Flavobacterium psychrophilum is the etiological agent of bacterial coldwater disease and the rainbow trout fry syndrome in salmonid aquaculture worldwide. However, there have been few studies into the capacity of F. psychrophilum to form biofilms and how these cellular accretions differ from planktonic cells or how they affect potential virulence. We evaluated the biofilm formation by three Chilean isolates of F. psychrophilum (LM-02-Fp, LM-06-Fp and LM-13-Fp) and two non-Chilean strains (JIP02/86 and NCMB1947T), and compared biofilm and planktonic states to obtain insights into expression differences of virulence- and biofilm-related genes (VBRGs). Our findings are based on scanning confocal laser microscopy (SCLM) and LIVE/DEAD staining, enzymatic reactions, reverse transcription-quantitative PCR (RT-qPCR) of genes encoding putative virulence factors, and transcriptomes (RNA-Seq). The LM-02-Fp and NCMB1947T strains were the strongest and weakest biofilm producers, respectively. The strong-biofilm producer showed different physiological cell states distributed in different layers of mature biofilms, whereas the NCMB1947T biofilms consisted of cells arranged in a monolayer. WGA-binding exopolysaccharides would be the main components of their corresponding extracellular matrices. Transcriptomes of F. psychrophilum NCMB1947T and LM-02-Fp were clustered by state (biofilm versus planktonic) rather than by strain, indicating important state-dependent differences in gene expression. Analysis of differentially expressed genes between states identified putative VBRGs involved in polysaccharide biosynthesis, lateral gene transfer, membrane transport (e.g. for drugs and Fe3+), sensory mechanisms, and adhesion, and indicated that about 60 % to 100 % of VBRGs involved in these processes was significantly upregulated in the biofilm state. Conversely, upregulated motility-related genes in the biofilm state were not identified, whereas a lower fraction of proteolysis-related genes (33 %) was upregulated in biofilms. In summary, F. psychrophilum strains that produce different biofilm phenotypes show global transcriptional activity in the mature biofilm state that differs significantly from their planktonic counterparts. Also, different biofilm phenotypes share a genetic potential for virulence that is transcriptionally enhanced with respect to free-living cells. Our results suggest that the F. psychrophilum biofilm lifestyle acts as a reservoir for a given set of putative virulence factors, and recommend a deeper understanding of which could help prevent recurring infections in salmonid farms.

Piscirickettsia salmonis is the causative agent of piscirickettsiosis, a disease with high socio-economic impacts for Chilean salmonid aquaculture. The identification of major environmental reservoirs for P. salmonis has long been ignored. Most microbial life occurs in biofilms, with possible implications in disease outbreaks as pathogen seed banks. Herein, we report on an in vitro analysis of biofilm formation by P. salmonis Psal-103 (LF-89-like genotype) and Psal-104 (EM-90-like genotype), the aim of which was to gain new insights into the ecological role of biofilms using multiple approaches. The cytotoxic response of the salmon head kidney cell line to P. salmonis showed interisolate differences, depending on the source of the bacterial inoculum (biofilm or planktonic). Biofilm formation showed a variable-length lag-phase, which was associated with wider fluctuations in biofilm viability. Interisolate differences in the lag phase emerged regardless of the nutritional content of the medium, but both isolates formed mature biofilms from 288 h onwards. Psal-103 biofilms were sensitive to Atlantic salmon skin mucus during early formation, whereas Psal-104 biofilms were more tolerant. The ability of P. salmonis to form viable and mucus-tolerant biofilms on plastic surfaces in seawater represents a potentially important environmental risk for the persistence and dissemination of piscirickettsiosis.

The strong seasonal variability in physical-chemical conditions of the Eastern South Pacific Ocean creates an ideal setting to study spatiotemporal distribution of key marine microbial communities. We herein report a nearly 4-year-long time series of the variability in amoA gene counts of ammonia oxidizing archaea (AOA) and bacteria (betaproteobacteria, bAOB) by quantitative PCR, GI.1a Thaumarchaeota and MG-II Euryarchaeota by CARD-FISH, and the picoplanktonic community by flow cytometry for this area. During spring-summer, non-photosynthetic picoplankton such as MG-II Euryarchaeota and GI.1a Thaumarchaeota peaked at the surface and deeper waters, respectively. General AOA and bAOB achieved higher abundances at the oxycline mainly in summer (up to 10⁵–10⁴ amoA copies mL^–1). Generalized additive models for location, scale, and shape (GAMLSS) indicated that season and depth account for 19–46% of variations in the abundance of the groups studied, particularly GI.1a Thaumarchaeota and AOA. The oxygen and nitrite concentration were statistically meaningful predictors for the studied groups. GAMLSS models indicate that ammonia oxidizing assemblage’s variability is coupled with ammonia, nitrite, and nitrate variations. Our results indicate that microbial abundances fluctuation is associated with upwelling variability and oxygen-deficient water conditions that shape the substrates availability and metabolic response of marine microbes, including keystone ammonia oxidizing assemblages and their ecological interactions. Overall, our results support planktonic nitrification activity and its contribution to nitrous oxide excess production in the time series off Concepción and the ecological dynamics regarding AOA and bAOB in coastal waters.