A novel Gram-stain-negative, aerobic, motile bacterial strain, D13-10-4-6T, was isolated from the bark sample of Populus × euramericana. The strain could grow at 15–35°C, at pH 6–10 and in 0–4% (w/v) NaCl, and the strain tested positive for oxidase and catalase activities. The main polar lipids were phosphatidylmonomethylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, and phosphatidylethanolamine. The main respiratory quinone was Q-10, and the predominant fatty acid was C18:1 ω7c. The phylogenetic analyses showed that the strain belonged to the genus Pseudogemmobacter of the family Rhodobacteraceae. The family Rhodobacteraceae is an ecologically diverse group that includes bacteria from aquatic to terrestrial ecosystems. As a consequence, the classification of the family Rhodobacteraceae is difficult, not least when the early taxonomy work relied heavily on 16S rRNA gene analysis. Recently, the taxonomic status of many members of the family has been revised based on the genome analysis; however, there are still some classification conflicts due to the lack of genome sequences and parallel publication time. In this study, phylogenetic trees based on 16S rRNA gene, gyrB gene, and 120 concatenated proteins, the average amino acid identity (AAI) and percentage of conserved proteins (POCP) have been used for the analysis of strain D13-10-4-6T and other members of 15 genera within the family to further clarify their taxonomic relationships. For the data of phylogeny, AAI, and POCP, the taxonomic proposals are (1) reclassification of Rhodobacter tardus as the type species of a novel genus, Stagnihabitans gen. nov., as Stagnihabitans tardus comb. nov.; (2) reclassification of Tabrizicola alkalilacus, Tabrizicola sediminis, Tabrizicola algicola into a novel genus, Pseudotabrizicola gen. nov., as Pseudotabrizicola alkalilacus comb. nov., Pseudotabrizicola sediminis comb. nov., Pseudotabrizicola algicola comb. nov.; (3) reclassification of Rhodobacter sediminicola into the genus Cereibacter as Cereibacter sediminicola comb. nov.; (4) reclassification of Rhodobacter flagellatus, Rhodobacter thermarum, and Xinfangfangia soli into the genus Tabrizicola as Tabrizicola flagellatus comb. nov., Tabrizicola thermarum comb. Nov., and Tabrizicola soli comb. nov.; (5) reclassification of Xinfangfangia humi into the genus Pseudogemmobacter as Pseudogemmobacter humicola comb. nov.; (6) classification of strain D13-10-4-6T as a novel species of the genus Pseudogemmobacter, for which the name P. hezensis sp. nov. is proposed, the type strain is D13-10-4-6T (= CFCC 12033T = KCTC 82215T).

The introduction of the pine wood nematode (PWN), Bursaphelenchus xylophilus, to new areas has impacted on the international economy. Therefore, accurate and reliable detection methods for PWN are essential for the control and management of this pest. A rapid and economic method for detecting PWN may be developed focusing on the PWN vector (Monochamus alternatus). This work standardized a loop-mediated isothermal amplification (LAMP) method using newly designed primer sequences based on the syg-2 gene, which encodes the synaptogenesis protein syg-2. Loopmediated isothermal amplification was conducted at 63°C for 60 min using six sets of primers. The result was confirmed by visual observation. A positive reaction was confirmed by SYBR Green I fluorescence dye under light thermal cycling. The lower limit of DNA detection was 51.4 pg/μl in both LAMP and 51.4 ng/μl in PCR. Therefore, the LAMP was 1,000 times more sensitive in DNA detection than PCR. The LAMP is a relatively new, highly accurate and rapid molecular technique that can rapidly detect infectious agents in the field without requiring sophisticated instruments, giving a visually readable result. The method greatly improves detection, without requiring professional knowledge and expensive, sophisticated equipment. Therefore, this system is suitable for quarantine and field detection. negative reaction (no target DNA template), the dye remained orange ( Figure 4). | DISCUSSIONBursaphelenchus xylophilus is an important pathogenic organism vectored between pine hosts by M. alternatus. Therefore, the number of M. alternatus is a crucial indicator of disease development. Unless B. xylophilus in M. alternatus is specifically treated in the primary phase of infection, PWD is fatal. The efficient, rapid detection of B. xylophilus from M. alternatus is necessary for monitoring and managing outbreaks, and in implementing specific therapeutics (Cardoso, Fonseca, & Abrantes, 2012; Wang et al., 2009). As the death of pines infected by PWD is very rapid, the novel PWD-detection technique must also be rapid.In the present work, we designed specific primers for rapid detection of B. xylophilus from M. alternatus using the LAMP. For this purpose, we targeted the conserved syg-2 gene reported by Gou (2014). The LAMP assay successfully detected all isolates of B. xylophilus, giving negative reactions in all other species tested. The high specificity of the LAMP is conferred by the six specially designed primers that recognize six regions on the syg-2 gene sequence, which are specific to B. xylophilus. The lower limit of DNA detection in the LAMP-based technique was 51.4 pg/μl of genomic DNA, compared with 51.4 ng/μl in conventional PCR. Therefore, the LAMP was 1,000-fold more sensitive than conventional PCR (Gou et al., 2015) and can rapidly detect B. xylophilus from M. alternatus.Given the high sensitivity of LAMP, post-amplification operations should be performed in a separate room, away from the reactions and reagents used in the PCR and LAMP, to reduce the chance of conta...

