Stenotrophomonas maltophilia, a ubiquitous Gram-negative γ-proteobacterium, has emerged as an important opportunistic pathogen responsible for nosocomial infections. A major characteristic of clinical isolates is their high intrinsic or acquired antibiotic resistance level. The aim of this study was to decipher the genetic determinism of antibiotic resistance among strains from different origins (i.e., natural environment and clinical origin) showing various antibiotic resistance profiles. To this purpose, we selected three strains isolated from soil collected in France or Burkina Faso that showed contrasting antibiotic resistance profiles. After whole-genome sequencing, the phylogenetic relationships of these 3 strains and 11 strains with available genome sequences were determined. Results showed that a strain’s phylogeny did not match their origin or antibiotic resistance profiles. Numerous antibiotic resistance coding genes and efflux pump operons were revealed by the genome analysis, with 57% of the identified genes not previously described. No major variation in the antibiotic resistance gene content was observed between strains irrespective of their origin and antibiotic resistance profiles. Although environmental strains generally carry as many multidrug resistant (MDR) efflux pumps as clinical strains, the absence of resistance–nodulation–division (RND) pumps (i.e., SmeABC) previously described to be specific to S. maltophilia was revealed in two environmental strains (BurA1 and PierC1). Furthermore the genome analysis of the environmental MDR strain BurA1 showed the absence of SmeABC but the presence of another putative MDR RND efflux pump, named EbyCAB on a genomic island probably acquired through horizontal gene transfer.

The occurrence of Pseudomonas aeruginosa was monitored at a broad spatial scale in French agricultural soils, from various soil types and under various land uses to evaluate the ability of soil to be a natural habitat for that species. To appreciate the impact of agricultural practices on the potential dispersion of P. aeruginosa, we further investigated the impact of organic amendment at experimental sites in France and Burkina Faso. A real-time quantitative PCR (qPCR) approach was used to analyze a set of 380 samples selected within the French RMQS (‘Réseau de Mesures de la Qualité des Sols’) soil library. In parallel, a culture-dependent approach was tested on a subset of samples. The results showed that P. aeruginosa was very rarely detected suggesting a sporadic presence of this bacterium in soils from France and Burkina Faso, whatever the structural and physico-chemical characteristics or climate. When we analyzed the impact of organic amendment on the prevalence of P. aeruginosa, we found that even if it was detectable in various manures (at levels from 103 to 105 CFU or DNA targets (g drywt)-1 of sample), it was hardly ever detected in the corresponding soils, which raises questions about its survival. The only case reports were from a vineyard soil amended with a compost of mushroom manure in Burgundy, and a few samples from two fields amended with raw urban wastes in the sub-urban area of Ouagadougou, Burkina Faso. In these soils the levels of culturable cells were below 10 CFU (g drywt)-1.

Aims To develop a qPCR approach for the detection of Pseudomonas aeruginosa in soil and manure and explore its efficacy and limitations compared with that of a classical culture-dependent approach. Methods and Results A Ps. aeruginosa ecfX qPCR assay was developed. This assay was optimized for soils of contrasting physico-chemical properties and evidenced a three-log dynamic range of detection [5 × 10⁴ – 5 × 10⁶ cells (g drywt soil)⁻¹] in inoculated microcosms. Sensitivity was determined to be around 5 × 10⁴ cells (g drywt soil)⁻¹. In parallel, the minimum detection limit was estimated in the range of 10–100 CFU (g drywt soil)⁻¹ using a culture-dependent approach based on the use of a selective medium (cetrimide agar base medium supplemented with nalidixic acid), coupled to ecfX gene amplification to confirm isolate identity. These soil samples led to the growth of abundant non-Ps. aeruginosa colonies mainly belonging to other Pseudomonas species but also some beta-Proteobacteria. These bacteria strongly impacted the detection threshold of this approach. Efficacy of these approaches was compared for Ps. aeruginosa enumeration among manure and agricultural soil samples from various sites in France, Tunisia and Burkina Faso. Conclusions The developed qPCR assay enabled a specific detection of Ps. aeruginosa in soil and manure samples. The culture-based approach was usually found more sensitive than the qPCR assay. However, abundance of non-Ps. aeruginosa species among the indigenous communities able to grow on the selective medium affected the sensitivity of this latter approach. Significance and Impact of the Study This study describes the first specific and sensitive qPCR assay for the detection and enumeration of Ps. aeruginosa in soil and manure and shows its complementarity with a culture-based approach.