New universal ITS2 primers for high-resolution herbivory analyses using DNA metabarcoding in both tropical and temperate zones

DNA metabarcoding is a rapidly growing technique for obtaining detailed dietary information. Current metabarcoding methods for herbivory, using a single locus, can lack taxonomic resolution for some applications. We present novel primers for the second internal transcribed spacer of nuclear ribosomal DNA (ITS2) designed for dietary studies in Mauritius and the UK, which have the potential to give unrivalled taxonomic coverage and resolution from a short-amplicon barcode. In silico testing used three databases of plant ITS2 sequences from UK and Mauritian floras (native and introduced) totalling 6561 sequences from 1790 species across 174 families. Our primers were well-matched in silico to 88% of species, providing taxonomic resolution of 86.1%, 99.4% and 99.9% at the species, genus and family levels, respectively. In vitro, the primers amplified 99% of Mauritian (n = 169) and 100% of UK (n = 33) species, and co-amplified multiple plant species from degraded faecal DNA from reptiles and birds in two case studies. For the ITS2 region, we advocate taxonomic assignment based on best sequence match instead of a clustering approach. With short amplicons of 187–387 bp, these primers are suitable for metabarcoding plant DNA from faecal samples, across a broad geographic range, whilst delivering unparalleled taxonomic resolution

Observational and metabarcoding approaches reveal the ecology, natural history and conservation status of Scolopendra abnormis, a threatened centipede endemic to Mauritius

The Serpent Island centipede Scolopendra abnormis is a threatened centipede species found on only 2 small islands in the Indian Ocean: Round Island, located 22.5 km northeast of Mauritius, and Serpent Island, 4 km northwest of Round Island. Current understanding of its ecology is based on limited direct observations from 30 yr ago. Round Island has since undergone significant habitat restoration. Hyperabundant non-native ants are also present, which may impact centipede nesting behaviour, ecology, and survival. Recent methodological advances, such as high-throughput sequencing of dietary DNA, can extend our understanding of invertebrate ecology and provide data complementary to direct observation. Using a combination of dietary metabarcoding and observational approaches, we provide new insights into the ecology and natural history of this threatened invertebrate predator. S. abnormis nest most consistently in the root network found beneath endemic Pandanus vandermeeschii trees. They are also found in areas with good soil cover, herbaceous growth, and areas of bare rock slab. Only 4 of 43 centipedes in this study were found near an ant foraging trail, which may have significant implications for S. abnormis nesting habits. These centipedes primarily consume insect prey (particularly taxa within Lepidoptera, Hymenoptera, Diptera), irrespective of centipede body size. A quarter of centipedes also consumed endemic lizards. We also found marked differences in diet composition between wet and dry seasons arising from the changing availability of prey. We provide additional natural history observations and conclude by suggesting conservation actions that would help better understand and safeguard S. abnormis populations

Impacts of herbivory by ecological replacements on an island ecosystem

The use of ecological replacements (analogue species to replace extinct taxa) to restore ecosystem functioning is a promising conservation tool. However, this approach is controversial, in part due to a paucity of data on interactions between analogue species and established taxa in the ecosystem. We conducted ecological surveys, comprehensively DNA barcoded an ecosystem's flora and inferred the diet of the introduced Aldabra giant tortoise, acting as an ecological replacement, to understand how it might have modified island plant communities on a Mauritian islet. Through further dietary analyses, we investigated consequential effects on the threatened endemic Telfair's skink. Dietary overlap between tortoises and skinks was greater than expected by chance. However, there was a negative correlation between tortoise and skink preferences in herbivory and minimal overlap in the plants most frequently consumed by the reptiles. Changes in the plant community associated with 7 years of tortoise grazing were characterised by a decrease in the percentage cover of native herbs and creepers, and an increase in the cover of an invasive herb when compared to areas without tortoises. However, tortoise dietary preferences themselves did not directly drive changes in the plant community. Tortoises successfully dispersed the seeds of an endemic palm, which in time may increase the extent of unique palm-rich habitat. We found no evidence that tortoises have increased the extent of plant species hypothesised to be part of a lost Mauritian tortoise grazed community. Synthesis and applications. Due to a negative correlation in tortoise and skink dietary preferences and minimal overlap in the most frequently consumed taxa, the presence of tortoises is unlikely to have detrimental impacts on Telfair's skinks. Tortoise presence is likely to be beneficial to skinks in the long term by increasing the extent of palm-rich habitat. Although tortoises are likely to play a role in controlling invasive plants, they are not a panacea for this challenge. After 7 years, tortoises have not resurrected a lost tortoise grazed community that we hypothesise might have existed in limited areas on the islet, indicating that further interventions may be required to restore this plant community

Possible control of introduced giant African land snails (Achatina spp.) by the reintroduced endemic skink Leiolopisma telfairii, Ile aux Aigrettes, Mauritius

The giant African land snail (Achatinafulica) is one of the world’s worst invasive species, out‐competing endemic snails, consuming native vegetation and potentially altering nutrient cycles. Attempts to eradicate the snail from islands have only been successful with incipient populations. We present correlative evidence that native island predators may act as an effective control agent for the snail. In 2000 a population of between 37,300 and 45,100 African land snails was estimated on the 26ha nature reserve island of Ile aux Aigrette, Mauritius. Between 2006 and 2007, 260 endemic Telfair’s skink Leiolopisma telfairii were reintroduced to the reserve. Snail population surveys in 2008 and 2009 showed that the introduced snail population had declined to 5,569 (± 3,630) and 6,871 (±5,379), respectively. Previous studies showed that the introduced snails were selective over other invertebrate prey items. We suggest that predation by the endemic skink has been an important causal factor behind the snail population decline

