The Ambrosia Symbiosis: From Evolutionary Ecology to Practical Management

Background During the last 15 years the internal transcribed spacer (ITS) of nuclear DNA has been used as a target for analyzing fungal diversity in environmental samples, and has recently been selected as the standard marker for fungal DNA barcoding. In this study we explored the potential amplification biases that various commonly utilized ITS primers might introduce during amplification of different parts of the ITS region in samples containing mixed templates ('environmental barcoding'). We performed in silico PCR analyses with commonly used primer combinations using various ITS datasets obtained from public databases as templates. Results Some of the ITS primers, such as ITS1-F, were hampered with a high proportion of mismatches relative to the target sequences, and most of them appeared to introduce taxonomic biases during PCR. Some primers, e.g. ITS1-F, ITS1 and ITS5, were biased towards amplification of basidiomycetes, whereas others, e.g. ITS2, ITS3 and ITS4, were biased towards ascomycetes. The assumed basidiomycete-specific primer ITS4-B only amplified a minor proportion of basidiomycete ITS sequences, even under relaxed PCR conditions. Due to systematic length differences in the ITS2 region as well as the entire ITS, we found that ascomycetes will more easily amplify than basidiomycetes using these regions as targets. This bias can be avoided by using primers amplifying ITS1 only, but this would imply preferential amplification of 'non-dikarya' fungi. Conclusions We conclude that ITS primers have to be selected carefully, especially when used for high-throughput sequencing of environmental samples. We suggest that different primer combinations or different parts of the ITS region should be analyzed in parallel, or that alternative ITS primers should be searched for.

Ectosymbioses among bark beetles (Curculionidae, Scolytinae) and fungi (primarily ophiostomatoid Ascomycetes) are widespread and diverse. Associations range from mutualistic to commensal, and from facultative to obligate. Some fungi are highly specific and associated only with a single beetle species, while others can be associated with many. In addition, most of these symbioses are multipartite, with the host beetle associated with two or more consistent partners. Mycangia, structures of the beetle integument that function in fungal transport, have evolved numerous times in the Scolytinae. The evolution of such complex, specialized structures indicates a high degree of mutual dependence among the beetles and their fungal partners. Unfortunately, the processes that shaped current day beetle-fungus symbioses remain poorly understood. Phylogeny, the degree and type of dependence on partners, mode of transmission of symbionts (vertical vs. horizontal), effects of the abiotic environment, and interactions among symbionts themselves or with other members of the biotic community, all play important roles in determining the composition, fidelity, and longevity of associations between beetles and their fungal associates. In this review, I provide an overview of these associations and discuss how evolution and ecological processes acted in concert to shape these fascinating, complex symbioses.