Wet biofilms associated with medical devices have been widely studied and their link with healthcare-associated infections (HCAIs) is well recognized. Little attention has been paid to the presence of dry biofilms on environmental surfaces in healthcare settings. To investigate the occurrence, prevalence, and diversity of dry biofilms on hospital surfaces. Sixty-one terminally cleaned items were received from three different UK hospitals. The presence of dry biofilm was investigated using culture-based methods and scanning electron microscopy (SEM). Bacterial diversity within biofilms was investigated using ribosomal RNA intergenic spacer analysis (RISA)-polymerase chain reaction and next-generation sequencing. Multi-species dry biofilms were recovered from 95% of 61 samples. Abundance and complexity of dry biofilms were confirmed by SEM. All biofilms harboured Gram-positive bacteria including pathogens associated with HCAI; 58% of samples grew meticillin-resistant Staphylococcus aureus. Dry biofilms had similar physical composition regardless of the type of items sampled or the ward from which the samples originated. There were differences observed in the dominance of particular species: dry biofilms from two hospitals contained mostly staphylococcal DNA, whereas more Bacillus spp. DNA was found on surfaces from the third hospital. The presence of dry biofilms harbouring bacterial pathogens is virtually universal on commonly used items in healthcare settings. The role of dry biofilms in spreading HCAIs may be underestimated. The risk may be further exacerbated by inefficient cleaning and disinfection practices for hospital surfaces.

Pseudomonas aeruginosa causes chronic lung infections in people with cystic fibrosis (CF) and acute opportunistic infections in people without CF. Forty-two P. aeruginosa strains from a range of clinical and environmental sources were collated into a single reference strain panel to harmonise research on this diverse opportunistic pathogen. To facilitate further harmonized and comparable research on P. aeruginosa, we characterized the panel strains for growth rates, motility, virulence in the Galleria mellonella infection model, pyocyanin and alginate production, mucoid phenotype, LPS pattern, biofilm formation, urease activity, and antimicrobial and phage susceptibilities. Phenotypic diversity across the P. aeruginosa panel was apparent for all phenotypes examined, agreeing with the marked variability seen in this species. However, except for growth rate, the phenotypic diversity among strains from CF versus non-CF sources was comparable. CF strains were less virulent in the G. mellonella model than non-CF strains (P = 0.037). Transmissible CF strains generally lacked O-antigen, produced less pyocyanin and had low virulence in G. mellonella. Furthermore, in the three sets of sequential CF strains, virulence, O-antigen expression and pyocyanin production were higher in the earlier isolate compared to the isolate obtained later in infection. Overall, this full phenotypic characterization of the defined panel of P. aeruginosa strains increases our understanding of the virulence and pathogenesis of P. aeruginosa and may provide a valuable resource for the testing of novel therapies against this problematic pathogen.

An antimicrobial activity screen of Burkholderia gladioli BCC0238, a clinical isolate from a cystic fibrosis patient, led to the discovery of gladiolin, a novel macrolide antibiotic with potent activity against Mycobacterium tuberculosis H37Rv. Gladiolin is structurally related to etnangien, a highly unstable antibiotic from Sorangium cellulosum that is also active against Mycobacteria. Like etnangien, gladiolin was found to inhibit RNA polymerase, a validated drug target in M. tuberculosis. However, gladiolin lacks the highly labile hexaene moiety of etnangien and was thus found to possess significantly increased chemical stability. Moreover, gladiolin displayed low mammalian cytotoxicity and good activity against several M. tuberculosis clinical isolates, including four that are resistant to isoniazid and one that is resistant to both isoniazid and rifampicin. Overall, these data suggest that gladiolin may represent a useful starting point for the development of novel drugs to tackle multidrug-resistant tuberculosis. The B. gladioli BCC0238 genome was sequenced using Single Molecule Real Time (SMRT) technology. This resulted in four contiguous sequences: two large circular chromosomes and two smaller putative plasmids. Analysis of the chromosome sequences identified 49 putative specialized metabolite biosynthetic gene clusters. One such gene cluster, located on the smaller of the two chromosomes, encodes a trans-acyltransferase (trans-AT) polyketide synthase (PKS) multienzyme that was hypothesized to assemble gladiolin. Insertional inactivation of a gene in this cluster encoding one of the PKS subunits abrogated gladiolin production, confirming that the gene cluster is responsible for biosynthesis of the antibiotic. Comparison of the PKSs responsible for the assembly of gladiolin and etnangien showed that they possess a remarkably similar architecture, obfuscating the biosynthetic mechanisms responsible for most of the structural differences between the two metabolites.

Pseudomonas aeruginosa is a highly versatile, antibiotic-resistant Gram-negative bacterium known for causing opportunistic infections and contamination of industrial products. Despite extensive genomic analysis of clinical P. aeruginosa strains, no genomes exist for preservative-tolerant industrial strains. A unique collection of 69 industrial isolates was assembled and compared to clinical and environmental strains; 16 genetically distinct industrial strains were subjected to array tube genotyping, multilocus sequence typing and whole-genome sequencing. The industrial strains possessed high preservative tolerance and were dispersed widely across P. aeruginosa as a species, but recurrence of strains from the same lineage within specific industrial products and locations was identified. The industrial P. aeruginosa genomes (mean=7.0 Mb) were significantly larger than those of previously sequenced environmental (mean=6.5 Mb; n=19) and clinical (mean=6.6 Mb; n=66) strains. Complete sequencing of the P. aeruginosa industrial strain RW109, which encoded the largest genome (7.75 Mb), revealed a multireplicon structure including a megaplasmid (555 265 bp) and large plasmid (151 612 bp). The RW109 megaplasmid represented an emerging plasmid family conserved in seven industrial and two clinical P. aeruginosa strains, and associated with extremely stress-resilient phenotypes, including antimicrobial resistance and solvent tolerance. Here, by defining the detailed phylogenomics of P. aeruginosa industrial strains, we show that they uniquely possess multireplicon, megaplasmid-bearing genomes, and significantly greater genomic content worthy of further study.

Preventing microbial contamination of non-food products is a major area of industrial microbiology where preservatives are used to stop microbial growth. However, microorganisms occasionally overcome product preservation, causing recalls and the implementation of multiple procedures to prevent further contamination. Correct reporting of microbial contamination in non-food industrial products is vital, especially if spoilage organisms are antimicrobial resistant and pose a health threat. Gram-negative bacteria such as Pseudomonas, Burkholderia and Enterobacteriaceae are frequently reported as non-food product contaminants, including species that overlap current antimicrobial resistance priorities. Historical analysis of recall databases highlighted that for greater than 15% of contamination incidents, the causative microbial agents are reported as unidentified. Here we review the current antimicrobial resistant bacterial species associated with non-food product contamination and evaluate recall reporting in Europe from 2005 to 2018. Our review shows that 49% of microbial contaminants are reported as unidentified despite frequent detection of antimicrobial resistant pathogens; in contrast, 98% of food-related microbial contaminants are classified. Recommendations to fill this microbial identification gap in non-food product recalls are made. Overall, reporting standards for microbial contamination in non-food products must be improved to enable surveillance and for understanding the risks associated with antimicrobial resistant microorganisms