Wednesday, 6 November 2019

Assessing the impact of the introduced and highly invasive Marbled Crayfish on freshwater ecosystems in Madagascar.

The Marbled Crayfish, Procambarus virginalis, is the only known Decapod Crustacean that reproduces by obligate parthenogenesis (i.e. all members of the species are female, and eggs are produced without sex). While the precise origin of the animals remains to be identified, the first record of marbled crayfish is from the German aquarium trade in 1995, indicating a very recent evolutionary origin. Based on morphological characters and genetic data, the sexually reproducing Slough Crayfish from Florida, Procambarus fallax, has been identified as the most closely related species. Additional evidence strongly suggests that marbled crayfish separated from Procambarus fallax by major genetic changes, which may have occurred as recently as 25 years ago. The combination of obligate parthenogenesis and very young evolutionary age has generated a population that can be considered a single genetic clone. In theory a lack of genetic variation severely curtails the ability of a species to adapt and proliferate. However, the genetically homogeneous Marbled Crayfish has been described as a successful invasive species in various countries. This is exemplified by the situation in Madagascar, where Marbled Crayfish were first introduced around 2005. By 2008, the animals had already spread considerably and became widely recognised in the area around the capital city Antananarivo. In 2017, Marbled Crayfish had colonised an area of approximately 100 000 km², stretching from the highland to the coast. This area includes several habitats that are not inhabited by the relatively narrowly distributed native Crayfish species of Madagascar.

In a paper publihed in the journal BMC Ecology on 6 February 2019, Ranja Andriantsoa and Sina Tönges of the Division of Epigenetics at the German Cancer Research Center, Jörn Panteleit and Kathrin Theissinger of the Institute for Environmental Sciences at the University of Koblenz-Landau, Vitor Coutinho Carneiro, also of the Division of Epigenetics at the German Cancer Research Center, Jeanne Rasamy of the Mention Zoologie et Biodiversité Animale at the Université d’Antananarivo, and Frank Lyko again of the Division of Epigenetics at the German Cancer Research Center, present the results of a study of Marbled Crayfish on freshwater ecosystems in Madagascar.

The invasiveness of Marbled Crayfish represents a key feature to define their overall impact. However, additional factors can contribute to this picture. For example, marbled crayfish may transmit the Crayfish Plague agent, the Oomycete Aphanomyces astaci, which has eradicated major Crayfish populations in Europe. On the other hand, freshwater Crayfish can also function as biocontrol agents for Human diseases, such as Schistosomiasis (Bilharzia). This is exemplified by their effects on the Snail populations that function as intermediate hosts for Schistosoma Flatworms and includes the direct predation of the Snails and the consumption of aquatic Plants that are used by the Snails for shelter, as oviposition sites and as food. Lastly, freshwater Crayfish also represent an increasingly important source of nutritional protein for Human consumption.

Despite their invasive spread, habitats of Marbled Crayfish have not been analysed systematically yet. Madagascar is characterised by a high climate and habitat diversity. The broad range of aquatic ecosystems renders the island ideally suited to better understand Marbled Crayfish ecology. Furthermore, Madagascar hosts a unique and diverse flora and fauna, including seven endemic Crayfish species (i.e. species of Crayfish found nowhere else) from the genus Astacoides, which are potentially threatened by Crayfish Plague outbreaks. Finally, Marbled Crayfish have been spreading in Madagascar for the past 10 years but their potential for Human use has not been elucidated. Andriantsoa et al. aim to provide a detailed description of the large population of Marbled Crayfish in Madagascar. Their results shed light on the animals’ ability to colonise new environments, their impact on local freshwater ecosystems and their increasing commercial importance.

The study was carried out from October 2017 to March 2018 in Madagascar. After an initial field survey that determined the distribution area of Marbled Crayfish, Andriantsoa et al. performed a more detailed follow-up analysis. We investigated five aquatic ecosystems consisting of three lentic (still water) and two lotic (running water) environments in four out of the five bioclimatic zones of Madagascar. The lentic ecosystems include a randomly selected pond in the middle of a village on the east coast (Ampasimpotsy), a lake on the highlands in the centre of a big city impacted by human activities (Ranomaimbo) and a rice field (Anjingilo) in a relatively isolated area in the south. Furthermore, two lotic habitats were included: a slow-flowing highland river located in the Ranomafana National Park rainforest (Andragnaroa) and a slow-flowing lowland river located near a large city in south-central Madagascar (Ihosy).

