North Carolina State University Raleigh, North Carolina, United States
Abstract Text: Root-knot nematodes are an economically important plant pathogen that have had a severe impact on national and North Carolina agriculture. The guava root-knot nematode (GRKN), Meloidogyne enterolobii, can infect crops such as cotton, sweetpotato, and soybean causing severe galling on the roots, leading to yield loss. Species identification of GRKN in soil is crucial in combating yield loss and damage, but can be difficult and time consuming. Identification methods include observing morphological characteristics and molecular techniques (PCR), but first the GRKN must be separated from the soil (e.g. Whitehead tray extraction), which is laborious and requires a lengthy incubation period. In this new molecular identification procedure, the GRKN remains in the soil matrix and DNA is extracted directly using a series of buffers, hot bath incubation, Fe3O4 nanoparticles, purification steps, and identification using species-specific PCR. This increases the efficiency and precision since it bypasses the need to isolate individuals from soil and is used in tandem with PCR. Sensitivity testing for samples inoculated with 100, 500, 1,000, 5,000, and 10,000 second-stage juveniles reveal that the lower limit of sensitivity with species-specific primers is approx. 100 J2 per 40 g soil. Results were obtained within 1.5 days of receiving the soil samples, nearly half the time required for standard Whitehead tray extraction followed by plucking and individual PCRs. This illustrates the efficiency and effectiveness of the procedure in interpreting GRKN populations in infected fields.
