Genus Leifsonia Research Articles

Comparative Genomic Understanding of Gram-Positive Plant Growth-Promoting Leifsonia

Root-associated bacteria play critical roles in shaping plant health for abiotic and biotic stress tolerance. The Leifsonia genus includes pathogenic and beneficial bacteria but limited research is available comparatively across species regarding their association with plants. We recently identified the plant growth-promoting Leifsonia sp. C5G2 isolated from the rhizosphere of Plectranthus scutellarioides (coleus) that increased stress tolerance and quality of water-stressed greenhouse ornamentals. Using this beneficial strain as a model, we performed comparative genomics with beneficial, commensal, and pathogenic Leifsonia spp. to better understand plant association and growth promotion under abiotic stress. Comparative phylogenetic analyses showed that beneficial and commensal Leifsonia spp. clustered separately from pathogenic Leifsonia xyli subsp. xyli. In addition, the beneficial or commensal strains share biosynthetic gene and orthologous protein clusters that are not shared with known phytopathogens in our analysis. The phytopathogenic Leifsonia strains also share noticeably higher percent homology with plant-associated virulence factors in closely related Clavibacter spp. than the beneficial or commensal strains. Genome analysis supplemented with in vitro assays validated the findings that our model growth-promoting Leifsonia sp. strain C5G2 was highly prolific on diverse carbon sources. This is unlike known fastidious growth behaviors of phytopathogenic Leifsonia spp. Beneficial Leifsonia sp. C5G2 imparts water stress tolerance in plants, and multiple genes were identified in the genomes of beneficial Leifsonia strains that are potentially involved in bacterial osmotic stress and the production of osmoprotectants and vitamin B9. This work sheds light on the diversity of the genus Leifsonia, with a better understanding of potential mechanisms employed by poorly understood beneficial bacteria.

Leifsonia flava sp. nov., a novel actinobacterium isolated from the rhizosphere of Aquilegia viridiflora.

SYP-B2174T is a yellow-pigmented, Gram-positive, non-motile, and rod-shaped actinobacterium isolated from the rhizospheric soil of Aquilegia viridiflora Pall. collected from the Xinjiang uygur autonomous region of China. The strain's growth temperature ranges from 1 to 35°C, with an optimal growth being observed at 28°C. Growth occurs from 0 to 5% NaCl and at pH 6-8, with optimal growth being observed in 1% NaCl at pH 7. Comparative 16S rRNA gene sequence-based phylogenetic analysis placed the strain in a clade with the species Leifsonia kafniensis JCM 17021T and Leifsonia psychrotolerans DSM 22824T with similarities of 97.8 and 97.6%, respectively. The DNA-DNA hybridization values of the strain SYP-B2174T to its closest phylogenetic neighbors were significantly lower than 35.7%. The strain was identified as a novel species of the genus Leifsonia judging by the coryneform morphology, peptidoglycans based upon 2,4-diaminobutyric acid, principal phospholipids phosphatidylglycerol and diphosphatidylglycerol, major menaquinone MK-11, predominant fatty acids of anteiso-C15:0, anteiso-C17:0, and iso-C16:0, and a DNA G + C base composition of 68.7 mol%, for which the name Leifsonia flava sp. nov. is proposed. The type strain is SYP-B2174T (= CGMCC 1.15856T = DSM 105144T = KCTC 39963T).

Molecular Detection of Diverse Leifsonia Strains Associated With Sugarcane.

Leifsonia xyli subsp. xyli, causal agent of ratoon stunting disease (RSD) of sugarcane (Saccharum interspecific hybrids), is the most well-known member of the Microbacteriaceae genus Leifsonia. However, the presence of other Leifsonia strains associated with sugarcane has not been reported. A total of 697 Australian and 40 Indonesian sugarcane fields were screened by leaf sheath biopsy (LSB) PCR using primers specific for L. xyli subsp. xyli, in addition to primers designed to amplify DNA from other members of the genus Leifsonia. While L. xyli subsp. xyli was detected in 126 fields, a total of 37 distinct and novel Leifsonia and non-Leifsonia strains were detected in 116 fields. Representatives of these strains were also detected in multiple samples of expressed xylem sap. Sequencing and phylogenetic analyses demonstrated the presence of a broad complex of novel Leifsonia strains, in addition to strains closely related to the recently erected Cnuibacter genus. Attempts to isolate Leifsonia strains were unsuccessful; however, one strain related to Cnuibacter was recovered from expressed xylem sap. Among the genetically diverse lineages discovered, identical genotypes were present in multiple sugarcane varieties growing in disparate regions in different years, strongly suggesting an ongoing association with sugarcane. The epidemiological significance of these strains is unknown, but there is evidence that they can interfere with serological and microscopic RSD diagnostics, and there is the potential that they may represent new and distinct pathologies of sugarcane.