Possible control of introduced giant African land snails (Achatina spp.) by the reintroduced endemic skink Leiolopisma telfairii, Ile aux Aigrettes, Mauritius

The giant African land snail (Achatina fulica) is one of the world’s worst invasive species, out‐competing endemic snails, consuming native vegetation and potentially altering nutrient cycles. Attempts to eradicate the snail from islands have only been successful with incipient populations. We present correlative evidence that native island predators may act as an effective control agent for the snail. In 2000 a population of between 37,300 and 45,100 African land snails was estimated on the 26ha nature reserve island of Ile aux Aigrette, Mauritius. Between 2006 and 2007, 260 endemic Telfair’s skink Leiolopisma telfairii were reintroduced to the reserve. Snail population surveys in 2008 and 2009 showed that the introduced snail population had declined to 5,569 (± 3,630) and 6,871 (±5,379), respectively. Previous studies showed that the introduced snails were selective over other invertebrate prey items. We suggest that predation by the endemic skink has been an important causal factor behind the snail population decline

New universal ITS2 primers for high-resolution herbivory analyses using DNA metabarcoding in both tropical and temperate zones

DNA metabarcoding is a rapidly growing technique for obtaining detailed dietary information. Current metabarcoding methods for herbivory, using a single locus, can lack taxonomic resolution for some applications. We present novel primers for the second internal transcribed spacer of nuclear ribosomal DNA (ITS2) designed for dietary studies in Mauritius and the UK, which have the potential to give unrivalled taxonomic coverage and resolution from a short-amplicon barcode. In silico testing used three databases of plant ITS2 sequences from UK and Mauritian floras (native and introduced) totalling 6561 sequences from 1790 species across 174 families. Our primers were well-matched in silico to 88% of species, providing taxonomic resolution of 86.1%, 99.4% and 99.9% at the species, genus and family levels, respectively. In vitro, the primers amplified 99% of Mauritian (n=169) and 100% of UK (n=33) species, and co-amplified multiple plant species from degraded faecal DNA from reptiles and birds in two case studies. For the ITS2 region, we advocate taxonomic assignment based on best sequence match instead of a clustering approach. With short amplicons of 187–387 bp, these primers are suitable for metabarcoding plant DNA from faecal samples, across a broad geographic range, whilst delivering unparalleled taxonomic resolution.</p

New universal ITS2 primers for high-resolution herbivory analyses using DNA metabarcoding in both tropical and temperate zones

DNA metabarcoding is a rapidly growing technique for obtaining detailed dietary information. Current metabarcoding methods for herbivory, using a single locus, can lack taxonomic resolution for some applications. We present novel primers for the second internal transcribed spacer of nuclear ribosomal DNA (ITS2) designed for dietary studies in Mauritius and the UK, which have the potential to give unrivalled taxonomic coverage and resolution from a short-amplicon barcode. In silico testing used three databases of plant ITS2 sequences from UK and Mauritian floras (native and introduced) totalling 6561 sequences from 1790 species across 174 families. Our primers were well-matched in silico to 88% of species, providing taxonomic resolution of 86.1%, 99.4% and 99.9% at the species, genus and family levels, respectively. In vitro, the primers amplified 99% of Mauritian (n=169) and 100% of UK (n=33) species, and co-amplified multiple plant species from degraded faecal DNA from reptiles and birds in two case studies. For the ITS2 region, we advocate taxonomic assignment based on best sequence match instead of a clustering approach. With short amplicons of 187–387 bp, these primers are suitable for metabarcoding plant DNA from faecal samples, across a broad geographic range, whilst delivering unparalleled taxonomic resolution.</p

Threatened endemic arthropods and vertebrates partition their diets 1 with non-native ants in an isolated island ecosystem

The success of non-native species depends on their ability to find food, which may ultimately lead to competition with native species and contribute to biodiversity loss in invaded ecosystems. Understanding which food resources are consumed is therefore crucial for evaluating how non-native species mechanistically fit into native biological communities. Non-native species may be predators or competitors of native species or may be consumed by native species as a novel source of nutrition, for example, and this can occur between both closely and distantly related species. Studies examining competitive interactions between non-native species and distantly related native taxa are relatively rare, largely because it is difficult to compare their diets using traditional methods. However, dietary DNA metabarcoding overcomes these limitations by enabling the construction of highly detailed food webs. Here, we use dietary DNA metabarcoding between two generalist native consumers – a reptile (Telfair’s skink) and a Scolopendra centipede (Serpent Island centipede) – and the hyperabundant non-native ant community to test which consumer groups prey upon one another and partition food resources. To determine how non-native ants fit into a native community, we calculated dietary composition, niche overlap, and dietary diversity of ants, centipedes, and skinks on Round Island, a small 2.19 km² oceanic island located 22.5 km north-east of Mauritius. We observed distinct partitioning of food resources among the three consumer groups—skinks, centipedes, and ants—and found that the level of predation between these groups varied. Skinks and centipedes frequently consumed non-native ants, which may represent an important nutritional resource for both native consumers. Dietary differences persisted through seasons despite large shifts in the availability of food and concomitant diet composition for all three consumers. We conclude that non-native ants fit into the biological community of Round Island as both prey for native consumers and extreme omnivorous generalists, but not necessarily at the expense of the native consumers because it is unlikely the consumers are competing for food resources. Our results suggest that abundant non-native generalists, which are highly invasive in much of their introduced range, can infiltrate native food webs without exerting strong competitive forces on other common native generalist species