Krolicka, A. | Search bioRxiv

bioRxiv2021

Sea lice (Lepeophtherius salmonis) detection and quantification around aquaculture installations using environmental DNA

The naturally occurring ectoparasite salmon lice (Lepeophtherirus salmonis) poses a great challenge for the salmon farming industry, as well as for wild salmonids in the Northern hemisphere. To better control the infestation pressure and protect the production, there is a need to provide fish farmers with sensitive and efficient tools for rapid early detection and monitoring of the parasitic load. This can be achieved by targeting L. salmonis DNA in environmental samples. Here, we developed and tested a new L. salmonis specific DNA-based assay (qPCR assay) for detection and quantification from seawater samples using an analytical pipeline compatible with the Environmental Sample Processor (ESP) for autonomous water sample analysis of gene targets. Specificity of the L. salmonis qPCR assay was demonstrated through in-silico DNA analyses covering sequences of different L. salmonis isolates. Seawater was spiked with known numbers of nauplii and copepodite free-swimming (planktonic) stages of L. salmonis to investigate the relationship with the number of marker gene copies (MGC). Finally, field samples collected at different times of the year in the vicinity of a salmon production farm in Western Norway were analyzed for L. salmonis detection and quantification. The assay specificity was high and a high correlation between MGC and planktonic stages of L. salmonis was established in the laboratory conditions. In the field, L. salmonis DNA was consequently detected, but with MGC number below that expected for one copepodite or nauplii. We concluded that only L. salmonis tissue or eDNA residues were detected. This novel study opens for a fully automatized L. salmonis DNA quantification using ESP robotic to monitor the parasitic load, but the challenge remains the adequate sampling of a volume of seawater sufficiently large to be representative of outbreaks and load around fish farms.

ecology↗

bioRxiv2019

Discovery of functional gene markers of bacteria for monitoring hydrocarbon pollution in the marine environment - a metatranscriptomics approach

The use of natural marine bacteria as "oil sensors" for the detection of pollution events can be suggested as a novel way of monitoring oil occurrence at sea. Nucleic acid-based devices generically called genosensors are emerging as potentially promising tools for in situ detection of specific microbial marker genes suited for that purpose. Functional marker genes are particularly interesting as targets for oil-related genosensing but their identification remains a challenge. Here, seawater samples, collected in tanks with oil addition mimicking a realistic oil spill scenario, were filtered and archived by the Environmental Sample Processor (ESP), a fully robotized genosensor, and the samples were then used for post-retrieval metatranscriptomic analysis. After extraction, RNA from ESP-archived samples at start, day 4 and day 7 of the experiment was used for sequencing. Metatranscriptomics revealed that several KEGG pathways were significantly enriched in samples exposed to oil. However, these pathways were highly expressed also in the non-oil-exposed water samples, most likely as a result of the release of natural organic matter from decaying phytoplankton. Temporary peaks of aliphatic alcohol and aldehyde dehydrogenases and monoaromatic ring-degrading enzymes (e.g. ben, box, and dmp clusters) were observed on day 4 in both control and oil tanks. Few alkane 1-monooxygenase genes were upregulated on oil, mostly transcribed by families Porticoccaceae and Rhodobacteraceae, together with aromatic ring-hydroxylating dioxygenases, mostly transcribed by Rhodobacteraceae. Few transcripts from obligate hydrocarbonoclastic genera of Alcanivorax, Oleispira and Cycloclasticus, were significantly enriched in the oil-treated tank in comparison to control, and these were mostly transporters and genes involved in nitrogen and phosphorous acquisition. This study highlights the importance of seasonality, i.e., phytoplankton occurrence and senescence leading to organic compound release which can be used preferentially by bacteria over oil compounds, delaying the latter process. As a result, such seasonal effect can reduce the sensitivity of genosensing tools employing bacterial functional genes to sense oil. A better understanding of the use of natural organic matter by bacteria involved in oil-biodegradation is needed to develop an array of functional markers enabling the rapid and specific in situ detection of anthropogenic pollution.

microbiology↗