Metabarcoding, direct stomach observation and stable isotope analysis reveal a highly diverse diet for the invasive green crab in Atlantic Patagonia

The European green crab Carcinus maenas and its sister species C. aestuarii are highly invasive species causing damage to coastal ecosystems and contributing to severe economic losses worldwide. C. maenas was first detected at the Atlantic Patagonian coast in 2001. In this work, we studied the diet of the green crab in a recently invaded location in Golfo Nuevo, using three complementary techniques: direct stomach observation, metabarcoding of gut content and stable isotope analysis. Direct stomach observation and metabarcoding showed that green crabs have a broad omnivorous diet, ingesting most of the phyla present in the study area. Gut content metabarcoding allowed a detailed description of algal diversity and revealed other taxa that went unnoticed in the visual stomach analysis. Stable isotope analysis showed that the major contribution to the crabs' diet was from the phytoplankton chain (by bivalve consumption) and not directly from algae. This study approach combining three complementary techniques also allowed us to detect some differences in the diet between sexes, which suggests that male and female crabs are not as ecologically equivalent as previously thought. Besides, we detected sequences corresponding to C. aestuarii suggesting that the green crab Patagonian population is a hybrid of both sister species. These findings are key to understanding the impacts green crabs can have on the local ecosystem.Fil: Cordone, Georgina Florencia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Centro Nacional Patagónico. Centro para el Estudio de Sistemas Marinos; ArgentinaFil: Lozada, Mariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Centro Nacional Patagónico. Instituto de Biología de Organismos Marinos; ArgentinaFil: Vilacoba, Elisabet. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Museo Argentino de Ciencias Naturales "Bernardino Rivadavia"; ArgentinaFil: Thalinger, Bettina. University of Guelph; CanadáFil: Bigatti, Gregorio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Centro Nacional Patagónico. Instituto de Biología de Organismos Marinos; ArgentinaFil: Lijtmaer, Dario Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Museo Argentino de Ciencias Naturales "Bernardino Rivadavia"; ArgentinaFil: Steinke, Dirk. University of Guelph; CanadáFil: Galvan, David Edgardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Centro Nacional Patagónico. Centro para el Estudio de Sistemas Marinos; Argentin

Rapid Plant Identification Using Species- and Group-Specific Primers Targeting Chloroplast DNA

Plant identification is challenging when no morphologically assignable parts are available. There is a lack of broadly applicable methods for identifying plants in this situation, for example when roots grow in mixture and for decayed or semi-digested plant material. These difficulties have also impeded the progress made in ecological disciplines such as soil- and trophic ecology. Here, a PCR-based approach is presented which allows identifying a variety of plant taxa commonly occurring in Central European agricultural land. Based on the trnT-F cpDNA region, PCR assays were developed to identify two plant families (Poaceae and Apiaceae), the genera Trifolium and Plantago, and nine plant species: Achillea millefolium, Fagopyrum esculentum, Lolium perenne, Lupinus angustifolius, Phaseolus coccineus, Sinapis alba, Taraxacum officinale, Triticum aestivum, and Zea mays. These assays allowed identification of plants based on size-specific amplicons ranging from 116 bp to 381 bp. Their specificity and sensitivity was consistently high, enabling the detection of small amounts of plant DNA, for example, in decaying plant material and in the intestine or faeces of herbivores. To increase the efficacy of identifying plant species from large number of samples, specific primers were combined in multiplex PCRs, allowing screening for multiple species within a single reaction. The molecular assays outlined here will be applicable manifold, such as for root- and leaf litter identification, botanical trace evidence, and the analysis of herbivory

faeces_molecular_prey_detection

The excel table "faeces_molecular_prey_detection" shows the results of the molecular screening of cormorant faeces produced in the feeding trial

consumed fish

An excel table of all fish individuals consumed by the four cormorants during the three experimental runs including fish species, total length, mass and start/end of consumption per fish species and cormorant

Data from: The influence of meal size on prey DNA detectability in piscivorous birds

Molecular methods allow non-invasive assessment of vertebrate predator-prey systems at high taxonomic resolution by examining dietary samples such as faeces and pellets. To facilitate the interpretation of field-derived data, feeding trials, investigating the impacts of biological, methodological, and environmental factors on prey DNA detection have been conducted. The effect of meal size, however, has not yet been explicitly considered for vertebrate consumers. Moreover, different non-invasively obtained sample types remain to be compared in such experiments. Here, we present a feeding trial on abundant piscivorous birds, Great Cormorants (Phalacrocorax carbo), to assess meal size effects on post-feeding prey DNA detection success. Faeces and pellets were sampled twice a day after the feed of large (350-540 g), medium (190-345 g), and small (15-170 g) fish meals contributing either a large (>79%) or small (<38%) share to the daily consumption. Samples were examined for prey DNA and fish hard parts. Molecular analysis of faeces revealed that both large meal size and share had a significantly positive effect on prey DNA detection rate post-feeding. Furthermore, large meals were detectable for a significantly longer time span with a detection limit at ~76 h and a 50% detection probability at ~32 h post-feeding. In pellets, molecular methods reliably identified the meal consumed the previous day, which was not possible via morphological analysis or when examining individual faeces. The less reliable prey DNA detection of small meals or meal shares in faeces signifies the importance of large numbers of dietary samples to obtain reliable trophic data

