Characterization, Phylogenetic Analyses, and Pathogenicity of Enterobacter cloacae on Rice Seedlings in Heilongjiang Province, China

Abstract

Rice is used as a staple food in different areas of world, especially in China. In recent years, rice seedlings have been affected seriously by symptoms resembling bacterial palea browning (BPB) in Heilongjiang Province. To isolate and identify the pathogenic bacteria responsible for the disease, 40 bacterial strains were isolated from diseased rice seedlings collected from the four major accumulative-temperature zones of rice fields cultivated in Heilongjiang Province, and these were identified as 13 species based on morphological characteristics and 16S ribosomal RNA (rRNA) gene sequences. Inoculation of all the isolates on healthy rice seedlings showed that the nine Enterobacter cloacae isolates were the pathogens causing typical symptoms of BPB, including yellowing to pale browning, stunting, withering, drying, and death. Moreover, the nine E. cloacae isolates could also cause symptoms of bacterial disease on the seedlings of soybean (Glycine max), maize (Zea mays L.), and tomato (Solanum lycopersicum). Phylogenetic analysis based on the 16S rRNA gene sequences and phenotypic and biochemical characteristics indicated that these nine pathogenic isolates were E. cloacae. In addition, analysis of the sequences of four housekeeping genes (rpoB, gyrB, infB, and atpD) from the selected strain SD4L also assigned the strain to E. cloacae. Therefore, E. cloacae is the pathogen causing disease of rice seedlings in Heilongjiang Province, which we propose to classify as a form of BPB. To the best of our knowledge, this is the first study to identify E. cloacae as a causal agent of BPB in rice.

Microbacterium helvum sp. nov., a novel actinobacterium isolated from cow dung

“…The pH range for growth (pH 4-12, at intervals of 1 pH units) was tested in GY broth. The buffer systems (Cao et al 2020;Zhao et al 2019). NaCl tolerance was determined in GY broth supplemented with 0-10 % NaCl (w/v, with an interval of 1 % w/v) at 28 ºC for 14 days on an rotary shaker.…”

Microbispora sitophila sp. nov., a novel actinobacterium isolated from rhizosphere soil of wheat (Triticum aestivum L.)

“…at 28 °C. The buffer systems were: pH 4.0–5.0, 0.1 M citric acid/0.1 M sodium citrate; pH 6.0–8.0, 0.1 M KH 2 PO 4 /0.1 M NaOH; pH 9.0–10.0, 0.1 M NaHCO 3 /0.1 M Na 2 CO 3 ; and pH 11.0–12.0, 0.2 M KH 2 PO 4 /0.1 M NaOH [15, 16]. Hydrolysis of Tweens (20, 40 and 80) and production of urease were tested as described by Smibert and Krieg [17].…”

“…Growth at different temperatures (4, 10,15,18,20,25,28,35,37,40,45 and 50 °C) was determined on ISP 3 agar after incubation for 14 days. The pH range for growth (pH 4.0-12.0, at intervals of 1.0 pH unit) was tested in ISP 2 broth and NaCl tolerance (0, 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5 and 7 % w/v) for growth was determined after [15,16]. Hydrolysis of Tweens (20, 40 and 80) and production of urease were tested as described by Smibert and Krieg [17].…”

“…The buffer systems were: pH 4.0-5.0, 0. [15,16]. Hydrolysis of Tweens (20, 40 and 80) and production of urease were tested as described by Smibert and Krieg [17].…”

Nocardia bovistercoris sp. nov., an actinobacterium isolated from cow dung

“…Growth at different temperatures (10,15,18,20,25,28,32,35,38,40 and 45 °C) was determined on ISP 2 agar after incubation for 14 days. Growth tests were performed in the pH range of pH 4.0-12.0 (with intervals of 1.0 pH unit) using a buffer system described by Zhao et al [16] and Cao et al [17]. NaCl tolerance (0, 1, 2, 3, 4, 5 %, w/v) was tested in ISP 2 broth at 28 °C for 14 days on a rotary shaker.…”