eDNA Inactivation and Biofilm Inhibition by the Polymeric Biocide Polyhexamethylene Guanidine Hydrochloride (PHMG-Cl)

The choice of effective biocides used for routine hospital practice should consider the role of disinfectants in the maintenance and development of local resistome and how they might affect antibiotic resistance gene transfer within the hospital microbial population. Currently, there is little understanding of how different biocides contribute to eDNA release that may contribute to gene transfer and subsequent environmental retention. Here, we investigated how different biocides affect the release of eDNA from mature biofilms of two opportunistic model strains Pseudomonas aeruginosa ATCC 27853 (PA) and Staphylococcus aureus ATCC 25923 (SA) and contribute to the hospital resistome in the form of surface and water contaminants and dust particles. The effect of four groups of biocides, alcohols, hydrogen peroxide, quaternary ammonium compounds, and the polymeric biocide polyhexamethylene guanidine hydrochloride (PHMG-Cl), was evaluated using PA and SA biofilms. Most biocides, except for PHMG-Cl and 70% ethanol, caused substantial eDNA release, and PHMG-Cl was found to block biofilm development when used at concentrations of 0.5% and 0.1%. This might be associated with the formation of DNA–PHMG-Cl complexes as PHMG-Cl is predicted to bind to AT base pairs by molecular docking assays. PHMG-Cl was found to bind high-molecular DNA and plasmid DNA and continued to inactivate DNA on surfaces even after 4 weeks. PHMG-Cl also effectively inactivated biofilm-associated antibiotic resistance gene eDNA released by a pan-drug-resistant Klebsiella strain, which demonstrates the potential of a polymeric biocide as a new surface-active agent to combat the spread of antibiotic resistance in hospital settings.

Mapping of crown rust (Puccinia coronata f. sp. avenae) resistance gene Pc54 and a novel quantitative trait locus effective against powdery mildew (Blumeria graminis f. sp. avenae) in the oat (Avena sativa) line Pc54

The Pc54 oat line carries the crown rust resistance gene ‘Pc54’ and an unknown gene effective against powdery mildew. In this study two recombinant inbred line populations were developed to identify the genomic locations of the two genes and producing lists of molecular markers with a potential for marker assisted selection. The RILs and parents were phenotyped for crown rust and powdery mildew in a controlled environment. They were also genotyped using the 6K Illumina Infinium iSelect oat SNP chip. Multiple interval mapping placed Pc54 on the linkage group Mrg02 (chromosome 7D) and the novel powdery mildew QTL ‘QPm.18’ on Mrg18 (chromosome 1A) both in the mapping and validating population. A total of nine and 31 significant molecular markers were identified linked with the Pc54 gene and QPm.18, respectively. Reactions to crown rust inoculations have justified separate identity of Pc54 from other genes and QTL that have previously been reported on Mrg02 except for ’qPCRFd’. Pm3 is the only powdery mildew resistance gene previously mapped on Mrg18. However, the pm3 differential line, Mostyn was susceptible to the powdery mildew race used in this study suggesting that Pm3 and QPm.18 are different genes. Determining the chromosomal locations of Pc54 and QPm.18 is helpful for better understanding the molecular mechanism of resistance to crown rust and powdery mildew in oats. Furthermore, SNPs and SSRs that are closely linked with the genes could be valuable for developing PCR based molecular markers and facilitating the utilization of these genes in oat breeding programs.

Reference genome-assisted identification of stem rust resistance gene Sr62 encoding a tandem kinase

The wild relatives and progenitors of wheat have been widely used as sources of disease resistance (R) genes. Molecular identification and characterization of these R genes facilitates their manipulation and tracking in breeding programmes. We developed a reference-quality genome assembly of the wild diploid wheat relative Aegilops sharonensis and used positional mapping, mutagenesis, RNA-Seq and transgenesis to identify the stem rust resistance gene Sr62, which was also transferred to common wheat. This gene encodes a tandem kinase, homologues of which exist across multiple taxa in the plant kingdom. Stable Sr62 transgenic wheat lines showed high levels of resistance against diverse isolates of the stem rust pathogen, highlighting the utility of Sr62 for deployment as part of a polygenic stack to maximize the durability of stem rust resistance.