Pseudomonas cichorii as the causal agent of midrib rot, an emerging disease of greenhouse-grown butterhead lettuce in Flanders.
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.
Cottyn
Bart
B
Plant-Crop Protection, Institute for Agricultural and Fisheries Research (ILVO), Burg. Van Gansberghelaan 96, B-9820 Merelbeke, Belgium. bart.cottyn@ilvo.vlaanderen.be
Heylen
Kim
K
Heyrman
Jeroen
J
Vanhouteghem
Katrien
K
Pauwelyn
Ellen
E
Bleyaert
Peter
P
Van Vaerenbergh
Johan
J
Höfte
Monica
M
De Vos
Paul
P
Maes
Martine
M
D015373:Bacterial Typing Techniques; D001530:Belgium; D004269:DNA, Bacterial / Q000302:isolation & purification; D004275:DNA, Ribosomal / Q000032:analysis; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D005227:Fatty Acids / Q000032:analysis; D018545:Lactuca / Q000382:microbiology*; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010935:Plant Diseases / Q000382:microbiology*; D016133:Polymerase Chain Reaction / Q000379:methods; D011133:Polysaccharide-Lyases / Q000378:metabolism; D011549:Pseudomonas / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification* / Q000502:physiology; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D017422:Sequence Analysis, DNA
D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D005227:Fatty Acids; D012336:RNA, Ribosomal, 16S; D012321:DNA-Directed RNA Polymerases; C105078:RNA polymerase beta subunit; D011133:Polysaccharide-Lyases; C023911:pectate lyase