Screen of Pinus massoniana for Resistance to Pinewood Nematode: In Vitro Propagation and Evaluation of Regenerated Microshoots

Abstract

To accelerate breeding of Pinus massoniana Lamb. resistance to pine wilt disease (PWD), caused by the pinewood nematode (PWN), Bursaphelenchus xylophilus, a protocol was established for the in vitro propagation of P. massoniana and the evaluation of resistance of regenerated microshoots from different clones to PWN. Axillary bud induction was achieved by culturing cotyledonary node explants from 3-week-old seedlings in Gupta and Durzan (DCR) medium that was supplemented with 4 mg L−1 6-benzyladenine (BA) and 0.2 mg L−1 α-naphthaleneacetic acid (NAA). Explants with induced buds were transferred to DCR medium without a plant growth regulator to facilitate elongation. Stem segments from elongated shoots were used as propagules for further shoot multiplication. Six-month-old regenerated shoots that met the requirements for a nematode resistance test were inoculated with aseptic PWN (500 PWNs/shoot). The wilting rate varied between clones from 20% to 100%, 18 days after inoculation. Except for Clone 227, which showed the highest resistance with a wilting rate of 0%, other clones showed wilting to various degrees 30 d after inoculation. The number of nematodes that were recovered from Clone 227 was significantly lower than from other clones. This study promotes the resistance breeding of P. massoniana to pine wilt disease and provides an effective method to study the host/pathogen interaction between PWN and P. massoniana.