Leifsonia psychrotolerans sp. nov., a psychrotolerant species of the family Microbacteriaceae from Livingston Island, Antarctica

A cold-tolerant, yellow-pigmented, Gram-positive, motile, facultatively anaerobic bacterial strain, LI1(T), was isolated from a moss-covered soil from Livingston Island, Antarctica, near the Bulgarian station St. Kliment Ohridski. Comparative 16S rRNA gene sequence-based phylogenetic analysis placed the strain in a clade with the species Leifsonia kafniensis KFC-22(T), Leifsonia pindariensis PON10(T) and Leifsonia antarctica SPC-20(T), with which it showed sequence similarities of 99.0, 97.9 and 97.9 %, respectively. DNA-DNA hybridization revealed a reassociation value of 2.7 % with L. kafniensis LMG 24362(T). The DNA G+C content of strain LI1(T) was 64.5 mol%. The growth temperature range was -6 to 28 °C, with optimum growth at 16 °C. Growth occurred in 0-5 % NaCl and at pH 4.5-9.5, with optimum growth in 1-2 % NaCl and at pH 5.5-6.5. The predominant fatty acids were anteiso-C(15 : 0), C(18 : 0) and iso-C(15 : 0). The polar lipids were phosphatidylglycerol and diphosphatidylglycerol. Physiological and biochemical tests clearly differentiated strain LI1(T) from L. kafniensis. Therefore, a novel cold-tolerant species within the genus Leifsonia is proposed: Leifsonia psychrotolerans sp. nov. (type strain LI1(T) = DSM 22824(T) = NCCB 100313(T)).

Reclassification of Leifsonia ginsengi (Qiu et al. 2007) as Herbiconiux ginsengi gen. nov., comb. nov. and description of Herbiconiux solani sp. nov., an actinobacterium associated with the phyllosphere of Solanum tuberosum L.

In the context of studying the effects of transgenic fructan-producing potatoes on the community structure of phyllosphere bacteria, a group of strains closely related to the species Leifsonia ginsengi was isolated. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the new isolates and L. ginsengi DSM 19088(T) formed a lineage at the genus level and this finding was supported by chemotaxonomic characterization. The peptidoglycan type of the representative isolate, K134/01(T), and L. ginsengi DSM 19088(T) was B2γ, with d- and l-diaminobutyric acid as the diagnostic diamino acid and glycine, alanine and threo-3-hydroxyglutamic acid. The almost-complete substitution of glutamic acid by threo-3-hydroxyglutamic acid supported the differentiation of the new strains from recognized species of the genus Leifsonia. Furthermore, the detection of substantial amounts of the fatty acid cyclohexyl-C(17 : 0) in the new isolates and L. ginsengi DSM 19088(T) was a prominent chemotaxonomic feature for a clear demarcation of these strains from all genera of the family Microbacteriaceae that display the B2γ cell-wall type. Comparative phylogenetic and phenotypic analyses of the isolates and L. ginsengi DSM 19088(T) revealed the separate species status of the isolates. On the basis of these results, it is proposed that L. ginsengi should be classified as the type species of a novel genus, Herbiconiux gen. nov., with the name Herbiconiux ginsengi gen. nov., comb. nov. (type strain wged11(T) = CGMCC 4.3491(T) = JCM 13908(T) = DSM 19088(T) = NBRC 104580(T)). The phyllosphere isolates are assigned to a novel species, Herbiconiux solani sp. nov. (type strain K134/01(T) = DSM 19813(T) = LMG 24387(T) = NBRC 106740(T)).

