Cousseau-Suhard, Pascaline

Abstract‘Candidatus Liberibacter solanacearum’ (Lso) has emerged as a serious threat on solanaceous and apiaceous crops worldwide. Five Lso haplotypes (LsoA, LsoB, LsoC, LsoD and LsoE) have been identified so far. To decipher genetic relationships between Lso strains, a MLSA study of seven housekeeping genes (acnA, atpD, ftsZ, glnA, glyA, gnd and groEL) was performed on a representative bacterial collection of 49 Lso strains. In all, 5415 bp spanning the seven loci were obtained from each of the 49 strains of our bacterial collection. Analysis of sequence data was consistent with a clonal population structure with no evidence of recombination. Phylogenies reconstructed from individual genes, and with concatenated data, were globally congruent with each other. In addition to the five highly supported and distinct genetic clusters, which correspond to the five established haplotypes, our phylogenetic data revealed the presence of a sixth haplotype, designated ‘LsoG’. This new haplotype is currently represented by two strains from France which had distinct sequences in four out of the seven tested housekeeping genes. Altogether, the data presented here provide new information regarding the genetic structure of Lso and the evolutionary history of the haplotypes defined within this bacterial species.

‘Candidatus Liberibacter solanacearum’ is a bacterium associated with several vegetative disorders on solanaceous and apiaceous crops. Following the recent detection of the bacterium in carrots in Europe, and particularly carrot plants used for seed production in France, two independent laboratories conducted experiments on the transmission of this pathogen by seed and had discordant results: one study showed no bacterial transmission to plants, and the other showed transmission to carrot seedlings starting from the fourth month of culture. To test the hypothesis that growing conditions affect seed transmission efficiencies, trials were renewed in 2015 on four lots of 500 carrot seeds naturally contaminated with ‘Ca. L. solanacearum’ and two lots of 100 healthy seeds. The plants were grown for 6 months in an insect-proof NS2 greenhouse. Sets of 108 plants from the contaminated lots and 24 plants from the healthy lots were individually analyzed each month using real-time PCR to detect the bacterium. The detection tests on seeds and plants from healthy lots were always negative. During the 6 months of the trial, no plants from the contaminated seed lots tested positive for the bacterium or showed any infection symptoms. These results indicate that transmission of ‘Ca. L. solanacearum’ by carrot seed is rare and difficult to reproduce.

‘Candidatus Liberibacter solanacearum’ (Lso) is an emerging phytopathogenic bacterium that causes significant crop losses worldwide. This bacterium has been identified in association with diseases of several solanaceous crops in the United States and New Zealand, and with carrot and celery crops in several European countries. Five Lso haplotypes (LsoA, LsoB, LsoC, LsoD, and LsoE) have now been described worldwide. In France, symptoms of Lso were observed on plants of the Apiaceae family in several regions. One hundred and ninety-two samples of apiaceous plants were collected from 2012 to 2016 in different geographical regions and were tested for the occurrence of Lso by real-time PCR assay. In addition to carrot and celery, Lso was detected in four other apiaceous crops: chervil, fennel, parsley, and parsnip. These new findings suggest that Lso has a wider natural host range within the Apiaceae family than expected. To identify the Lso haplotypes present in France, we sequenced and analyzed the 16S rRNA gene and the 50S ribosomal protein rpIJ-rpIL gene region from a representative bacterial collection of 44 Lso-positive samples. Our SNP analysis revealed the occurrence of two distinct bacterial lineages that correspond to haplotypes D and E. Then, we assessed the phylogenetic relationships between strains isolated from France and a worldwide collection of Lso isolates by using the rpIJ-rpIL gene region sequences. The neighbor-joining tree constructed delineated five clusters corresponding to the five Lso haplotypes, with LsoD and LsoE being closely related phylogenetically. Altogether, the data presented here constitute a first step toward a better understanding of the genetic diversity among Lso haplotypes in France, and provide new insights into the host range of this emerging bacterial species.