Bacterial Dry Stalk Rot In Maize, A New Disease

Maize (Zea mayz L.) is an important crop with enhanced production in recent years. In maize production, pests and diseases are the main limiting factors of high and stable yield. The occurrence of new pests and diseases or those previously occurred in limited areas have brought new threats to maize production in China. In July 2006, a new maize stalk rot in the paternal parent PS056 was observed in the hybrid seed production fields in Xinjiang and Gansu provinces, China, which was different from the previous reports. The diseased plants were scattered throughout the fields and 80% to 100% plants of the male parental line PS056 were infected, which resulted in serious yield loss because of poor pollen shed. Disease symptoms included slow growth and dwarf in diseased plants. The infected tissues showed dry rot symptoms. The symptoms of this disease were different from those of the maize bacterial stalk rot in previous reports. Thus, the new disease on maize was designated bacterial dry stalk rot. In this study, the newly found disease was discussed in details. The main results are as follows:1. From 2006 to 2009, a new bacterial disease with dry rot symptom on stalks has frequently occurred in the maize hybrid seed production fields in Xinjiang and Gansu provinces, China. The typical symptoms of the disease were the infected plants grew slower and became dwarf, and irregular dark brown spot on cortex and dry rot on stalk of maize. Sometimes, the infected parts of stalks became black and formed concavity due to browned cortex of stalk disappeared. The diseased tissues showed typical dry rot symptoms. The diseased plants were lopsided and easily broken because only one side of the stalk was infected. Based on examination results of microscope, electron microscopy, pathogenicity assay, physiological and biochemical tests,16S rDNA sequence analysis and whole-cell fatty acid identification, etc., the pathogenic bacteria isolated from the infected stalk that caused bacterial stalk rot on Xinjiang and Gansu maize was identified as Pantoea agglomerans. This is the first report of dry stalk rot disease on maize caused by P. agglomerans.2. A pair of specific primers Pa1/Pa2 (forward primer: 5'GCTACAATGGCGCATACAA 3', reverse primer:Pa2: 5'CGTCCTGCTCGTTACTTTT 3') were obtained by screening bacterial 16S rDNA sequences 16S-23S rDNA ITS sequences, which can specifically detect P. agglomerans in the target bacteria and reference strains. The PCR amplified production from the target bacteria was 375 bp. A molecular detection technique for P. agglomerans was established, and the specific primers Pa1/Pa2 can detect the target bacteria in the maize seeds and soil.3. Seeds infection results indicated that paternal PS056 and F, Jinyu 9856 seeds only carried bacteria internally but not externally; maternal OSL190 seeds carried bacteria neither internally nor externally. PCR amplification with specific primers showed that the same specific target band (375 bp) can be amplified from isolates Pagl isolated from paternal F1 Jinyu 9856 and isolates Pag2 isolated from PS056. The greenhouse and field experiments showed that the pathogenic bacteria can be transmitted through seeds. The P. agglomerans of soil was also one of the main sources of infection, and the pathogenic bacteria of pollen maybe the main source of infection in the transmittion from male parent to hybrid variety F1 seeds. 4. A transposon mutant library of the pathogenic strain P. agglomerans XJ1 was constructed by transposon Tn5 random insertion mutation. Eleven mutants which lost pathogenicity were obtained by injecting inoculation on maize stalks ex vivo. By amplifying the flank sequences of Tn5 insertion sites, it was confirmed that non-pathogenic mutation strains PA 121 and PA71 were due to mutation of genes yhfK and flhD. The mutation strains PA 121 and PA71 had a single-copy of Tn5 transposon. By regulating gene hrpA transcription, gene yhfK induced lost pathogenicity of P. agglomerans on maize. The mutation of gene flhD caused the loss of flagellum development and swimming motility of P. agglomerans, thus the mutation of gene flhD induced lost pathogenicity of P. agglomerans on maize.5. Using the F2 population derived from the cross between the resistant parent OSL190 and the susceptible parent PS056 and relative parental lines, the inheritance of resistance to P. agglomerans of maize was studied. Inoculation with the strain XJ 1 of P. agglomerans indicated that all the plants of the paternal parent PS056 were susceptible, but the plants of the maternal parent OSL190 were all resistant. The plants of reciprocal Fj cross were also resistant. The segregation of resistant to susceptible plants fitted the ratio of 3:1, indicating that OSL190 carries a single dominant gene for resistance and PS056 carries a single recessive gene for susceptibility. The inbred lines 78599-1, Shen137, Qi319, E28, and XI78 in the pedigree of lne PS056 were all resistant, while Dan 341 and Dan 340 were severely susceptible. This indicates that the susceptibilility was most probably derived from Dan 341.