Leifsonia soli sp. nov., a yellow-pigmented actinobacterium isolated from teak rhizosphere soil

Two yellow-pigmented, Gram-stain-positive, aerobic, motile, short rod-shaped bacteria were isolated from natural teak tree rhizosphere soil and their taxonomic positions were determined by using a polyphasic approach. Comparative 16S rRNA gene sequence analysis showed that strains TG-S248(T) and TG-S240 formed a distinct phyletic line within the genus Leifsonia. 16S rRNA gene sequence analysis of strain TG-S248(T) with sequences from Leifsonia shinshuensis DB 102(T), L. poae VKM Ac-1401(T), L. naganoensis DB 103(T), L. aquatica DSM 20146(T) and L. xyli subsp. cynodontis JCM 9733(T) revealed pairwise similarities ranging from 98.7 to 99.1 %. The major fatty acids were anteiso-C(15 : 0), anteiso-C(17 : 0) and iso-C(16 : 0). The G+C content of the DNA of the type strain was 69.4 mol%. DNA-DNA hybridization experiments revealed low levels of DNA-DNA relatedness (32 % or less) between strain TG-S248(T) and its closest relatives. Based on differences in phenotypic and genotypic characteristics, strain TG-S248(T) (=LMG 24767(T) =JCM 15679(T)) is designated the type strain of a novel species of the genus Leifsonia, for which the name Leifsonia soli sp. nov. is proposed.

Description of Leifsonia kafniensis sp. nov. and Leifsonia antarctica sp. nov.

Strains KFC-22(T) and SPC-20(T) are yellow-pigmented, Gram-positive, aerobic, non-motile, rod-shaped bacteria that were isolated from a soil sample near the Kafni glacier in the Himalayan mountain ranges in India, and from a spade core sediment sample from the Antarctic Ocean at Larsemann Hill, respectively. In both cases, the cell-wall peptidoglycan contained 2,4-diaminobutyric acid as the diamino acid, anteiso-C(15 : 0), anteiso-C(17 : 0) and iso-C(16 : 0) were the predominant fatty acids and MK-11 was the major isoprenoid quinone in the cell membrane. On the basis of the above-mentioned characteristics, both strains can be assigned to the genus Leifsonia. The strains share 16S rRNA gene sequence similarity of 97.7 % and DNA relatedness of only 10 %, indicating that they represent different species. A blast analysis indicated that Leifsonia pindariensis PON10(T) was the closest phylogenetic neighbour of strains SPC-20(T) and KFC-22(T), showing 16S rRNA gene sequence similarities of 97.3 and 97.7 %, respectively. However, at the whole-genome level, strains KFC-22(T) and SPC-20(T) shared 42 and 11 % DNA-DNA relatedness, respectively, with L. pindariensis PON10(T). In addition, both strains exhibited several phenotypic differences with respect to L. pindariensis PON10(T). Thus, on the basis of the differences that the two strains exhibited with respect to L. pindariensis, both were identified as representing novel species of the genus Leifsonia, for which the names Leifsonia kafniensis sp. nov. (type strain KFC-22(T) =NCCB 100216(T) =LMG 24362(T)) and Leifsonia antarctica sp. nov. (type strain SPC-20(T) =NCCB 100227(T) =LMG 24541(T)) are proposed.

Leifsonia lichenia sp. nov., isolated from lichen in Japan

An actinobacterial strain, 2SbT, isolated from lichen was characterized taxonomically using a polyphasic approach. Strain 2SbT was Gram-positive, strictly aerobic, rod-shaped and non-motile. Phylogenetic analysis based on 16S rRNA gene sequence revealed that strain 2SbT was located in the genus Leifsonia. Level of 16S rRNA gene sequence similarity between the isolated strains and the type strains of Leifsonia species were 95.0-99.2%. The value of DNA-DNA relatedness between strain 2SbT and Leifsonia poae, a phylogenetically related and phenotypically similar species, was 39.9/25.2%. The DNA G+C content of the strain 2SbT was 61.7 mol%. Cell wall peptidoglycan type (2,4-diaminobutyric acid), major cellular fatty acids (anteiso-C(15:0), anteiso-C(17:0) and iso-C(16:0)) and quinone type (MK-11 and MK-10) of the isolate support their affiliation to the genus Leifsonia. On the basis of phylogenetic position, physiological and chemotaxonomic properties, strain 2SbT represent a novel species of the genus Leifsonia, for which the name Leifsonia lichenia sp. nov. is proposed. The type strain is 2SbT (=IAM 15426T =JCM 23226T =KCTC 13122T).

