Fig. 2

Fungal amplification using TEF primers is more efficient and displays reduced species bias compared to ITS1. A. Quantitative PCR efficiency (left) was significantly higher with TEF compared to ITS1 for all three species (P < 0.0001, 2-way ANOVA). Species efficiencies were significantly different from one another for ITS1 (*; P < 0.05, ***; P < 0.001 by 2-way ANOVA Tukey’s multiple comparisons). Relative quantitation of (A) fumigatus, C. albicans and L. prolificans using the ITS1 (middle) and TEF (right) PCR assays shows under-representation of L. prolificans with ITS1. (B) Quantitative PCR of 50,000 haploid genome equivalents of twenty one fungal species shows species quantitation is more variable with ITS1 than TEF. Data is visualised as difference from mean Ct value. Ct values were significantly different between species for TEF and ITS1 (P < 0.001 and P < 0.0001, respectively, by ANOVA. Of the 210 Tukey’s multiple comparisons tests performed, 6 and 81 were significant for TEF and ITS1, respectively (P < 0.05, not indicated)