Gnomoniopsis (Gnomoniaceae, Diaporthales) is a well-classified genus inhabiting leaves, branches and fruits of the hosts in three plant families, namely Fagaceae, Onagraceae and Rosaceae. In the present study, eighteen Gnomoniopsis isolates were obtained from diseased leaves of Fagaceae hosts collected from Fujian, Guangdong, Hainan, Henan, Jiangxi and Shaanxi provinces in China. Morphology from the cultures and phylogeny based on the 5.8S nuclear ribosomal DNA gene with the two flanking internally transcribed spacer (ITS) regions, the translation elongation factor 1-alpha (tef1) and the beta-tubulin (tub2) genes were employed to identify these isolates. As a result, seven species were revealed, viz. Gnomoniopsis castanopsidis, G. fagacearum, G. guangdongensis, G. hainanensis, G. rossmaniae and G. silvicola spp. nov, as well as a known species G. daii. In addition, G. daii was firstly reported on the host Quercus aliena.

A Gram-stain-negative, yellow-pigmented, ovoid to rod-shaped, strictly aerobic bacterial strain, designated 100921-2T, was isolated from air at the foot of Xiangshan Mountain. Phylogenetic and phenotypic analysis of the organism revealed that the isolate belongs to the genus Altererythrobacter. Strain 100921-2T showed high 16S rRNA gene sequence similarity (96.01-94.70 %) to other type strains of the genus Altererythrobacter, with the highest similarity to Altererythrobactermarensis MSW-14T. Growth of strain 100921-2T was observed at 4-50 °C (optimum, 30 °C), at pH 4.5-10.0 (optimum, pH 7.0) and at salinities of 0-10 % (w/v) NaCl (optimum 0-0.5 %). The major fatty acids were C18 : 1ω7c (27.8 %), C17 : 1ω6c (23.1 %), 11-methyl C18 : 1ω7c(11.9 %), summed feature 3 (9.1 %) and C15 : 0 2-OH (7.9 %). The predominant respiratory quinone was ubiquinone-10 (Q-10). Polar lipid analysis indicated the presence of diphosphatidylglycerol, sphingoglycolipid, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylcholine, two unknown phospholipids, five unknown polar lipids and two unknown glycolipids. The DNA G+C content of the type strain was 67.5 mol%. On the basis of the data from the polyphasic characterization, strain 100921-2T represents a novel species, for which the name Altererythrobacter aerius sp. nov. is proposed. The type strain is 100921-2T (=CFCC 14287T=KCTC 42844T).

Five Gran-stain-negative, facultatively anaerobic, motile, bacterial strains were isolated from symptomatic bark tissue of Populus¾euramericana canker. Strains grew at 4-41 6C, pH 4-10 and 0-6 % (w/v) salinity. They were positive with respect to catalase activity and negative for oxidase activity, nitrate reduction and the Voges-Proskauer reaction. Analysis of 16S rRNA gene sequences indicated that these five poplar isolates belong to the genus Brenneria, having highest sequence similarity of 95.98 % with Brenneria goodwinii LMG 26270 T . These five isolates formed a single cluster based on multilocus sequence analysis, indicating that they all belong to a single taxon within the genus Brenneria, which was confirmed by DNA-DNA hybridization. The DNA G+C content was 54.9-55.7 mol%, and the main fatty acids were C 16 : 0 , C 18 : 1 v7c, C 17 : 0 cyclo and C 16 : 1 v7c/iso-C 15 : 0 2-OH. Based on these results, we describe a novel species of the genus Brenneria with the proposed name Brenneria populi sp. nov. The type strain is D9-5 T (5CFCC 11963 T 5KCTC 42088 T ).The genus Brenneria was first established by Hauben et al. (1998a). At the time of writing, the genus Brenneria comprises five species with validly published names. Almost all species of the genus are plant pathogens. Brenneria salicis (the type species of the genus) causes willow (Salix spp.) watermark disease, and occurs mainly in the xylem vessels of the host plants (Hauben et al., 1998b). There are also three species which can cause plant bark canker disease. Brenneria nigrifluens and Brenneria rubrifaciens are the causal agents of bark canker and deep bark canker of walnut, respectively (Loreti et al., 2008;McClean et al., 2008). Brenneria alni is the causal agent of bark canker of alder (Surico et al., 1996). Moreover, another two species, namely Brenneria goodwinii and 'Brenneria roseae' (including two subspecies), which were isolated from symptomatic oak tissues, are both associated with acute oak decline (Denman et al., 2012;Brady et al., 2014). Here, we describe the phenotypic and genotypic properties of five novel strains representing the genus Brenneria isolated from symptomatic bark of Populus6euramericana canker in Henan Province of China. The Populus6euramericana canker with abundant white, sour exudates on poplar trees more than five years old were observed for the first time in China's Henan and Shandong provinces in 2006. Diseased plants had stem or branch bark that cracked and exuded frothy fluid. When the disease progressed, many cankers (50-15063-8 cm, width by length) appeared rapidly (Li et al., 2014). Lonsdalea quercina subsp. populi proved to be the causal agent of the bark canker (Li et al., 2014).During our research on pathogen isolation from Populus6euramericana canker, besides the pathogen L. quercina subsp. populi, five other strains (D9-5 T 5CFCC

