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Genomics-informed LAMP-PCR design for detection of Xanthomonas arboricola pv. pruni and Erwinia amylovora. Andreas Bühlmann JF Pothier, THM Smits, JE Frey, B Duffy (ACW, Switzerland). 24-Month Meeting Wien (A) – 12-13.6.2012. www.qdetect.org. XAP LAMP assay.
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Genomics-informed LAMP-PCR design for detection of Xanthomonas arboricola pv. pruni and Erwinia amylovora Andreas Bühlmann JF Pothier, THM Smits, JE Frey, B Duffy (ACW, Switzerland) 24-Month Meeting Wien (A) – 12-13.6.2012 www.qdetect.org
XAP LAMP assay Xanthomonas arboricola pv. pruni Bacterial spot of stone fruit (Prunus) • Assay designed • Validated • Ring tested • Paper submitted/ accepted • EPPO validation? Andreas Bühlmann – 12-13.6.2012 Q-Detect 24-Month Meeting, Wien, A
COST873 STSM in MelbourneAustralian isolates of Xap (DPI Melbourne)Talks about LAMP stuff with DPI and AQIS – they seem to be enthusiastic about the method, interested in: Viruses, Bacteria, Insects
XAP Ring test ACW Wädenswil, CITA Zaragoza, IVIA Valencia, APR Lisse Inter- Laboratory concordance Andreas Bühlmann – 12-13.6.2012 Q-Detect 24-Month Meeting, Wien, A
XAP Ring test ACW Wädenswil, CITA Zaragoza, IVIA Valencia, APR Lisse Inter- Assay concordance Cohen J. 1960. A coefficient of agreement for nominal scales. Euc. Psychol. Meas 20 (1): 37-46. Landis J.R., Koch G.G. 1997. The measurement of observer agreements for categorical data. Biomedics 33: 159-174. Andreas Bühlmann – 12-13.6.2012 Q-Detect 24-Month Meeting, Wien, A
XAP Ring test ACW Wädenswil, CITA Zaragoza, IVIA Valencia, APR Lisse Sensitivity and Specificity SE: Standard error; lo IC 95%: lower limit of the 95% confidence interval; up IC 95%: upper limit of the 95% confidence interval. Andreas Bühlmann – 12-13.6.2012 Q-Detect 24-Month Meeting, Wien, A
XAP Ring test ACW Wädenswil, CITA Zaragoza, IVIA Valencia, APR Lisse Talk or poster on next SEF congress (in spanish) Andreas Bühlmann – 12-13.6.2012 Q-Detect 24-Month Meeting, Wien, A
Ea LAMP assay Erwinia amylovora Fire blight of pome fruit (Spiraeoideae) • Assay designed • Validated • Field tested (only CH) • Paper about to submit • EPPO validation • Further sequencing efforts – 96 bacterial strains to further improve knowledge about presence/ absence of diagnostic targets Andreas Bühlmann – 12-13.6.2012 Q-Detect 24-Month Meeting, Wien, A
Planned/current assays for plant pathogenic bacteria • Comparative genomics, primer design, validation with partners on… • Pantoea ananatis • Pseudomonas syringae pv. actinida • Pseudomonas syringae pv. aesculi • Xanthomonas oryzae pv. oryzae • Xanthomonas oryzae pv. oryzicola • Ralstonia solanacearum (necessary?) • Suggestions? Xanthomonas fragariae? (Neil and colleagues) Andreas Bühlmann – 12-13.6.2012 Q-Detect 24-Month Meeting, Wien, A
Import diagnostics • Swiss quarantine inspection service is planning to buy two Genies… • Would need: • Thrips palmi • Bactrocera spp. • Bemisisa tabaci • Liriomyza trifolii/sativae (>80% of samples) • Multiplexable by melt curve: • Single tube assays make • inspectors lifes easier • Works only if no mixed samples • i.e. single insects Xa 88.2°C Ea 86.0°C Bt 83.3°C Tp 81.8°C Andreas Bühlmann – 12-13.6.2012 Q-Detect 24-Month Meeting, Wien, A
Streptomycin resistance monitoring with LAMP? vs. TaqMan • Point mutations on codon 43 of Ea rpsL leads to streptomycin resistance: • AAA Lysine wild type susceptible • AGA Arginine strep resistant • ACA Threonine strep resistant • AAY Asparigine strep resistant Andreas Bühlmann – 12-13.6.2012 Q-Detect 24-Month Meeting, Wien, A
