Coastal zones are exposed to various anthropogenic impacts, such as different types of wastewater pollution, e.g., treated wastewater discharges, leakage from sewage systems, and agricultural and urban runoff. These various inputs can introduce allochthonous organic matter and microbes, including pathogens, into the coastal marine environment. The presence of fecal bacterial indicators in the coastal environment is usually monitored using traditional culture-based methods that, however, fail to detect their uncultured representatives. We have conducted a year-around in situ survey of the pelagic microbiome of the dynamic coastal ecosystem, subjected to different anthropogenic pressures to depict the seasonal and spatial dynamics of traditional and alternative fecal bacterial indicators. To provide an insight into the environmental conditions under which bacterial indicators thrive, a suite of environmental factors and bacterial community dynamics were analyzed concurrently. Analyses of 16S rRNA amplicon sequences revealed that the coastal microbiome was primarily structured by seasonal changes regardless of the distance from the wastewater pollution sources. On the other hand, fecal bacterial indicators were not affected by seasons and accounted for up to 34% of the sequence proportion for a given sample. Even more so, traditional fecal indicator bacteria (Enterobacteriaceae) and alternative wastewater-associated bacteria (Lachnospiraceae, Ruminococcaceae, Arcobacteraceae, Pseudomonadaceae and Vibrionaceae) were part of the core coastal microbiome, i.e., present at all sampling stations. Microbial source tracking and Lagrangian particle tracking, which we employed to assess the potential pollution source, revealed the importance of riverine water as a vector for transmission of allochthonous microbes into the marine system. Further phylogenetic analysis showed that the Arcobacteraceae in our data set was affiliated with the pathogenic Arcobacter cryaerophilus, suggesting that a potential exposure risk for bacterial pathogens in anthropogenically impacted coastal zones remains. We emphasize that molecular analyses combined with statistical and oceanographic models may provide new insights for environmental health assessment and reveal the potential source and presence of microbial indicators, which are otherwise overlooked by a cultivation approach.

Background Vibrio spp. is ubiquitously present in the marine environment. Numerous Vibrio species are known pathogens affecting animal welfare and ecosystem function, causing significant economic losses. Hence, monitoring their presence accurate taxonomic identification and understanding their functional potential are important for disease control. All of this information is contained in the microorganism’s genome, which is easiest obtained from bacterial culture. Whole-genome sequencing in combination with new bioinformatics tools enables high phylogenetic resolution and provides insights into the full genetic potential of the microorganism, beyond traditional microbiological approaches. Results We assembled a complete genome of Vibrio campbellii isolated from the coastal Adriatic Sea. We showed that comparable genome sequences could be assembled from pure and non-axenic cultures (Average Nucleotide Identity value 99.98%) using a set of different bioinformatics tools, overcoming the problems of culture contamination frequently encountered in environmental microbiology. Based on the complete genome, we were able to accurately determine taxonomy, which often cannot be reliably determined with commonly applied genetic markers (e.g., 16S rRNA sequences). Pangenomic analyses with 10 reference genomes suggested that our newly assembled genome clusters in the sublineages of V. campbellii associated with diseases of marine organisms (e.g., Acute Hepatopancreatic Necrosis Disease in shrimps). Furthermore, our analysis revealed unique genomic features mainly associated with a new unique plasmid connected to bacterial defense (e.g., type I restriction-modification systems, CRISPR Csa3 proteins) and with the propagation of genetic material (e.g., transposases associated with horizontal gene transfer, ardC genes increasing plasmids host range). Conclusions Our completed genome assembly from a non-axenic culture highlights the possibility of obtaining complete genomic information when encountering contamination events, or beyond our study, even in the case of co-cultures. With this new complete V. campbellii genome we are contributing novel insights into the genomic characteristics and functional potential of this potential pathogen. In the light of projected human pressures on coastal regions and associated marine ecosystem services, pathogen surveillance and control of possible disease outbreaks are crucial. Our study reveals a novel, unique mobile element associated with V. campbellii that could potentially ease the propagation of its associated genetic material across different microbes.

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