Map of Madagascar indicating the location of the 5 selected sites. Area colors indicate bioclimates, Dark green: humid, light green: sub-humid, grey: montane, beige: dry, brown: sub-arid. Scale bar is 100 km. Andriantsoa in Andriantsoa et al. (2019).

Pictures of habitats were taken using a Canon PowerShot D30 digital camera or a Samsung Galaxy S6 camera. For each habitat, Andriantsoa et al. chose sampling stations of 20 to 150 cm depth and 50 to 1000 cm width. Collections were done in the morning from 8.00 to 11.00 for 5 to 7 days.  Crayfish were caught without release, either by the traditional fishing tool 'tandroho' (50 cm × 30 cm × 30 cm) or with a net (200 × 400 cm) or manually in burrows. Carapace length and total length were measured using a manual calliper and weight was recorded using a portable scale with 0.1 g precision. Female sex was morphologically confirmed by the presence of the gonopores on the base of the third pair of legs and the ovaries under the carapace. Abdominal musculature samples from three animals per site were preserved in ethanol for genotyping, while soft cuticles, uropods and legs were preserved for the molecular detection of the Crayfish Plague agent Aphanomyces astaci. After data collection, all animals were sacrificed according to current Malagasy legislation, which prohibits the release or live transportation of Marbled Crayfish. All maps were drawn with QGIS.

For each sampling site, bottom sediments (mud, clay, sand) were visually identified and water temperature was recorded with a multiparameter device (Hanna Instruments HI991300) 10 cm below the surface between 8.00 and 10.00 in the morning. The same device was used to record the conductivity and the pH with a calibration at 25 °C. Neighbouring vegetation and animals co-collected in the fishing tools were recorded for each site. Finally, water samples were collected at each site, and stored in a cold and dark place. Barium, sodium, and nitrate levels were determined by Raiffeisen-Laborservice of Ormont, Germany.

Genomic DNA was isolated and purified from 100 mg abdominal musculature using a Qiagen Tissue Ruptor, followed by proteinase K digestion and ethanol precipitation. Genotyping was performed by sequencing of a polymerase chain reaction amplicon from the mitochondrial cytochrome b gene. Sequencing of polymerase chain reaction amplicons from three randomly collected animals from each of the five sites showed complete identity with the Marbled Crayfish reference sequence and thus provided genetic authentication for the populations analysed in this study.

In total, Andriantsoa et al. collected 6641 Crayfish and obtained data for 2458 animals (192–817 animals per site) Catch per hour results differed considerably between individual sites, ranging from 154 animals in Ihosy to less than 20 animals in Andragnaroa. These results suggest that population densities differ between the analysed sites. Measurements established carapace lengths that were often between 10 and 35 mm and total lengths between 30 and 80 mm. Animal weights usually ranged from 1 to 10 g. Marbled Crayfish were significantly larger and heavier in the Ihosy river as compared to the remaining sites The reasons for these differences remain to be established.

Subsequent analyses revealed remarkable differences for several fundamental ecological parameters. For example, while Marbled Crayfish were initially described to inhabit the central highland of Madagascar, the population in the Ampasimpotsy pond was located almost at sea level. In fact, Andriantsoa et al. detected Marbled Crayfish at a wide range of altitudes (3–1491 m above sealevel). Additional examples for the ability of marbled crayfish to colonise different habitats were provided by the Anjingilo Rice fields that are irrigated by thermal water and the Ranomaimbo lake located in the city centre of Antsirabe. The water of Anjinglio is characterised by a particularly high temperature of 37°C and elevated Barium concentrations, which are characteristic of thermal water. The water of the Ranomaimbo lake was characterised by particularly high conductivity levels and a high concentration of dissolved solids, such as sodium (157 mg/l) and nitrate (20 mg/l). These values reflect the high levels of pollution that are often associated with urban settlements. Taken together, our findings suggest that Marbled Crayfish can tolerate substantial variation in ecological parameters.

Ampasimpotsy coastal pond, which is located 3 m above sea level. Andriantsoa in Andriantsoa et al. (2019).

