholderia bacteria produce multiple potentially novel molecules that inhibit carbapenem-resistant gram-negative bacterial pathogens

Antimicrobial resistance in Gram-negative pathogens represents a global threat to human health. This study determines the antimicrobial potential of a taxonomically and geographically diverse collection of 263 Burkholderia (sensu lato) isolates and applies natural product dereplication strategies to identify potentially novel molecules. Antimicrobial activity is almost exclusively present in Burkholderia sensu stricto bacteria and rarely observed in the novel genera Paraburkholderia, Caballeronia, Robbsia, Trinickia, and Mycetohabitans. Fourteen isolates show a unique spectrum of antimicrobial activity and inhibited carbapenem-resistant Gram-negative bacterial pathogens. Dereplication of the molecules present in crude spent agar extracts identifies 42 specialized metabolites, 19 of which represented potentially novel molecules. The known identified Burkholderia metabolites include toxoflavin, reumycin, pyrrolnitrin, enacyloxin, bactobolin, cepacidin, ditropolonyl sulfide, and antibiotics BN-227-F and SF 2420B, as well as the siderophores ornibactin, pyochelin, and cepabactin. Following semipreparative fractionation and activity testing, a total of five potentially novel molecules are detected in active fractions. Given the molecular formula and UV spectrum, two of those putative novel molecules are likely related to bactobolins, and another is likely related to enacyloxins. The results from this study confirm and extend the observation that Burkholderia bacteria present exciting opportunities for the discovery of potentially novel bioactive molecules

Clinical course, treatment and visual outcome of an outbreak of Burkholderia contaminans endophthalmitis following cataract surgery

Background - Postoperative endophthalmitis is a rare but dreaded complication of intraocular surgery and often results in severe visual impairment or blindness. The present study describes the clinical course, treatment and visual outcome of an outbreak of Burkholderia contaminans endophthalmitis following cataract surgery. Methods - Among 290 patients who underwent uneventful phacoemulsification cataract surgery at one outpatient clinic between January 4th and 28th 2019, 6 cases developed Burkholderia contaminans endophthalmitis. Clinical data were collected by retrospective review of patient records. Microbiological samples from vitreous aspirates, intraocular lenses (IOL) and lens capsules were cultured, and recA and draft whole genome sequences analysed. Results - The recA sequences of all Burkholderia contaminans isolates and the allelic profile of the isolates were identical. All cases had a similar clinical presentation with rapid development of endophthalmitis symptoms with variable time to onset. The mean time to admission was 34 days (12–112 days). All cases had a seemingly favourable response to intravitreal antibiotics. However, acute recurrences occurred after long time periods (12–71 days). The cases experienced between 0 and 3 recurrences. Due to persistent infection, the cases received between 5 and 15 treatments (mean 7.8) including IOL and lens capsule explantation in 5 of 6 cases. Burkholderia contaminans was detected in all explanted lens capsules. The final corrected distance visual acuity (CDVA, Snellen chart) was between 0.8 and 1.2 and all cases had final CDVA ≥0.8. Conclusions - A persistent and intensive treatment approach including total lens capsule and IOL explantation is recommended for Burkholderia contaminans endophthalmitis following cataract surgery and may lead to a favourable visual result

Burkholderia cepacia complex taxon K : where to split?

