Phytochemical analysis and anti-infective potential of fungal endophytes isolated from Nigella sativa seeds.

Endophytic fungi, particularly from higher plants have proven to be a rich source of antimicrobial secondary metabolites. The purpose of this study is to examine the antimicrobial potential of three endophytic fungi Aspergillus sp. SA1, Aspergillus sp. SA2, and Aspergillus sp. SA3, cultivated from Nigella sativa seeds against Staphylococcus aureus (ATCC 9144), Escherichia coli (ATCC 25922), Pseudomonas aeruginosa (ATCC 27853), Klebsiella pneumoniae (ATCC 13883), MRSA (ATCC 33591), and human pathogen Candida albicans (ATCC 10231). Furthermore, the most active cultivated endophytic fungi were molecularly identified via internal transcribed spacer (ITS) sequencing. HR-ESIMS guided approach has been used successfully in chemical profiling of 26 known bioactive secondary metabolites (1-26), which belongs to different classes of natural compounds such as polyketides, benzenoids, quinones, alcohols, phenols or alkaloids. Finally, in-silico interactions within active site of fungal Cyp51 and bacterial DNA gyrase revealed possibility of being a hit-target for such metabolites as antimicrobials.

Anti-infective potential of the endophytic fungus Aspergillus sp. associated with Aptenia cordifolia root supported by metabolomics analysis and docking studies.

Endophytic fungi are known to be a rich source for anti-infective drugs. In this study, Aptenia cordifolia associated endophytic fungi were explored for the first time. Seven isolates were identified morphologically followed by screening of these fungi by plug diffusion assay which revealed their potential activity against Staphylococcus aureus (ATCC 9144), Bacillus cereus (ATCC 14579), Serratia marcescens (ATCC 14756), Fusarium oxysporum (ATCC 48112), and Aspergillus flavus (ATCC 22546). Additionally, the crude ethyl acetate extract of the most potent three isolates in plug diffusion assay showed that Aspergillus sp. ACEFR2 was the most potent as anti-infective in disc diffusion assay; Accordingly, Aspergillus sp. ACEFR2 was investigated using phylogenetic analysis and LC-HR-ESI-MS. The phylogenetic analysis placed the strain into the Aspergillus section Niger close related to few species including A. niger. Whereas the metabolomic profiling revealed the presence of diverse pool of metabolites. Furthermore, in silico molecular docking study was carried out to predict which compounds most likely responsible for the anti-infective activity.

Paenibacillus plantiphilus sp. nov. from the plant environment of Zea mays.

A Gram-strain positive, aerobic, endospore-forming bacterial strain (JJ-246T) was isolated from the rhizosphere of Zea mays. The 16S rRNA gene sequence similarity comparisons showed a most closely relationship to Paenibacillus oenotherae DT7-4T (98.4%) and Paenibacillus xanthinolyticus 11N27T (98.0%). The pairwise average nucleotide identity and digital DNA-DNA hybridisation values of the JJ-246T genome assembly against publicly available Paenibacillus type strain genomes were below 82% and 33%, respectively. The draft genome of JJ-246T shared many putative plant-beneficial functions contributing (PBFC) genes, related to plant root colonisation, oxidative stress protection, degradation of aromatic compounds, plant growth-promoting traits, disease resistance, drug and heavy metal resistance, and nutrient acquisition. The quinone system of strain JJ-246T, the polar lipid profile and the major fatty acids were congruent with those reported for members of the genus Paenibacillus. JJ-246T was shown to represent a novel species of the genus Paenibacillus, for which the name Paenibacillus plantiphilus sp. nov. is proposed, with JJ-246T (= LMG 32093T = CCM 9089T = CIP 111893T) as the type strain.

Paenibacillus auburnensis sp. nov. and Paenibacillus pseudetheri sp. nov., isolated from the rhizosphere of Zea mays.

Two Gram-stain-positive, aerobic, endospore-forming bacterial strains, isolated from the rhizosphere of Zea mays were studied for their detailed taxonomic allocation. Based on 16S rRNA gene sequence similarity comparisons, both strains JJ-7T and JJ-60T were shown to be members of the genus Paenibacillus. Strain JJ-7T was most closely related to the type strains of Paenibacillus tianjinensis (99.6 %) and P. typhae (98.7 %), and strain JJ-60T to Paenibacillus etheri (99.5 %). The 16S rRNA gene sequence similarities to all other Paenibacillus species were ≤98.4 %. Both strains JJ-7T and JJ-60T showed 97.6 % 16S rRNA gene sequence similarity between each other. Genomic comparisons showed that the average nucleotide identity and digital DNA-DNA hybridization values to next related type strain genomes were always <94 and <56 %, respectively. The polar lipid profiles of both strains contain a number of phospholipids including diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine, which is in accord with the genus Paenibacillus. The major quinone was MK-7 in both strains. Major fatty acids were iso- and anteiso-branched. Physiological and biochemical characteristics allowed a further phenotypic differentiation of strains JJ-7T and JJ-60T from the most closely related species. Thus, each strain represents a novel species of the genus Paenibacillus, for which the names Paenibacillus auburnensis sp. nov. and Paenibacillus pseudetheri sp. nov. are proposed, with JJ-7T (=CIP 111892T=DSM 111785T=LMG 32088T=CCM 9087T) and JJ-60T (=CIP 111894T=DSM 111787T=LMG 32090T=CCM 9086T) as the type strains, respectively.

