Phylogenetic tree of <i>Bacillus</i> species based on 16S rRNA gene sequences.

<p>The tree was constructed using the neighbor-joining method and genetic distances were generated using the Kimura 2-parameter method. The numbers at the branches are bootstrap confidence percentages from 1000 bootstrapped trees. <i>Alicyclobacillus acidocaldarius</i> (GenBank accession no. AB089859) was used as the outgroup. The numbers in parentheses indicate the GenBank accession numbers.</p

Bacterial strains and plasmids used in this study.

a<p>Species attributes of native <i>Bacillus</i> strains were identified based on Biolog analysis except for strain OF3-16, which was identified based on physiological and biochemical characteristics described in the Bergey's Manual of Systematic Bacteriology <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0042124#pone.0042124-Sneath1" target="_blank">[39]</a>.</p>b<p>ATCC, American Type Culture Collection Center; BCRC, Bioresource Collection and Research Center, Taiwan; DSM, Leibniz Institute DSMZ-German Collection of Microorganisms and Cell Cultures; NCBI, National Center for Biotechnology Information.</p

Antagonistic activity of <i>Bacillus</i> species against <i>X. axonopodis</i> pv. <i>citri</i> XW19.

<p>A 20 µl aliquot of <i>X. axonopodis</i> pv. <i>citri</i> XW19 suspension (OD₆₂₀ = 0.3) was spread on an SYB agar plate. Following overnight incubation at 30°C, 20 µl of <i>Bacillus</i> suspension (OD₆₂₀ = 0.3) was spotted (A) inside the stainless steel ring or (B) on a paper disc. The plates were incubated at 30°C, for 5 days. CK, 20 µl of sterile Milli-Q water was used as a control. The results represent the means and standard deviations (error bars) of a representative experiment. Different lowercase letters indicate significant differences (p<0.05) according to Tukey's HSD test.</p

tDNA-PCR fingerprint and UPGMA cluster analysis of <i>Bacillus</i> species.

<p>(A) tDNA-PCR fingerprint and (B) UPGMA cluster analysis. The UPGMA cluster analysis was based on tDNA-PCR. M, GeneRuler™ 100 bp plus DNA ladder (Fermentas, Taipei, Taiwan).</p

The effect of application time and frequency of <i>B. subtilis</i> strain TKS1-1 on symptom development and disease incidence of citrus bacterial canker on navel orange grown in the greenhouse.

<p><i>B. subtilis</i> strain TKS1-1 endospore formulation (T, 10⁹ CFU/ml) and <i>X. axonopodis</i> pv. <i>citri</i> XW19 (B, 10⁸ CFU/ml) were used. Treatment T-B-T/1WK, strain TKS1-1 endospores were applied 24 h prior to inoculation with <i>X. axonopodis</i> pv. <i>citri</i> XW19, and then weekly post-pathogen inoculation for 4 weeks; treatment T-B-D/1WK, strain TKS1-1 endospores were applied 24 h prior to <i>X. axonopodis</i> pv. <i>citri</i> XW19 inoculation; treatment D-B-T/1WK, strain XW19 was inoculated, then strain TKS1-1 endospores were sprayed every week post-pathogen inoculation for 4 weeks; treatment D-B-D/1WK, only strain XW19 was inoculated. Without <i>Bacillus</i> treatment, Milli-Q water (D) was sprayed on the leaf surface. (A) Symptoms on upper (left) and lower (right) leaf surfaces after different treatments. Scale bars, 1 cm. (B) Disease incidence was rated 4 weeks post-inoculation. Bars indicate standard deviations. Columns that are top-labeled with different letters are significantly different (p<0.05) according to one-way ANOVA and Tukey's HSD test.</p

ITS-PCR fingerprint and UPGMA cluster analysis of <i>Bacillus</i> species.

<p>(A) ITS-PCR fingerprint and (B) UPGMA cluster analysis. The UPGMA cluster analysis was based on ITS-PCR. M, GeneRuler™ 100 bp plus DNA ladder (Fermentas, Taipei, Taiwan).</p