BIOCHEMICAL FINGERPRINTING OF PARASITIC SEA LICE Lepeophtheirus salmonis USING MID-INFRARED SPECTROSCOPY AND STABLE ISOTOPE ANALYSIS FOR HOST ORIGIN ASSIGNMENT

Background

The salmon louse, Lepeophtheirus salmonis, is associated with substantial losses to the salmon farming industry. These ectoparasites feed on the blood, gills, and skin of their hosts, facilitating pathogen entry and reducing immune function, leading to secondary infections. Adverse health outcomes and mass mortalities are increasing alongside the emergence of treatment resistance; therefore, the sustainability of salmon aquaculture depends on a shift towards prevention-focused sea lice control. At present, a limited understanding of sea lice transmission dynamics represents a major barrier to effective prevention. Due to the highly dispersive nature of free-living larvae and migratory salmonids, sea lice from distinct sources exhibit weak genetic differentiation (Jacobs et al., 2018). Accurate determination of host origin at individual louse level would substantially advance our understanding of sea lice transmission dynamics, relationship between farmed and wild salmonid host populations, and provide evidence base for more targeted and effective management interventions.

Alternative profiling approaches to traditional genetics, such as stable isotope analysis (SIA) and mid-infrared reflectance spectroscopy (MIR), remain underexplored for assigning the parasite origin of individual sea lice. Parasite isotopic signatures reflect host diet; initial studies suggest δ¹³C and δ¹⁵N values can distinguish farmed from wild lice due to contrasting feed sources (Butterworth et al., 2004). However, SIA analysis is costly, and studies in other organisms suggest infrared spectroscopy may provide a cost-effective proxy (Sim et al., 2023). MIR can also detect differences in parasite tissue composition associated with variation in dietary lipid and carbohydrate content. This pilot study used adult sea lice from farmed salmonid hosts (n=35) and wild salmonid hosts (n=106) in Loch Etive, Scotland, to compare the discriminatory power of SIA and MIR in determining sea lice host origin. All samples underwent carbon and nitrogen SIA and MIR spectroscopy, and a subset (n=40) also underwent low-coverage whole genome sequencing, and mitochondrial haplotype frequencies were compared to provide a genetic benchmark.

Results and Discussion

Wild- and farmed-host derived sea lice shared the same fundamental spectral peaks, reflecting identical biochemical classes, but differed consistently in absorbance intensity (figure 1). MIR spectroscopy discriminated sea lice from farmed and wild hosts using partial least square discriminant analysis with a balanced accuracy of 0.838 under leave-one-out cross-validation, confirmed as non-random by permutation testing (p<0.001). The primary discriminating signal localised to the CH2/CH3 bending region (1350–1480 cm^-1; Cohen's d = 1.311), with lice from farmed hosts exhibiting consistently higher aliphatic lipid absorbance across 100% of wavenumbers in this region after FDR correction. Supporting differences were observed in the amide II (1480–1580 cm^-1; Cohen's d = 1.024) and phosphodiester/amide III (1200–1280 cm^-1; Cohen's d = 0.719) sub-regions at 63% and 56% significance, respectively. In contrast, sea lice from wild hosts showed higher absorbance in the O–H/N–H stretching (3200–3600 cm^-1) and low-wavenumber (400–600 cm^-1) regions, consistent with greater hydration and variability in their biochemical composition. Mean isotope values differed between groups, with δ¹⁵N, δ¹³C, and δ³⁴S in farmed-origin lice depleted than in wild-origin lice by 3.92±0.15‰, 4.34±0.18‰, and 2.96±0.16‰, respectively.

These differences are consistent with expectations of a more uniform and lipid-rich diet in farmed salmonid hosts and greater environmental variability in wild salmonids. Together, this work provides proof of concept for using biochemical markers to trace sea lice origin, supporting improved understanding of transmission pathways and more targeted strategies to improve salmon health.

Figure

Figure 1. Biochemical interpretation of spectral differences between farmed and wild sea lice (Lepeoptheirus salmonis).

Acknowledgement

This study was supported by the Erasmus+: Key Action 1 – Erasmus Mundus Joint Master (EMJM) (Project No.: 101080968, Topic: ERASMUS-EDU-2022-PEX-EMJM-MOB) for the EMJMD in Aquaculture, Environment and Society STAR (EMJMD ACES-STAR), and the BBSRC project E(tive)lice: Revolutionizing Sea Lice Detection for Sustainable Salmon Farming and Conservation (Grant No. BB/Z515292/1).

References

Butterworth, K. G., Li, W., & McKinley, R. S. (2004). Carbon and nitrogen stable isotopes: A tool to differentiate between Lepeophtheirus salmonis and different salmonid host species? Aquaculture, 241(1–4), 529–538. https://doi.org/10.1016/j.aquaculture.2004.07.021

Jacobs, A., De Noia, M., Praebel, K., Kanstad-Hanssen, Ø., Paterno, M., Jackson, D., McGinnity, P., Sturm, A., Elmer, K. R., & Llewellyn, M. S. (2018). Genetic fingerprinting of salmon louse (Lepeophtheirus salmonis) populations in the North-East Atlantic using a random forest classification approach. Scientific Reports, 8, 1203. https://doi.org/10.1038/s41598-018-19323-z

Sim, J., McGoverin, C., Oey, I., Frew, R., & Kebede, B. (2023). Near-infrared reflectance spectroscopy accurately predicted isotope and elemental compositions for origin traceability of coffee. Food Chemistry, 427, 136695. https://doi.org/10.1016/j.foodchem.2023.136695