A cytogenetic and molecular characterization of Meloidogyne-induced giant cell nuclei

Abstract

DNA content of Meloidogyne-induced giant cell nuclei in several host species was measured microspectrophotometrically. Nuclei from mature giant cells in pea and tomato averaged over a 14-fold increase in DNA compared to nuclei from uninfected root tip cells. Nuclei from broad bean and lettuce showed a mean increase in DNA content of 8.3- and 6.5-fold, respectively, when compared to uninfected root tip nuclei. DNA content of giant cells was highly variable (coefficients of variation > 41%) while DNA content of root tip cells showed little variation (coefficients of variation < 8%). DNA content in giant cell nuclei from pea increased linearly from the time of inoculation until development of mature females three weeks after inoculation. In giant cells from lettuce, DNA content increased 6.5-fold during the first week after inoculation with no further increase. Cytogenetic studies were conducted on pea giant cell nuclei. Chromosome counts of metaphase-arrested nuclei revealed a majority of aneuploid nuclei with counts ranging from 17 (2n=14) to over 100. No pattern to the counts emerged. Squashes of giant cell nuclei in late anaphase and telophase revealed lagging chromosomes and chromatin bridges, respectively. Also observed were telophase nuclei in the process of fusion. To determine whether selective sequence amplification has occurred in giant cell nuclei, slot blots were conducted on DNA from uninfected roots and DNA enriched in the giant cell DNA. The probes used in the blots were (1) a 2.1 kilobase (kb) fragment of the soybean ribulose bis phosphate decarboxylase small subunit gene, (2) a 3 kb fragment of a soybean actin gene, (3) a 415 bp histone 3 fragment from Arabidopsis thaliana, and (4) a 4 kb ribosomal gene fragment from soybean. Densitometry on autoradiographs failed to show any selective sequence amplification of the tested probes. The results of this study showed Meloidoavne-induced nuclei to have highly elevated DNA levels when compared to root tip nuclei and exhibit aneuploid chromosome numbers. Additionally, slot blots with four selected probes failed to show any selective sequence amplification.