Leifsonia kribbensis sp. nov., isolated from soil

A yellow-pigmented actinobacterium, designated strain MSL-13T, was isolated from a soil sample collected from Bigeum Island, Republic of Korea, and its taxonomic position was determined by using a polyphasic approach. Strain MSL-13T showed phenotypic and chemotaxonomic properties consistent with its classification in the genus Leifsonia. 16S rRNA gene sequence analysis of strain MSL-13T with sequences from Leifsonia naganoensis DB103T, Leifsonia aquatica DSM 20146T, Leifsonia xyli subsp. cynodontis JCM 9733T, Leifsonia poae VKM Ac-1401T and Leifsonia shinshuensis DB102T revealed similarities of 96.22, 96.19, 95.77, 95.44 and 95.37 %, respectively, with differences of 39-65 nt among 1483 total nucleotides aligned. Based on differences in phenotypic and genotypic characteristics, strain MSL-13T (=KCTC 19267T =DSM 19272T) is designated as the type strain of a novel species of the genus Leifsonia, for which the name Leifsonia kribbensis sp. nov. is proposed.

Leifsonia pindariensis sp. nov., isolated from the Pindari glacier of the Indian Himalayas, and emended description of the genus Leifsonia

Strain PON10(T) is a yellow-pigmented, Gram-positive, aerobic, motile, rod-shaped bacterium isolated from the Pindari glacier of the Indian Himalayas. The cell-wall peptidoglycan contained DL-diaminobutyric acid as the diamino acid. The predominant fatty acids were anteiso-C(15:0), anteiso-C(17:0) and iso-C(16:0) and the major isoprenoid quinones were MK-10 and MK-11. Based on the above characteristics, strain PON10(T) was assigned to the genus Leifsonia. blast sequence similarity results indicated that Leifsonia ginsengi and Leifsonia poae were the nearest relatives, with 16S rRNA gene sequence similarity of 97.0 and 96.8% to the respective type strains. A difference of 3% in the 16S rRNA gene sequence indicated that PON10(T) represents a novel species of the genus Leifsonia, and therefore DNA-DNA hybridization was not done. In addition, PON10(T) showed a number of differences from Leifsonia ginsengi and Leifsonia poae with respect to phenotypic and chemotaxonomic characteristics. Thus, based on the differences it exhibited from Leifsonia ginsengi and Leifsonia poae, strain PON10(T) was identified as representing a novel species named Leifsonia pindariensis sp. nov. The type strain is PON10(T) (=LMG 24222(T) =MTCC9128(T)). An emended description of the genus Leifsonia is also presented.

Leifsonia bigeumensis sp. nov., isolated from soil on Bigeum Island, Korea

A Gram-positive, motile, mesophilic bacterium that formed yellowish-white, circular, convex, glistening colonies, strain MSL-27T, was isolated from a soil sample from Bigeum Island, Korea, and its taxonomic position was investigated by a polyphasic approach. Cells of strain MSL-27T are non-spore-forming, irregular rods, usually fragmenting into shorter rods or coccoid elements. The growth temperature and pH ranges are 20-37 degrees C and pH 6.0-12.0, with optimum growth at 28 degrees C and pH 8.0. MK-11 is the major menaquinone and dl-2,4-diaminobutyric acid is found in the cell-wall peptidoglycan. The major fatty acids are i-C16:0 (39 %), ai-C15:0 (36 %), ai-C17:0 (18 %) and i-C15:0 (2 %). The G+C content is 69.0 mol%. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that strain MSL-27T should be placed in the genus Leifsonia. Levels of similarity between the 16S rRNA gene sequence of strain MSL-27T and those of the type strains of the other members of the genus Leifsonia ranged from 95.8 to 96.7 %. Based on phenotypic characteristics, biochemical properties and chemotaxonomic features, strain MSL-27T (=KCTC 19268T =DSM 19322T) represents the type strain of a novel species of Leifsonia, for which the name Leifsonia bigeumensis sp. nov. is proposed.

