Fusarium wilt‐resistant lines of Brazil banana (Musa spp., AAA) obtained by EMS‐induced mutation in a micro‐cross‐section cultural system

This study combined the micro‐cross‐section cultural system with in vitro mutagenesis induced by ethyl methanesulphonate (EMS) to screen for fusarium wilt‐resistant lines of Brazil banana (Musa spp., AAA). The results indicated that the optimum EMS concentration and duration for the treatment of micro‐cross‐sections cut from the pseudostem of tissue‐cultured plantlet were 300 mm and 60 min, respectively. Under the optimal treatment, an average of 2·2 regenerated shoots were produced from each explant. One hundred regenerated plantlets were used for screening for fusarium wilt‐resistant lines by the early screening technique. The initial disease symptom – yellowing in lower leaves of susceptible plantlets – was observed 2 weeks after inoculation. After 2 months, only six plants survived – the putative fusarium wilt‐resistant lines. The fusarium wilt pathogen Fusarium oxysporum f. sp. cubense race 4, was identified in the preliminary test field by a SCAR marker technique. Of the six putative resistant lines, five survived the preliminary field test. The regenerated plantlets from these five fusarium wilt‐resistant lines were subjected to early screening again, where they showed markedly reduced disease incidences compared with regenerated plantlets of Brazil banana (control). It was concluded that EMS‐induced mutation of banana through the micro‐cross‐section cultural system is potentially useful for banana improvement.     

Development of a high‐throughput real‐time PCR assay for the detection of the R81T mutation in the nicotinic acetylcholine receptor of neonicotinoid‐resistant Myzus persicae

BACKGROUND: Myzus persicae is a globally important aphid pest that is mainly controlled through the application of chemical insecticides. Recently, a clone of M. persicae exhibiting control‐compromising levels of resistance to neonicotinoid insecticides was described. The resistance of this clone was associated with reduced affinity of imidacloprid for the target site (the nicotinic acetylcholine receptor) as a result of mutation of a key amino acid residue (R81T) in the loop D region of a nAChR β1 subunit. The potent levels of resistance conferred by this mechanism are cause for considerable concern, and the frequency and distribution of the mutation in worldwide populations of M. persicae require careful monitoring. In this study, a high‐throughput assay has been developed that allows detection of the mutation in individual aphids. RESULTS: A real‐time TaqMan assay to detect the R81T substitution was developed that proved to be sensitive and specific in tests of analytical sensitivity and in a blind genotyping trial of DNA extracted from individual aphids comprising the three possible genotypes. The assay was then used to examine the frequency of the R81T mutation in aphids collected and stored in ethanol from peach orchards in southern France. The R81T frequency varied from 33 to 100% in seven populations from the department of Gard, France. CONCLUSIONS: This study describes a rapid and sensitive assay that very effectively detects the R81T mutation in individual aphids. The results also have practical significance for the control of M. persicae in southern France and provide contemporary data to inform current resistance management strategies. Copyright © 2012 Society of Chemical Industry