The objective of the present study was to provide an updated classification for Burkholderia cepacia complex (Bcc) taxon K isolates. A representative set of 39 taxon K isolates were analyzed through multilocus sequence typing (MLST) and phylogenomic analyses. MLST analysis revealed the presence of at least six clusters of sequence types (STs) within taxon K, two of which contain the type strains of Burkholderia contaminans (ST-102) and Burkholderia lata (ST-101), and four corresponding to the previously defined taxa Other Bcc groups C, G, H and M. This clustering was largely supported by a phylogenomic tree which revealed three main clades. Isolates of B. contaminans and of Other Bcc groups C, G, and H represented a first clade which generally shared average nucleotide identity (ANI) and average digital DNA-DNA hybridization (dDDH) values at or above the 95–96% ANI and 70% dDDH thresholds for species delineation. A second clade consisted of Other Bcc group M bacteria and of four B. lata isolates and was supported by average ANI and dDDH values of 97.2 and 76.1% within this clade and average ANI and dDDH values of 94.5 and 57.2% toward the remaining B. lata isolates (including the type strain), which represented a third clade. We therefore concluded that isolates known as Other Bcc groups C, G, and H should be classified as B. contaminans, and propose a novel species, Burkholderia aenigmatica sp. nov., to accommodate Other Bcc M and B. lata ST-98, ST-103, and ST-119 isolates. Optimized MALDI-TOF MS databases for the identification of clinical Burkholderia isolates may provide correct species-level identification for some of these bacteria but would identify most of them as B. cepacia complex. MLST facilitates species-level identification of many taxon K strains but some may require comparative genomics for accurate species-level assignment. Finally, the inclusion of Other Bcc groups C, G, and H into B. contaminans affects the phenotype of this species minimally and the proposal to classify Other Bcc group M and B. lata ST-98, ST-103, and ST-119 strains as a novel Burkholderia species is supported by a distinctive phenotype, i.e., growth at 42°C and lysine decarboxylase activity

Comparative genomics of Pandoraea, a genus enriched in xenobiotic biodegradation and metabolism

Comparative analysis of partial gyrB, recA, and gltB gene sequences of 84 Pandoraea reference strains and field isolates revealed several clusters that included no taxonomic reference strains. The gyrB, recA, and gltB phylogenetic trees were used to select 27 strains for whole-genome sequence analysis and for a comparative genomics study that also included 41 publicly available Pandoraea genome sequences. The phylogenomic analyses included a Genome BLAST Distance Phylogeny approach to calculate pairwise digital DNA-DNA hybridization values and their confidence intervals, average nucleotide identity analyses using the OrthoANIu algorithm, and a whole-genome phylogeny reconstruction based on 107 single-copy core genes using bcgTree. These analyses, along with subsequent chemotaxonomic and traditional phenotypic analyses, revealed the presence of 17 novel Pandoraea species among the strains analyzed, and allowed the identification of several unclassified Pandoraea strains reported in the literature. The genus Pandoraea has an open pan genome that includes many orthogroups in the 'Xenobiotics biodegradation and metabolism' KEGG pathway, which likely explains the enrichment of these species in polluted soils and participation in the biodegradation of complex organic substances. We propose to formally classify the 17 novel Pandoraea species as P. anapnoica sp. nov. (type strain LMG 31117(T) = CCUG 73385(T)), P. anhela sp. nov. (type strain LMG 31108(T) = CCUG 73386(T)), P. aquatica sp. nov. (type strain LMG 31011(T) = CCUG 73384(T)), P. bronchicola sp. nov. (type strain LMG 20603(T) = ATCC BAA-110(T)), P. capi sp. nov. (type strain LMG 20602(T) = ATCC BAA-109(T)), P. captiosa sp. nov. (type strain LMG 31118(T) = CCUG 73387(T)), P. cepalis sp. nov. (type strain LMG 31106(T) = CCUG 39680(T)), P. commovens sp. nov. (type strain LMG 31010(T) = CCUG 73378(T)), P. communis sp. nov. (type strain LMG 31110(T) = CCUG 73383(T)), P. eparura sp. nov. (type strain LMG 31012(T) = CCUG 73380(T)), P. horticolens sp. nov. (type strain LMG 31112(T) = CCUG 73379(T)), P. iniqua sp. nov. (type strain LMG 31009(T) = CCUG 73377(T)), P. morbifera sp. nov. (type strain LMG 31116(T) = CCUG 73389(T)), P. nosoerga sp. nov. (type strain LMG 31109(T) = CCUG 73390(T)), P. pneumonica sp. nov. (type strain LMG 31114(T) = CCUG 73388(T)), P. soli sp. nov. (type strain LMG 31014(T) = CCUG 73382(T)), and P. terrigena sp. nov. (type strain LMG 31013(T) = CCUG 73381(T))

