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Although aquaculture is a major player in current and future food production, the routine use of antibiotics provides ample ground for development of antibiotic resistance. An alternative route to disease control is the use of probiotic bacteria such as the marine bacteria Phaeobacter inhibens which produces tropodithietic acid (TDA) that inhibit pathogens without affecting the fish. Improving conditions for the formation of biofilm and TDA-synthesis is a promising avenue for biocontrol in aquaculture. In this study, the biosynthesis of TDA by Phaeobacter inhibens grown on micro-structured polymeric surfaces in micro-fluidic flow-cells is investigated. The formation of biofilms on three surface topographies; hexagonal micro-pit-arrays, hexagonal micro-pillar-arrays, and planar references is investigated. The biomass on these surfaces is measured by a non-invasive confocal microscopy 3D imaging technique, and the corresponding TDA production is monitored by liquid chromatography mass spectrometry (LC-MS) in samples collected from the outlets of the microfluidic channels. Although all surfaces support growth of P. inhibens, biomass appears to be decoupled from total TDA biosynthesis as the micro-pit-arrays generate the largest biomass while the micro-pillar-arrays produce significantly higher amounts of TDA. The findings highlight the potential for optimized micro-structured surfaces to maintain biofilms of probiotic bacteria for sustainable aquacultures.

Bacillus subtilis produces a wide range of secondary metabolites providing diverse plant growth-promoting and biocontrol abilities. These secondary metabolites include nonribosomal peptides with strong antimicrobial properties, causing either cell lysis, pore formation in fungal membranes, inhibition of certain enzymes, or bacterial protein synthesis. However, the natural products of B. subtilis are mostly studied either in laboratory strains or in individual isolates, and therefore, a comparative overview of secondary metabolites from various environmental B. subtilis strains is missing. In this study, we isolated 23 B. subtilis strains from 11 sampling sites, compared the fungal inhibition profiles of wild types and their nonribosomal peptide mutants, followed the production of targeted lipopeptides, and determined the complete genomes of 13 soil isolates. We discovered that nonribosomal peptide production varied among B. subtilis strains coisolated from the same soil samples. In vitro antagonism assays revealed that biocontrol properties depend on the targeted plant pathogenic fungus and the tested B. subtilis isolate. While plipastatin alone is sufficient to inhibit Fusarium spp., a combination of plipastatin and surfactin is required to hinder growth of Botrytis cinerea. Detailed genomic analysis revealed that altered nonribosomal peptide production profiles in specific isolates are due to missing core genes, nonsense mutation, or potentially altered gene regulation. Our study combines microbiological antagonism assays with chemical nonribosomal peptide detection and biosynthetic gene cluster predictions in diverse B. subtilis soil isolates to provide a broader overview of the secondary metabolite chemodiversity of B. subtilis. IMPORTANCE Secondary or specialized metabolites with antimicrobial activities define the biocontrol properties of microorganisms. Members of the Bacillus

Secondary metabolites provide Bacillus subtilis with increased competitiveness towards other microorganisms. In particular, nonribosomal peptides (NRPs) have an enormous antimicrobial potential by causing cell lysis, perforation of fungal membranes, enzyme inhibition, or disruption of bacterial protein synthesis. This knowledge was primarily acquired in vitro when B. subtilis was competing with other microbial monocultures. However, our understanding of the true ecological role of these small molecules is limited. In this study, we have established soil-derived semisynthetic mock communities containing 13 main genera and supplemented them with B. subtilis P5_B1 WT, the NRP-deficient strain sfp, or single-NRP mutants incapable of producing surfactin, plipastatin, or bacillaene. Through 16S amplicon sequencing, it was revealed that the invasion of NRP-producing B. subtilis strains had no major impact on the bacterial communities. Still, the abundance of the two genera Lysinibacillus and Viridibacillus was reduced. Interestingly, this effect was diminished in communities supplemented with the NRP-deficient strain. Growth profiling of Lysinibacillus fusiformis M5 exposed to either spent media of the B. subtilis strains or pure surfactin indicated the sensitivity of this strain towards the biosurfactant surfactin. Our study provides a more in-depth insight into the influence of B. subtilis NRPs on semisynthetic bacterial communities and helps to understand their ecological role.

BACKGROUND: Anterior cruciate ligament reconstruction (ACL-RC) is often associated with moderate to severe postoperative pain even with a multimodal analgesic regimen. We aimed to compare the analgesic efficacy of low volume saphenous-obturator block with placebo and femoral-obturator block in patients undergoing ACL-RC.METHODS: In a randomized controlled trial eighty-two patients undergoing ACL-RC with hamstring autograft were allocated to either low volume saphenous-obturator block, placebo block or femoral-obturator block. Ropivacaine 0.75% was used for active blocks and saline for placebo. Primary outcome was pain-scores at rest quantified as area-under-the-curve 0-6 hr postoperatively. Secondary outcomes were postoperative opioid consumption and pain localization in the knee.RESULTS: No statistical difference existed between groups in area-under-the-curve 6 hr pain-scores. However, pain-scores were significantly lower in the two ropivacaine groups compared to placebo at emergence t=0 (P<0.018), at t=5 (P<0.042) and at t=6 hours (P<0.002) postoperatively. Furthermore, ropivacaine blocks exhibited significantly reduced total opioid consumption (15.81 and 18.44 mg) postoperatively compared with placebo (26.38 mg) (P<0.018). Patients receiving ropivacaine blocks localized pain in the posterolateral knee, whereas placebo block patients reported anteromedial and central pain. Other secondary outcomes were similar between groups.CONCLUSIONS: Low volume saphenous-obturator block is significantly more effective than placebo in reducing both early and late pain-scores as well as postoperative opioid consumption in patients undergoing ACL-RC. No statistical difference existed when comparing low volume saphenous-obturator block to femoral-obturator block regarding early and late pain-scores and postoperative opioid consumption.