Fig 1.
Flowchart of study design.
Table 1.
Sequences of primers and probes used during the SOP harmonization procedure.
Table 2.
Participants in the first round BU-LABNET EQA program.
Table 3.
Qualitative results in the first round BU-LABNET EQA program.
Table 4.
Laboratories from 9 African countries and activities in 2020 and 2021.
Fig 2.
Comparison results of DNA extraction methods.
(A) from four swabs, and (B) from four FNA. No significant difference was observed between the “in-house” method and the commercial kit (Dunn’s multiple comparisons test).
Fig 3.
Validation of the DNA extraction method using commercial kit (GenoLyse).
Nineteen positive samples were extracted with the “in house” method followed by purification on Qiagen column versus the commercial kit. Validation of the commercial kit for DNA extraction was evaluated with Spearman’s correlation test. Each dot corresponds to a sample. The x and y axes indicate the logarithm of M. ulcerans DNA concentration. Spearman’s correlation coefficient = 0.9702. ****, Correlation was statistically significant with p<0.0001. No significant difference was observed between the “in-house” method and the commercial kit (Wilcoxon test).
Fig 4.
Comparison of qPCR mixes and primers/probe sets for sensitivity and efficiency.
Serial 10-fold dilution from 6.4x106U/mL to 6.4x100U/mL of M. ulcerans DNA. The standard curve was generated by a linear regression of the threshold cycle (Ct) against the logarithm of M. ulcerans DNA concentration. Each 10-fold dilution was performed in duplicate for n = 3 experiments, one standard deviation (SD) is shown. (A) Primers/probe set 1: Brillant II (y = −3.16x + 41.37); Hot FirePol (y = −3.307x + 42.44). (B) Primers/probe set 2: Brillant II (y = −3.217x + 42.31); Hot FirePol (y = −3.147x + 41.84).
Fig 5.
Comparison of internal positive control: Standard curve for IS2404.
Serial 10-fold dilution from 6,4x106U/mL to 6.4x100U/mL of M. ulcerans DNA (in black). Serial 10-fold dilution from 1x107U/mL to 1x100U/mL of plasmid DNA (in blue). The standard curve was generated by a linear regression of the threshold cycle (Ct) against the logarithm of DNA concentration. Each 10-fold dilution was performed in duplicate, one standard deviation (SD) is shown. M. ulcerans DNA standard curve (y = −3.271x + 43.96); Plasmid standard curve (y = −3.074x + 43.56).