INVITED REVIEW: Molecular analysis of predation: a review of best practice for DNA‐based approaches

Abstract

Molecular analysis of predation, through polymerase chain reaction amplification of prey remains within the faeces or digestive systems of predators, is a rapidly growing field, impeded by a lack of readily accessible advice on best practice. Here, we review the techniques used to date and provide guidelines accessible to those new to this field or from a different molecular biology background. Optimization begins with field collection, sample preservation, predator dissection and DNA extraction techniques, all designed to ensure good quality, uncontaminated DNA from semidigested samples. The advantages of nuclear vs. mitochondrial DNA as primer targets are reviewed, along with choice of genes and advice on primer design to maximize specificity and detection periods following ingestion of the prey by the predators. Primer and assay optimization are discussed, including crossamplification tests and calibratory feeding experiments. Once primers have been made, the screening of field samples must guard against (through appropriate controls) cross contamination. Multiplex polymerase chain reactions provide a means of screening for many different species simultaneously. We discuss visualization of amplicons on gels, with and without incorporation of fluorescent primers. In more specialized areas, we examine the utility of temperature and denaturing gradient gel electrophoresis to examine responses of predators to prey diversity, and review the potential of quantitative polymerase chain reaction systems to quantify predation. Alternative routes by which prey DNA might get into the guts of a predator (scavenging, secondary predation) are highlighted. We look ahead to new technologies, including microarrays and pyrosequencing, which might one day be applied to this field.

Detecting arthropod intraguild predation in the field

“…1. The best practice for most parts of this procedure is reviewed by King et al (2008). Here, we highlight key aspects.…”

“…Because ladybirds possess taxonomically-specific defensive alkaloids (Daloze et al 1995), GC-MS of prey alkaloids in predator bodies is particularly appropriate for studying IGP of other ladybird species by H. axyridis (Hautier et al 2008;Sloggett et al 2009;Sloggett et al 2011) and to date are used in the only two published field studies on the ecological impact of H. axyridis (Hautier et al 2008;Hautier et al 2011). In addition to the above technique, the last decade has seen the development of a PCR DNA based approach to identifying the gut contents of predatory invertebrates (King et al 2008).…”

“…They showed that the largest fragment had a statistically shorter half life than the two smaller fragments. Studies from the mid to late 2000s have consistently targeted prey amplicons less than 250 bps (see King et al 2008). The main steps of a general procedure to design and test primers for use in a DNA detection system for IGP are summarized in Fig.…”

“…1 Basic procedure to design and test primers for use as molecular markers for intraguild predation studies under field conditions. Adapted from King et al (2008). For further details, see main text or King et al (2008) Field studies…”

“…The predator is starved beforehand, generally for 24 h (King et al 2008). The prey species of interest is the sole source of food for the predator, yet in the field it may represent only a small percentage of the predator's daily diet.…”