Abstract
In the never ending struggle against plant pathogenic bacteria, a major goal is the early identification and classification of infecting microorganisms. Xylella fastidiosa, a Gram-negative bacterium belonging to the family Xanthmonadaceae, is no exception as this pathogen showed a broad range of vectors and host plants, many of which may carry the pathogen for a long time without showing any symptom. Till the last years, most of the diseases caused by X. fastidiosa have been reported from North and South America, but recently a widespread infection of olive quick decline syndrome caused by this fastidious pathogen appeared in Apulia (south-eastern Italy), and several cases of X. fastidiosa infection have been reported in other European Countries. At least five different subspecies of X. fastidiosa have been reported and classified: fastidiosa, multiplex, pauca, sandyi, and tashke. A sixth subspecies (morus) has been recently proposed. Therefore, it is vital to develop fast and reliable methods that allow the pathogen detection during the very early stages of infection, in order to prevent further spreading of this dangerous bacterium. To this purpose, the classical immunological methods such as ELISA and immunofluorescence are not always sensitive enough. However, PCR-based methods exploiting specific primers for the amplification of target regions of genomic DNA have been developed and are becoming a powerful tool for the detection and identification of many species of bacteria. The aim of this review is to illustrate the application of the most commonly used PCR approaches to X. fastidiosa study, ranging from classical PCR, to several PCR-based detection methods: random amplified polymorphic DNA (RAPD), quantitative real-time PCR (qRT-PCR), nested-PCR (N-PCR), immunocapture PCR (IC-PCR), short sequence repeats (SSRs, also called VNTR), single nucleotide polymorphisms (SNPs) and multilocus sequence typing (MLST). Amplification and sequence analysis of specific targets is also mentioned. The fast progresses achieved during the last years in the DNA-based classification of this pathogen are described and discussed and specific primers designed for the different methods are listed, in order to provide a concise and useful tool to all the researchers working in the field.
Highlights
Xylella fastidiosa is a Gram-negative, slow growing and strictly aerobic bacterium in the family Xanthmonadaceae
Such approach was used for characterizing X. fastidiosa isolated from olive trees affected by olive quick decline syndrome (OQDS) in Italy and the results showed that olive strain is distinct from the four previously defined species fastidiosa, multiplex, sandyi and pauca, showing the highest level of similarity with the latter
X. fastidiosa has recently emerged as a potential threat to European agriculture after its outbreak in southern Italy were it was found associated to OQDS (Saponari et al, 2013; Cariddi et al, 2014; Elbeaino et al, 2014; Martelli et al, 2016)
Summary
Xylella fastidiosa is a Gram-negative, slow growing and strictly aerobic bacterium in the family Xanthmonadaceae. One of the major problems caused by X. fastidiosa infections is the decreased fruit quality and yield in commercially important crops such as coffee and olive (Rocha et al, 2010; Della Coletta et al, 2016) Plants affected by scorch disease usually show weak and stunt growth, may be more susceptible to environmental stresses, such as water or heat stress, but do not always die Instead, they can be removed to reduce the risk of new infections or because the weakened plant has become dangerous (e.g., ornamental trees). A multiprimer combination with three different targets was developed in order to differentiate strains of X. fastidiosa
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