In search of the unexpected: the first finding of an islandic population of the noble crayfish Astacus astacus in Greece redefines its distribution

This contribution presents the first record of the noble crayfish Astacus astacus (Linnaeus, 1758) in Euboea (Evia) Island, Greece. The crayfish was found during a fish monitoring expedition in the upper Mesapios River, in a reach that maintains flow throughout the year, while most of the river length desiccates completely during the dry season. The finding of A. astacus in an intermittent river of Euboea Island expands the species geographical range and calls for immediate conservation actions to protect its habitat and preserve this vulnerable population.

Evaluation of the ability of cholera vibrios to form a biofilm on the surface of the chitinous shell of a crayfish by real-time PCR

Introduction. Most of the bacteria exist in natural ecosystems not in the form of free floating cells; but in the form of biofilms attached to the substrate. One of the most ecologically important substrates is chitin. Vibrio cholerae; like most members of the Vibrionaceae family; has a chitinolytic complex and can degrade chitin. The ability of V. cholerae to form a biofilm on chitinous substrates can explain the mechanism of the formation of an ecological niche for the preservation and transfer of the pathogen to new regions with the likelihood of the formation of new foci of cholera.Aim — to determine the ability of V. cholerae to form a biofilm on the chitinous shell of crayfish (Astacus astacus) by means of real-time PCR.Materials and methods. A comparative analysis of the timing of biofilm formation by V. cholerae of different serogroups and toxigenicity was carried out.Results. In the course of the study; it was found that cholera vibrios were shown to be capable of forming a biofilm regardless the serogroup and toxigenicity. However; toxigenic tcpA+ strains have a higher intensity of biofilm formation than nontoxigenic ones; in which the tcpA gene is absent.

Proximate Composition and Amino Acid Profile of the Edible Muscle of European Crayfish Astacus astacus L. Inhabiting the Orchomenos Region in Central Greece

The aim of this study was to determine the proximate composition and amino acid profile of the edible muscle of European crayfish Astacus astacus L. Animals were collected from Orchomenos region in Central Greece and muscle tissue samples were collected for chemical analysis. Total crude protein and crude lipid content of muscle tissues were determined using the Kjeldahl method and Folch’s procedure, respectively. Amino acids profile was performed by Ultra Performance Liquid Chromatographic (UPLC) determination of the acid hydrolysed muscle extract after derivatization with AccQ-Tag reagent (Waters, USA). The results showed that protein and lipid content of crayfish muscle tissue were 16.55 ± 0.4 g 100 g-1 and 0.52 ± 0.20 g 100 g-1, respectively. Also, the moisture and ash content were 80.93 ± 0.36 g 100 g-1 and 1.25 ± 0.14 g 100 g-1, respectively. Amino acids analysis of muscle tissue revealed that the highest values were those of glutamic acid (2.98 g 100 g-1) and arginine (2.19 g 100g-1) as well as those for aspartic acid (1.88 g 100 g-1), lysine (1.47 g 100 g-1) and leucine (1.33 g 100 g-1). The lowest values were found for histidine (0.35 ± 0.03 g 100 g-1) and methionine (0.47 ± 0.01 g 100 g-1). The results of this study showed that the crayfish of Orchomenos region can be considered as an important source of essential nutrients in the human diet due to its protein and essential amino acids content, which are relatively comparable to those of farmed sea bream (Sparus aurata L.), as well as to farmed and wild sea bass (Dicentrarchus labrax). However, the lipid content of Orchomenos crayfish was found to be insignificant and negligible. Overall, the protein quality of European crayfish can be considered as a main factor in selecting this species as a candidate species for European aquaculture diversification.

Environmental DNA monitoring of noble crayfish Astacus astacus: Comparison and refining of methodology

During the past decade, environmental DNA (eDNA) methodology has become an important non-invasive tool to monitor aquatic micro- and macro-organisms, including freshwater crayfish. In Europe, noble crayfish Astacus astacus is the most widespread native freshwater crayfish. However, the species is threatened in its entire distribution range. It is therefore included on the International Union for Conservation Nature (IUCN) red list, and on several national red lists. Reliable monitoring is essential for implementation of conservation measures. For crayfish, traditional population trends have been obtained from catch per unit effort (CPUE) data. In order to successfully apply and use eDNA monitoring for noble crayfish, or any species, it is a prerequisite to know the strengths and weaknesses of the applied methods and how they perform compared to traditional methodology. Sampling strategy and analysis methodology also depends on choice of species to be monitored, and which questions to be answered. Further, refinement of the employed methods may improve the detection probability for eDNA monitoring. Here we report the results from 1) a recently published study on noble crayfish eDNA monitoring (Johnsen et al. 2020) and 2) an ongoing study comparing and optimising the methods used for monitoring noble crayfish. 1) We compared eDNA monitoring (transects with ten 5L samples) with traditional trapping (transects with 50 traps) for noble crayfish in lentic habitats, in order to evaluate detection probability and if eDNA concentration correlates with relative density of crayfish. We also compared two commonly used analytical methods [quantitative real-time PCR (qPCR) and droplet digital PCR (ddPCR)] for eDNA monitoring. We found that qPCR outperformed ddPCR in detection frequency (Fig. 1), most likely due to some inhibition in the ddPCR analysis. eDNA monitoring provided reliable presence/absence data for noble crayfish, even in lakes with very low crayfish densities. Detection frequency increased with increasing CPUE (Fig. 1). However, we did not observe any correlation between relative crayfish densities and eDNA concentrations of crayfish. eDNA concentrations were consistently very low, even in lakes with very high crayfish densities. For lakes with very low crayfish densities, we estimated that ~5 samples (5L samples) are needed for 95 % detection likelihood, while for lakes with high densities 2 samples were needed. 2) We compared two eDNA sampling strategies (sampling from bottom or the surface), commonly used for crayfish or fish in Norway to investigate how both strategies perform. The sampled filters were divided and two DNA extraction protocols were evaluated (CTAB based vs Column based). We found that the DNA yield was higher from the column based DNA extraction protocol, and that eDNA concentrations from fish (brown trout Salmon trutta, northern pike Esox lucius and European perch Perca fluviatilis) were significantly higher than for crayfish. For crayfish and brown trout, there was little difference between detection probability for bottom and surface samples, while for northern pike and European perch the detection probability was higher for the bottom samples. Currently, we are analysing eDNA samples collected with glass fibre filters and NatureMetrix filters for noble crayfish in both lentic and lotic habitats and the preliminary results will be presented. We conclude that eDNA monitoring cannot substitute CPUE monitoring for freshwater crayfish, but it offers reliable presence-absence data, provided sufficient sampling efforts. Thus, it is suitable for large scale monitoring of threatened crayfish and combined with eDNA analysis of alien crayfish and diseases such as crayfish plague, this is a cost-efficient supplement offering a more holistic approach for aquatic environments and native crayfish conservation. Furthermore, the synergy effect of using collected eDNA samples from different projects to monitor additional species is substantial.

Detection of signal crayfish Pacifastacus leniusculus v1

A species-specific assay was developed and tested by Agersnap et al. (2017) Crayfish play important role in freshwater ecosystems. Noble crayfish "Astacus astacus" is threatened to extinct by non-indigenous species such as signal crayfish "Pacifastacus leniusculus".