Pseudomonas cichorii as the causal agent of midrib rot, an emerging disease of greenhouse-grown butterhead lettuce in Flanders

Abstract

Bacterial midrib rot of greenhouse-grown butterhead lettuce (Lactuca sativa L. var. capitata) is an emerging disease in Flanders (Belgium) and fluorescent pseudomonads are suspected to play an important role in the disease. Isolations from infected lettuces, collected from 14 commercial greenhouses in Flanders, yielded 149 isolates that were characterized polyphasically, which included morphological characteristics, pigmentation, pathogenicity tests by both injection and spraying of lettuce, LOPAT characteristics, FAME analysis, BOX-PCR fingerprinting, 16S rRNA and rpoB gene sequencing, as well as DNA-DNA hybridization. Ninety-eight isolates (66%) exhibited a fluorescent pigmentation and were associated with the genus Pseudomonas. Fifty-five of them induced an HR+ (hypersensitive reaction in tobacco leaves) response. The other 43 fluorescent isolates were most probably saprophytic bacteria and about half of them were able to cause rot on potato tuber slices. BOX-PCR genomic fingerprinting was used to assess the genetic diversity of the Pseudomonas midrib rot isolates. The delineated BOX-PCR patterns matched quite well with Pseudomonas morphotypes defined on the basis of colony appearance and variation in fluorescent pigmentation. 16S rRNA and rpoB gene sequence analyses allowed most of the fluorescent isolates to be allocated to Pseudomonas, and they belonged to either the Pseudomonas fluorescens group, Pseudomonas putida group, or the Pseudomonas cichorii/syringae group. In particular, the isolates allocated to this latter group constituted the vast majority of HR+ isolates and were identified as P. cichorii by DNA-DNA hybridization. They were demonstrated by spray-inoculation tests on greenhouse-grown lettuce to induce the midrib rot disease and could be re-isolated from lesions of inoculated plants. Four HR+ non-fluorescent isolates associated with one sample that showed an atypical midrib rot were identified as Dickeya sp.

Pseudomonas viridiflava, a Multi Host Plant Pathogen with Significant Genetic Variation at the Molecular Level

“…Detailed characterization of the genetic variability among isolates belonging to P. viridiflava species was achieved by DNA fingerprinting, based on BOX-and ERIC-PCR (collectively known as rep-PCR) [23]–[25], as was previously discribed [52]. PCR reaction contained 150 ng template DNA, each of the deoxynucleoside triphosphates at a concentration of 250 nM, primers at a total concentration of 2.5 µM, 2.5 mM MgCl 2 , and 2 units of Taq DNA polymerase (Kapa Biosystems) in a total volume of 20 µl.…”

First report on the whole genome sequence of Pseudomonas cichorii strain JBC1 and comparison with other Pseudomonas species

“…For example, Cottyn et al . () reported that P. cichorii is the causal agent of midrib rot on greenhouse‐grown butterhead lettuce in Belgium and Mirik et al . () isolated P. cichorii from stem and pith necrosis of tomato and leaf spot of the dwarf umbrella tree in Turkey.…”

Epidemiology of Pseudomonas cichorii, the Cause of Lettuce Midrib Rot

“…The bacterial strains used are listed in Table 1. SF and AF strains were isolated from symptomatic endive and lettuce heads in Belgium as described by Cottyn et al. (2009).…”

“…2009) using overhead sprinkler irrigation. This intensive management has been accompanied by the emergence of a new disease termed bacterial midrib rot, caused by Pseudomonas cichorii (Cottyn et al. 2009).…”

“…Pseudomonas cichorii strains, from symptomatic lettuces, are a diverse group. Belgian P. cichorii strains, isolated from symptomatic butterhead lettuces, can be divided into three subgroups C1, C2 and C3 based on morphological characteristics and BOX‐PCR genomic fingerprinting, whereas a Californian ‘varnish spot’ isolate, P. cichorii 9D42, did not cluster with any of the three subgroups, although it was most related to subgroup C3 isolates (Cottyn et al. 2009).…”