Leifsonia ginsengi sp. nov., isolated from ginseng root

A Gram-positive, rod-shaped, non-motile bacterium, designated strain wged11T, was isolated from the root of ginseng, and its taxonomic position was established using a polyphasic approach. Phylogenetic analysis based on 16S rRNA gene sequences showed that this organism formed a robust clade with recognized species of the genus Leifsonia. Strain wged11T was characterized by a high content of omega-cyclohexylundecanoic and anteiso- and iso-branched saturated fatty acids, MK-11 as the major menaquinone and DL-2,4-diaminobutyric acid in its cell-wall peptidoglycan. The DNA G+C content of strain wged11T was 66.4 mol%. Levels of similarity between the 16S rRNA gene sequence of strain wged11(T) and those of the type strains of other members of the genus Leifsonia ranged from 94.7 to 97.6 %. The mean level of DNA-DNA relatedness between strain wged11T and Leifsonia poae DSM 15202T, its nearest phylogenetic neighbour, was 35.3 %. Based on these findings, strain wged11T (=CGMCC 4.3491T=JCM 13908T) is proposed as the type strain of a novel species of the genus Leifsonia, Leifsonia ginsengi sp. nov.

Identification of bacteria on the surface of clinically infected and non-infected prosthetic hip joints removed during revision arthroplasties by 16S rRNA gene sequencing and by microbiological culture

It has been postulated that bacteria attached to the surface of prosthetic hip joints can cause localised inflammation, resulting in failure of the replacement joint. However, diagnosis of infection is difficult with traditional microbiological culture methods, and evidence exists that highly fastidious or non-cultivable organisms have a role in implant infections. The purpose of this study was to use culture and culture-independent methods to detect the bacteria present on the surface of prosthetic hip joints removed during revision arthroplasties. Ten consecutive revisions were performed by two surgeons, which were all clinically and radiologically loose. Five of the hip replacement revision surgeries were performed because of clinical infections and five because of aseptic loosening. Preoperative and perioperative specimens were obtained from each patient and subjected to routine microbiological culture. The prostheses removed from each patient were subjected to mild ultrasonication to dislodge adherent bacteria, followed by aerobic and anaerobic microbiological culture. Bacterial DNA was extracted from each sonicate and the 16S rRNA gene was amplified with the universal primer pair 27f/1387r. All 10 specimens were positive for the presence of bacteria by both culture and PCR. PCR products were then cloned, organised into groups by RFLP analysis and one clone from each group was sequenced. Bacteria were identified by comparison of the 16S rRNA gene sequences obtained with those deposited in public access sequence databases. A total of 512 clones were analysed by RFLP analysis, of which 118 were sequenced. Culture methods identified species from the genera Leifsonia (54.3%), Staphylococcus (21.7%), Proteus (8.7%), Brevundimonas (6.5%), Salibacillus (4.3%), Methylobacterium (2.2%) and Zimmermannella (2.2%). Molecular detection methods identified a more diverse microflora. The predominant genus detected was Lysobacter, representing 312 (60.9%) of 512 clones analysed. In all, 28 phylotypes were identified: Lysobacter enzymogenes was the most abundant phylotype (31.4%), followed by Lysobacter sp. C3 (28.3%), gamma proteobacterium N4-7 (6.6%), Methylobacterium SM4 (4.7%) and Staphylococcus epidermidis (4.7%); 36 clones (7.0%) represented uncultivable phylotypes. We conclude that a diverse range of bacterial species are found within biofilms on the surface of clinically infected and non-infected prosthetic hip joints removed during revision arthroplasties.

Leifsonia rubra sp. nov. and Leifsonia aurea sp. nov., psychrophiles from a pond in Antarctica.

Two unique psychrophilic strains (CMS 76rT and CMS 81yT) were isolated from a cyanobacterial mat sample from a pond in Wright Valley, McMurdo, Antarctica. Both isolates were assigned to the genus Leifsonia, since they were gram-positive, curved rods, non-motile, catalase-positive, contained DL-2,4-diaminobutyric acid, menaquinone MK-11, phosphatidylglycerol and diphosphatidylglycerol, had a high content of anteiso- and iso-branched fatty acids and had a DNA G + C content of 64-66 mol%. In addition, both isolates were related to the five reported species of Leifsonia at a level of about 95-96% 16S rDNA sequence similarity and differed from one another by 2.5%. Strains CMS 76rT and CMS 81yT also differed from one another in many other phenotypic characteristics and exhibited only 30% relatedness at the DNA-DNA level, thus indicating that they represent two different species. Furthermore, these two isolates also showed many distinct differences with respect to the reported species of Leifsonia in terms of their phenotypic characteristics, biochemical properties, chemotaxonomic features, sensitivity to various antibiotics and 16S rDNA similarity, clearly indicating that strains CMS 76rT (= MTCC 4210T = DSM 15304T = CIP 107783T) and CMS 81yT (= MTCC 4657T = DSM 15303T = CIP 107785T) represent the type strains of two novel species of Leifsonia, for which the names Leifsonia rubra sp. nov. and Leifsonia aurea sp. nov. are proposed.