To explore the potential impact of these populations, Andriantsoa et al. first investigated their infection status with the Crayfish plague pathogen Aphanomyces astaci. A previous study had suggested low levels of Aphanomyces astaci in Marbled Crayfish populations from Germany. Andriantsoa et al. investigated 100 animals from the five study sites and from Antananarivo, where marbled crayfish were first detected on Madagascar. Quantitative polymerase chain reaction indicated undetectable or very low levels of Aphanomyces astaci DNA for the large majority (96%) of samples. Only four samples presented with somewhat higher levels. However, confirmatory sequencing and microsatellite analyses could not be carried out due to the low amounts of Aphanomyces astaci DNA in the tissues. Notably, we also found Marbled Crayfish in natural habitats of two Astacoides species: in the Andragnaroa river, in sympatry with Astacoides betsileoensis, and in a channel connected to a rice field in Sahavondronina, in sympatry with Astacoides granulimanus. Marbled Crayfish populations in those two locations have been known for at least two years, with no indications for Crayfish Plague outbreaks. Taken together, these results suggest that Marbled Crayfish do not trigger Crayfish Plague outbreaks in Madagascar.

The Anjingilo rice field. Andriantsoa in Andriantsoa et al. (2019).

Andriantsoa et al.also noticed that the distribution area of Marbled Crayfish showed a strong overlap with the freshwater Snail Biomphalaria pfeifferi, which acts as the main intermediate host of the parasitic Flatworm Schistosoma mansoni (the cause of Schistosomiasis) in Madagascar. However, they could not find Biomphalaria pfeifferi at the locations that were analysed for Marbled Crayfish, suggesting possible predation. To confirm this possibility, Andriantsoa et al. performed a laboratory experiment by placing Snails of different sizes in laboratory boxes with a single Marbled Crayfish. After the first night, all 28 Snails had been eaten by the Crayfish, and only leftover shells were found in the boxes. This suggests that Marbled Crayfish can act as effective predators of Biomphalaria pfeifferi

Laboratory experiment to test the predation of Biomphalaria pfeifferi Snails by Marbled Crayfish. Bar is 2 cm. Andriantsoa in Andriantsoa et al. (2019).

Finally, Andriantsoa et al. also addressed the emerging role of Marbled Crayfish as a food for Human consumption. Marbled Crayfish can be easily caught in rivers and ponds using traditional Malagasy fishing tools. Furthermore, the animals are also farmed in larger quantities on Rice fields. For commercial distribution, 60–80 kg of live animals are packed in large bags and then sold to consumers and/or local vendors. The measurement of 200 arbitrarily sampled animals from commercially distributed Marbled Crayfish established a median total length of 57 mm and a median weight of 4.8 g, thus illustrating the commercial relevance of relatively small animals. Marbled Crayfish currently represent an important component of the animal protein supply on local markets in all areas that were analysed in this study and are being sold both as live animals and as boiled and processed tail meat. Prices ranged from 500 to 1500 Malagasy ariary (US$0.15-0.40) per kg for live animals to 8000 Malagasy ariary (US$2.20) per kg for tail meat, which is comparable to the price of Rice, at approximately 2000 Malagasy ariary (US$0.55) per kg. Popular Marbled Crayfish foods include deep-fried beignets (fritters) and Rice with Marbled Crayfish in Tomato sauce. The increasing acceptance and popularity of Marbled Crayfish foods in Madagascar is likely to further increase their commercial demand and intentional propagation.

Marbled crayfish fishing in the Andragnaroa river, using a tandroho, a traditional Malagasy fishing tool. Andriantsoa in Andriantsoa et al. (2019).

The Marbled Crayfish is a newly emerging invasive species, but very little is known about its ecological interactions. Andriantsoa et al.'s study provides the first detailed description of habitats that were successfully colonised by Marbled Crayfish populations in Madagascar. Their findings suggest a substantial habitat diversity and thus convincingly establish the ecological plasticity of the species. The results also provide answers for additional, important questions. For example, they found no evidence for the transmission of the Crayfish Plague pathogen by Marbled Crayfish. They also show that the animals are active predators of the intermediate hosts for Human Schistosomiasis. Finally, they provide the first documentation for the commercial exploitation of Marbled Crayfish for Human consumption. Altogether, the study thus identifies key factors for the ecological assessment of this new invasive species.

Typical Marbled Crayfish-based food; Rice with Marbled Crayfish in Tomato sauce. Andriantsoa in Andriantsoa et al. (2019).

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