Unravelling bird nest arthropod community structure using metabarcoding

Bird nests are fascinating microcosms harboring a wide range of arthropods parasitizing the nesting birds or feeding on prey remains, feces, and the nest material. Studies of these communities have been entirely based on emergence traps which collect live organisms out of the nests. The analysis of nest contents and environmental DNA (eDNA) via metabarcoding could expand our knowledge and identify prey, exuviae, and other animal remains in bird nests. Here, we investigated the potential of arthropod remains, nest dust, and feathers to better describe arthropod diversity accumulated in 20 bird nests collected in Guelph (Canada). We used subsampling strategies and tested two extraction approaches to investigate the distribution of DNA in nests, account for low-quality DNA, and the presence of inhibitory substances. In total, 103 taxa were detected via metabarcoding. Arthropod remains delivered the highest number of taxa (n = 67), followed by nest dust (n = 29). Extractions with the PowerSoil kit outperformed DNeasy extractions coupled with PowerClean Pro inhibitor removal. Per nest, on average 5.5% taxonomic overlap between arthropod remains of different size classes was detected and subsamples of nest dust extracted with the PowerSoil kit showed 47.3% taxonomic overlap indicating a heterogeneous eDNA distribution in nests. Most detected species were either feeding in the nest, i.e., herbivorous / predatory, or bird food. We also detected molecular traces of 25 bird species, whose feathers were likely used as nest material. Consequently, the metabarcoding of bird nest materials provides a more complete picture of nest communities, which can enable future studies on functional diversity and better comparisons between nesting species

Alignment CHD1 cormorant

Sequence alignment with male (CHD1Z) and female (CHD1W) sequences of the chromodomain-helicase-DNA-binding protein 1 (CHD1) in 5’ – 3’ direction, including reference sequences from GenBank (two for CHD1Z and CHD1W each). sequences were obtained from DNA extracts of either muscle tissue of shot cormorants (K-samples) or regurgitate pellet samples collected between 29th March and 10th May 2012 at Chiemsee (S-samples)

molecular sexing molecular fish ID hard part analysis

data based on cormorant pellets collected in the field; excel file constisting of 2 tables: Table 1 has four parts providing per pellt: 1. molecular sexing of the pellets including peak heigth in capillary electrophoresis (RFU); females with 2 peaks, males with one peak; 2. molecular fish identification from pellets (presence absence data); fish species as column headers; 3. morphological analysis (count data) of fish hard parts identifiable to species level and measurable i.e. suitable for fish-length regression analysis; 4. morphological analysis: all hard parts including non-measurable ones and not to species identifiable; Table 2 contains per pellet the mean fish length (mm) per fish specie

Data from: Molecular prey identification in Central European piscivores

Diet analysis is an important aspect when investigating the ecology of fish-eating animals and essential for assessing their functional role in food webs across aquatic and terrestrial ecosystems. The identification of fish remains in dietary samples, however, can be time-consuming and unsatisfying using conventional morphological analysis of prey remains. Here, we present a two-step multiplex PCR system, comprised of six assays, allowing for rapid, sensitive and specific detection of fish DNA in dietary samples. This approach encompasses 78 fish and lamprey species native to Central European freshwaters and enables the identification of 31 species, six genera, two families, two orders and two fish family clusters. All targeted taxa were successfully amplified from 25 template molecules, and each assay was specific when tested against a wide range of invertebrates and vertebrates inhabiting aquatic environments. The applicability of the multiplex PCR system was evaluated in a feeding trial, wherein it outperformed morphological prey analysis regarding species-specific prey identification in faeces of Eurasian otters. Additionally, a wide spectrum of fish species was detected in field-collected faecal samples and regurgitated pellets of Common Kingfishers and Great Cormorants, demonstrating the broad applicability of the approach. In conclusion, this multiplex PCR system provides an efficient, easy to use and cost-effective tool for assessing the trophic ecology of piscivores in Central Europe. Furthermore, the multiplex PCRs and the primers described therein will be applicable wherever DNA of the targeted fish species needs to be detected at high sensitivity and specificity