Molecular detection of invertebrate prey in vertebrate diets: trophic ecology of Caribbean island lizards

Understanding community assembly and population dynamics frequently requires detailed knowledge of food web structure. For many consumers, obtaining precise information about diet composition has traditionally required sacrificing animals or other highly invasive procedures, generating tension between maintaining intact study populations and knowing what they eat. We developed 16S mitochondrial DNA sequencing methods to identify arthropods in the diets of generalist vertebrate predators without requiring a blocking primer. We demonstrate the utility of these methods for a common Caribbean lizard that has been intensively studied in the context of small island food webs: Anolis sagrei (a semi-arboreal ‘trunk-ground’ anole ecomorph). Novel PCR primers were identified in silico and tested in vitro. Illumina sequencing successfully characterized the arthropod component of 168 faecal DNA samples collected during three field trips spanning 12 months, revealing 217 molecular operational taxonomic units (mOTUs) from at least nine arthropod orders (including Araneae, Blattodea, Coleoptera, Hemiptera, Hymenoptera, Isoptera, Lepidoptera and Orthoptera). Three mOTUs (one beetle, one cockroach and one ant) were particularly frequent, occurring in ≥50% of samples, but the majority of mOTUs were infrequent (180, or 83%, occurred in ≤5% of samples). Species accumulation curves showed that dietary richness and composition were similar between size-dimorphic sexes; however, female lizards had greater per-sample dietary richness than males. Overall diet composition (but not richness) was significantly different across seasons, and we found more pronounced interindividual variation in December than in May. These methods will be generally useful in characterizing the diets of diverse insectivorous vertebrates.

解析群落组装与种群动态，通常需要对食物网结构具备详尽的认知。传统上，要获取多数消费者的食物组成精确信息，需通过宰杀动物或其他高侵入性操作，这使得维持完整研究种群与掌握其摄食内容之间产生了矛盾。本研究开发了16S线粒体DNA（16S mitochondrial DNA）测序方法，可在无需封闭引物的前提下，鉴定广谱脊椎动物捕食者食谱中的节肢动物。我们以小型岛屿食物网研究中被广泛关注的常见加勒比海蜥蜴——安乐蜥（Anolis sagrei，半树栖“树陆型”安乐蜥生态型）为对象，验证了该方法的实用性。研究通过计算机模拟筛选（in silico）获得了新型聚合酶链式反应（PCR）引物，并通过体外实验（in vitro）验证了其有效性。本次研究通过Illumina测序技术，对为期12个月的3次野外考察采集的168份粪便DNA样本中的节肢动物组分进行了成功解析，共鉴定出至少9个节肢动物目（包括蜘蛛目Araneae、蜚蠊目Blattodea、鞘翅目Coleoptera、半翅目Hemiptera、膜翅目Hymenoptera、等翅目Isoptera、鳞翅目Lepidoptera以及直翅目Orthoptera）下的217个分子操作分类单元（molecular operational taxonomic units, mOTUs）。其中3个mOTUs（分别对应1种甲虫、1种蟑螂和1种蚂蚁）出现频率极高，在≥50%的样本中均可检测到；但绝大多数mOTUs（共180个，占比83%）仅在≤5%的样本中出现。物种积累曲线分析显示，体型二态的雌雄个体间，其食谱丰富度与组成均较为相似；但雌性蜥蜴单样本的食谱丰富度显著高于雄性。整体食谱组成（而非丰富度）在不同季节间存在显著差异，且12月的个体间摄食差异比5月更为显著。该方法可广泛应用于解析多样食虫脊椎动物的食谱组成。