Diagnosis of Phytophthora in Soil Samples by Polymerase Chain Reaction
Journal Title: Current Investigations in Agriculture and Current Research - Year 2018, Vol 1, Issue 1
Abstract
Phytophthora infestans is the most notorious causing Late blight of potato and tomato, globally. The word Phytophthora is derived from the Greek word: Phyto = plant, phthora = destroyer. Potato is a native of the North Andes (South America) and the late blight of potato was initially an endemic disease but in the mid1800s, late blight caused wide spread crop failures throughout the Northern Europe including Ireland where it was responsible for the Irish famine [1]. The genus Phytophthora represents a large group of plant pathogenic fungi responsible for crop losses in temperate and tropical climate [2]. Many species of Phytophthora are soil borne pathogens and spread through the movement of infested soil, or by water flow through infested soil [3]. A key element in the management of such diseases is the ability to detect the pathogen in soil and water. However, DNA extracted from soil contains substances such as humic acids, lignins, carbohydrates, resins, and so on which are very inhibitory to PCR amplification [4,5]. The amounts of inhibitory substances will vary widely with soil type, vegetation type, and composition of the soil micro flora. As the micro flora varies even over small distances (1 m scale) [6], the efficiency of PCR amplification is likely to vary widely even over small distances. It is therefore critical that an internal standard is used for PCR analysis of soil samples [7]. Recorded positive detection of P. infestans for up to twelve months from soil in which infested leaf tissue had been buried and for up to 24 months from soil containing leaf tissue infected with both mating types. The difference is ascribed to the formation of sexual oospores in the doubly infected material. However [8] showed that infective propagules of P. cinnamomido not persist for long periods in soil. After incubation of infested soil for 12 weeks at 25°C the pathogen could not be detected by baiting, and although it could be detected by PCR the detection efficiency was significantly reduced. When the soil was incubated at 30°C for 12 weeks the pathogen could notbe detected by either baiting or PCR. For detection in soil, where there is no obvious lesiontissue to sample from, soil baiting has the advantage that it enables large amounts of soil (0.5-1 kg) to be tested whereas DNA extraction protocols are all based on extraction of small (1-10 g) samples. Consequently, with sampling for DNA extraction and analysis by PCR we may simply miss the pathogen. Baiting also has the advantage that it only detects viable pathogen. Recently [9] developed a new modified baiting method for detection P.infestans from potato agricultural fields. Despite the advantages, baiting is too slow and low throughput to be useful and can be subject to a high degree of false negatives also. The efficiency of detection of Phytophthora in soil by the baiting technique can be improved by sieving out the soil and rewetting it (double baiting demonstrating that, as with tissue sections, the pathogen, although present, will not always grow out of the sample [10,11] found that double baiting increased the recovery of positive samples [12].
Authors and Affiliations
Touseef Hussain
Keywords
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- EP ID EP567409
- DOI 10.32474/CIACR.2018.01.000103
- Views 132
- Downloads 0
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