Biosynthesis of ditropolonyl sulfide, an antibacterial compound produced by Burkholderia cepacia complex strain R-12632

Burkholderia cepacia complex strain R-12632 produces ditropolonyl sulfide, an unusual sulfur-containing tropone, via a yet-unknown biosynthetic pathway. Ditropolonyl sulfide purified from a culture of strain R-12632 inhibits the growth of various Gram-positive and Gram-negative resistant bacteria, with MIC values as low as 16 mu g/ml. In the present study, we used a transposon mutagenesis approach combined with metabolite analyses to identify the genetic basis for antibacterial activity of strain R-12632 against Gram-negative bacterial pathogens. Fifteen of the 8304 transposon mutants investigated completely lost antibacterial activity against Klebsiella pneumoniae LMG 2095. In these loss-of-activity mutants, nine genes were interrupted. Four of those genes were involved in assimilatory sulfate reduction, two were involved in phenylacetic acid (PAA) catabolism, and one was involved in glutathione metabolism. Via semipreparative fractionation and metabolite identification, it was confirmed that inactivation of the PAA degradation pathway or glutathione metabolism led to loss of ditropolonyl sulfide production. Based on earlier studies on the biosynthesis of tropolone compounds, the requirement for a functional PAA catabolic pathway for antibacterial activity in strain R-12632 indicated that this pathway likely provides the tropolone backbone for ditropolonyl sulfide. Loss of activity observed in mutants defective in assimilatory sulfate reduction and glutathione biosynthesis suggested that cysteine and glutathione are potential sources of the sulfur atom linking the two tropolone moieties. The demonstrated antibacterial activity of the unusual antibacterial compound ditropolonyl sulfide warrants further studies into its biosynthesis and biological role. IMPORTANCE Burkholderia bacteria are historically known for their biocontrol properties and have been proposed as a promising and underexplored source of bioactive specialized metabolites. Burkholderia cepacia complex strain R-12632 inhibits various Gram-positive and Gram-negative resistant pathogens and produces numerous specialized metabolites, among which is ditropolonyl sulfide. This unusual antimicrobial has been poorly studied and its biosynthetic pathway remains unknown. In the present study, we performed transposon mutagenesis of strain R-12632 and performed genome and metabolite analyses of loss-of-activity mutants to study the genetic basis for antibacterial activity. Our results indicate that phenylacetic acid catabolism, assimilatory sulfate reduction, and glutathione metabolism are necessary for ditropolonyl sulfide production. These findings contribute to understanding of the biosynthesis and biological role of this unusual antimicrobial

Outbreak of Burkholderia contaminans endophthalmitis traced to a clinic ventilation system

We describe the follow-up investigation of an outbreak of endophthalmitis due to Burkholderia contaminans following cataract surgery in a single clinic. Whole-genome sequence analysis of bacteria recovered from affected patients and the clinic identified the clinic's ventilation system as the source of infection

Burkholderia: an update on taxonomy and biotechnological potential as antibiotic producers

Burkholderia is an incredibly diverse and versatile Gram-negative genus, within which over 80 species have been formally named and multiple other genotypic groups likely represent new species. Phylogenetic analysis based on the 16S rRNA gene sequence and core genome ribosomal multilocus sequence typing analysis indicates the presence of at least three major clades within the genus. Biotechnologically, Burkholderia are well-known for their bioremediation and biopesticidal properties. Within this review, we explore the ability of Burkholderia to synthesise a wide range of antimicrobial compounds ranging from historically characterised antifungals to recently described antibacterial antibiotics with activity against multiresistant clinical pathogens. The production of multiple Burkholderia antibiotics is controlled by quorum sensing and examples of quorum sensing pathways found across the genus are discussed. The capacity for antibiotic biosynthesis and secondary metabolism encoded within Burkholderia genomes is also evaluated. Overall, Burkholderia demonstrate significant biotechnological potential as a source of novel antibiotics and bioactive secondary metabolites