Fusarium oxysporum f. sp. cubense (Foc) is a soil-borne fungus that causes Fusarium wilt, a destructive plant disease that has resulted in devastating economic losses to banana production worldwide. The fungus has a complex evolutionary history and taxonomic repute and consists of three pathogenic races and at least 24 vegetative compatibility groups (VCGs). Surveys conducted in Asia, Africa, the Sultanate of Oman and Mauritius encountered isolates of F. oxysporum pathogenic to banana that were not compatible to any of the known Foc VCGs. Genetic relatedness between the undescribed and known Foc VCGs were determined using a multi-gene phylogeny and diversity array technology (DArT) sequencing. The presence of putative effector genes, the secreted in xylem (SIX) genes, were also determined. Fourteen novel Foc VCGs and 17 single-member VCGs were identified. The multi-gene tree was congruent with the DArT-seq phylogeny and divided the novel VCGs into three clades. Clustering analysis of the DArT-seq data supported the separation of Foc isolates into eight distinct clusters, with the suite of SIX genes mostly conserved within these clusters. Results from this study indicates that Foc is more diverse than hitherto assumed.

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The polyphagous shot hole borer (PSHB) Euwallacea fornicatus is an ambrosia beetle that poses a serious threat to hundreds of tree species in invaded countries. In its invaded range, it relies on a mutualistic fungus, Fusarium euwallaceae, for colony establishment and nutrition. Together, the beetle–fungus complex can cause Fusarium dieback disease that can lead to the death of highly susceptible hosts. The key to mitigation of the spread and control of PSHB is its rapid detection. Current identification relies on DNA extraction and sequencing of barcoding genes of the beetle and/or the fungus, which is expensive and time‐consuming. Often the beetle is not present in a sample, and isolation and purification of the fungus is needed before employing molecular procedures. Because PSHB has a strong association with its fungal symbiont, at least in its invaded range, and the fungus can infect trees without successful PSHB colony establishment, a molecular tool that can rapidly detect F. euwallaceae from infected wood would aid monitoring efforts. Here we developed a PCR amplification protocol that targets a species‐specific gene present in F. euwallaceae from within infected wood of various hosts without prior fungal isolation, DNA purification or sequencing. The technique can detect F. euwallaceae within minimal woody material (c.15 mg) and even from degraded samples of many hosts. Additionally, identification of F. euwallaceae using lysate prepared from mycelia provided a reliable secondary measure. This method drastically reduces the time and costs associated with the identification of F. euwallaceae from field samples.