Leifsonia poae gen. nov., sp. nov., isolated from nematode galls on Poa annua, and reclassification of 'Corynebacterium aquaticum' Leifson 1962 as Leifsonia aquatica (ex Leifson 1962) gen. nov., nom. rev., comb. nov. and Clavibacter xyli Davis et al. 1984 with two subspecies as Leifsonia xyli (Davis et al. 1984) gen. nov., comb. nov.

The new genus Leifsonia gen. nov. with two new species, Leifsonia poae sp. nov. (type strain VKM Ac-1401T) and Leifsonia aquatica (ex Leifson 1962) nom. rev., comb. nov. (the type species, with VKM Ac-1400T = DSM 20146T = JCM 1368T as type strain), is proposed to accommodate bacteria found in Poa annua root gall, induced by the nematode Subanguina radicicola, and 'Corynebacterium aquaticum' Leifson 1962. Further, it is proposed to reclassify Clavibacter xyli Davis et al. 1984 with two subspecies in the new genus as Leifsonia xyli (Davis et al. 1984) comb. nov., Leifsonia xyli subsp. xyli (Davis et al. 1984) comb. nov. and Leifsonia xyli subsp. cynodontis (Davis et al. 1984) comb. nov. Members of the proposed genus are characterized by coryneform morphology, peptidoglycans based upon 2,4-diaminobutyric acid, the major menaquinone MK-11, phosphatidylglycerol and diphosphatidylglycerol as principal phospholipids, the high content of anteiso- and iso-branched saturated fatty acids, and a DNA G+C base composition of 66-73 mol%. They form a distinct phylogenetic branch attached to the line of descent of Agromyces spp. The new and reclassified species of the new genus clearly differ from each other phylogenetically and phenetically and can be recognized by their morphologies, the cell wall sugar composition, the requirement of complex media for growth, and numerous physiological characteristics, including the oxidase reaction.

Leifsonia gen. nov., a genus for 2,4-diaminobutyric acid-containing actinomycetes to accommodate "Corynebacterium aquaticum" Leifson 1962 and Clavibacter xyli subsp. cynodontis Davis et al. 1984.

"Corynebacterium aquaticum" was first proposed by Leifson in 1962 but not included in the approved lists of bacterial names in 1980. This species has been left from reclassification of the genus Corynebacterium because of the unusual chemotaxonomic characteristics such as 2,4-diaminobutyric acid (DAB) in the peptidoglycan and menaquinones of MK-10 and MK-11. A close relationship of "C. aquaticum" to the genera Agromyces and Rathayibacter has been pointed out from the viewpoint of chemotaxonomic profiles and phylogeny based on the 16S rDNA sequences. An analysis of DAB isomers of the peptidoglycan distinguished "C. aquaticum" clearly from these genera by possessing both L-DAB and D-DAB. We also found that the type strain of Clavibacter xyli subsp. cynodontis and two strains of amine-decomposing bacteria showed the similar chemotaxonomic features and formed a cluster with "C. aquaticum" in the phylogenetic tree based on 16S rDNA sequences in the family Microbacteriaceae. Considering these results, we propose a new genus Leifsonia to accommodate the four strains. The four species, Leifsonia aquatica sp. nov., nom. rev., comb. nov. (type species, type strain=JCM 1368), Leifsonia shinshuensis sp. nov. (type strain=DB102=JCM 10591), Leifsonia naganoensis sp. nov. (type strain=DB103=JCM 10592), and Leifsonia cynodontis comb. nov. (type strain=JCM 9733=ICMP 8790), were proposed here for the strains.