Table 2.

Comparison of the strength and weaknesses of available diet analysis techniques

		Molecular techniques		Traditional methods
	High-throughput sequencing (HTS)	Cloning (Zeale et al. 2011)	Fragments (Clare et al. 2011b)	Morphological—feces/stomach	Culled prey remains
Diet resolution	High: species level constrained by reference sequences database (can be mediated by MOTU)	High: species level constrained by reference sequences database (can be mediated by MOTU)	High: species level constrained by reference sequences database (can be mediated by MOTU)	Low: family order	High: species order but depends on taxa
Diet coverage	Better representation of the DNA extracted, but bias from primer binding biases, and relative DNA abundance of prey	Limited by the selected number of clones. Bias toward free floating DNA, DNA abundance, and primer bias	Selected fragments. Bias toward prey of low digestibility or fragment selection method	Greater bias toward hard-bodied prey of low digestibility	Bias toward large prey that requires culling
Diet quantification	Number of species/ MOTUs, no within samples quantification	Number of species/ MOTUs, no within samples quantification	Number of species/ MOTUs, no within samples quantification	Number of prey taxa and percent volume	Number of prey taxa and density
Rare dietary components	High potential to identify rare prey—high proportion of DNA variance sequenced	Selection of clones for sequencing reduces the chance of identifying rare prey	Potential of identification depends on number of fragments sequenced from each dropping (costs)	High potential to identify hard-bodied rare prey, but low potential to recover rare soft prey	Rare prey identified only if require culling
Accuracy	Does not require taxonomists to obtain accurate results, lower analyzer bias	Does not require taxonomists to obtain accurate results, lower analyzer bias	Does not require taxonomists to obtain accurate results, lower analyzer bias	Accuracy requires significant entomological training on the part of the identifier	Accuracy requires entomological training on the part of the identifier
Costs	Less limited by the amount of prey sequences or droppings per sequencing run. Lower cost per sequence but high initial costs and needs specialized facilities	Expensive cloning process, but less specialized facilities	Depends on number of fragments sequenced—each fragment requires separate sequencing. Less specialized facilities.	Very low—minimal consumables	Very low—minimal consumables
Applicability across feeding groups	High relevance across taxa, including herbivores	High relevance across taxa, including herbivores	Mainly relevant for predators and seed disperser	Mainly relevant for predators	Only relevant for predators that use feeding perches