Colletotrichum is an important plant pathogenic genus causing anthracnose on a wide range of host plants. During 2019 and 2021, Colletotrichum isolates were obtained during surveys of anthracnose on garden plants in China. Multi-gene phylogenetic analyses of internal transcribed spacer (ITS), glyceraldehyde-3-phosphate dehydrogenase (gapdh), chitin synthase 1 (chs-1), actin (act) and beta-tubulin (tub2) sequences coupled with morphological evidence support the introduction of two novel species namely Colletotrichum chinense sp. nov. from Yucca gloriosa in Beijing and C. quercicola sp. nov. from Quercus variabilis in Shaanxi Province. Phylogenetic inference revealed that two isolates of C. chinense belonged to the agaves species complex and were closely related to C. agaves, and differed from the other species within this species complex by shorter conidia and the host association. Molecular identification showed that two isolates of C. quercicola formed a highly supported lineage close to C. tanaceti in the destructivum species complex, which could be distinguished from C. tanaceti by straighter conidia. In pathogenicity tests, yellow spots and orange conidial masses displayed on the inoculated Y. gloriosa leaves and brown spots appeared on the inoculated Q. variabilis leaves. In addition, C. chinense and C. quercicola were re-isolated from spots of the tested leaves of Y. gloriosa and Q. variabilis.

Two Gram-negative, non-motile, rod-shaped bacterial strains, 2BJ1 T and 2C-3-1, were isolated from canker bark of Populus ¾ euramericana collected from different locations in Puyang City, Henan Province, China. The two strains were characterized using nutritional and physiological testing and DNA sequence analysis. They were found to produce acid from D-glucose. Haemolysis was not observed on agar medium supplemented with sheep erythrocytes. Phylogenetic analysis based on 16S rRNA, rpoB and gyrB gene sequences revealed that the strains formed a distinct cluster with 100 % bootstrap support within the genus Acinetobacter in all phylogenetic trees. The phenotypic characteristics most useful for the differentiation of the two strains from other species of the genus Acinetobacter were their ability to grow at 41 6C and to assimilate malonate, phenylacetate and trigonelline. Based on phenotypic, genotypic and phylogenetic characteristics, the two strains are considered to represent a novel species of the genus Acinetobacter, for which the name Acinetobacter qingfengensis sp. nov. is proposed. The type strain is 2BJ1 T (5CFCC 10890 T 5KCTC 32225 T ).

The family Rhizobiaceae contains 19 validly described genera including the rhizobia groups, many of which are important nitrogen-fixing bacteria. Early classification of Rhizobiaceae relied heavily on the poorly resolved 16S rRNA genes and resulted in several taxonomic conflicts. Although several recent studies illustrated the taxonomic status of many members in the family Rhizobiaceae, several para- and polyphyletic genera still needed to be elucidated. The rapidly increasing number of genomes in Rhizobiaceae has allowed for a revision of the taxonomic identities of members in Rhizobiaceae. In this study, we performed analyses of genome-based phylogeny and phylogenomic metrics to review the relationships of 155-type strains within the family Rhizobiaceae. The UBCG and concatenated protein phylogenetic trees, constructed based on 92 core genes and concatenated alignment of 170 single-copy orthologous proteins, demonstrated that the taxonomic inconsistencies should be assigned to eight novel genera, and 22 species should be recombined. All these reclassifications were also confirmed by pairwise cpAAI values, which separated genera within the family Rhizobiaceae with a demarcation threshold of ~86%. In addition, along with the phenotypic and chemotaxonomic analyses, a novel strain BDR2-2T belonging to a novel genus of the family Rhizobiaceae was also confirmed, for which the name Ectorhizobium quercum gen. nov., sp. nov. was proposed. The type strain is BDR2-2T (=CFCC 16492T = LMG 31717T).