Vagococcus luciliae sp. nov., isolated from the common green bottle fly Lucilia sericata.

The genus Vagococcus belongs to the family Enterococcaceae (order Lactobacillales) and is closely related to the genus Enterococcus. Currently, 19 species of the genus have been validly named. In this study, we isolated strain G314FT from the common green bottle fly Lucilia sericata collected in Germany. Sequencing of its almost-complete 16S rRNA gene revealed that the isolate belongs to the genus Vagococcus, being closely related to Vagococcus bubulae SS1994T with high sequence identity (99.50 %), followed by Vagococcus martis D7T301T (98.86 %), Vagococcus vulneris SS1995T (98.71 %), Vagococcus teuberi DSM 21459T (98.64 %), Vagococcus silagei 2B-2T (98.64 %) and Vagococcus penaei CD276T (98.64 %). Genome sequencing of strain G314FT was performed by a combination of Illumina and Oxford Nanopore technology, yielding a circular genome with a size of 2 139 468 bp and an 11 kb plasmid. Average nucleotide identity and digital DNA-DNA hybridization values were calculated between G314FT and its closest-related taxa, and found to be <91 % and <40 %, respectively, thus strongly supporting that strain G314FT represents a novel species of the genus Vagococcus. Phylogenetic and core protein-based phylogenomic trees revealed that G314FT was closely related to a group of three species, V. bubulae SS1994T, V. martis D7T301T and V. teuberi DSM 21459T. Comparatively, the genome of G314FT is the smallest in the group of the four related species, and the biochemical pathway comparison using BlastKOALA revealed that G314FT has lost some amino acid biosynthetic proteins; however, it has gained enzymes for carbohydrate metabolism. Phenotypically, G314FT was consistent with other species of the genus Vagococcus including a negative catalase reaction and non-motility. Using the polyphasic approach, our data supports that the isolate represents a new species, for which we propose the name Vagococcus luciliae G314FT (=DSM 112651T= CCM 9164T).

Description of two cultivated and two uncultivated new Salinibacter species, one named following the rules of the bacteriological code: Salinibacter grassmerensis sp. nov.; and three named following the rules of the SeqCode: Salinibacter pepae sp. nov., Salinibacter abyssi sp. nov., and Salinibacter pampae sp. nov.

Current -omics methods allow the collection of a large amount of information that helps in describing the microbial diversity in nature. Here, and as a result of a culturomic approach that rendered the collection of thousands of isolates from 5 different hypersaline sites (in Spain, USA and New Zealand), we obtained 21 strains that represent two new Salinibacter species. For these species we propose the names Salinibacter pepae sp. nov. and Salinibacter grassmerensis sp. nov. (showing average nucleotide identity (ANI) values < 95.09% and 87.08% with Sal. ruber M31T, respectively). Metabolomics revealed species-specific discriminative profiles. Sal. ruber strains were distinguished by a higher percentage of polyunsaturated fatty acids and specific N-functionalized fatty acids; and Sal. altiplanensis was distinguished by an increased number of glycosylated molecules. Based on sequence characteristics and inferred phenotype of metagenome-assembled genomes (MAGs), we describe two new members of the genus Salinibacter. These species dominated in different sites and always coexisted with Sal. ruber and Sal. pepae. Based on the MAGs from three Argentinian lakes in the Pampa region of Argentina and the MAG of the Romanian lake Fara Fund, we describe the species Salinibacter pampae sp. nov. and Salinibacter abyssi sp. nov. respectively (showing ANI values 90.94% and 91.48% with Sal. ruber M31T, respectively). Sal. grassmerensis sp. nov. name was formed according to the rules of the International Code for Nomenclature of Prokaryotes (ICNP), and Sal. pepae, Sal. pampae sp. nov. and Sal. abyssi sp. nov. are proposed following the rules of the newly published Code of Nomenclature of Prokaryotes Described from Sequence Data (SeqCode). This work constitutes an example on how classification under ICNP and SeqCode can coexist, and how the official naming a cultivated organism for which the deposit in public repositories is difficult finds an intermediate solution.

Aristophania vespae gen. nov., sp. nov., isolated from wasps, is related to Bombella and Oecophyllibacter, isolated from bees and ants.

Acetic acid bacteria (family Acetobacteraceae) are found in the gut of most insects. Two clades are currently recognized: Commensalibacter-Entomobacter and Bombella-Oecophyllibacter. The latter group is only found in hymenopteran insects and the described species have been isolated from bees and ants. In this study, two new strains DDB2-T1T (=KACC 21507T=LMG 31759T) and DM15PD (=CCM 9165=DSM 112731=KACC 22353=LMG 32454) were isolated from wasps collected in the Republic of Korea and Germany, respectively. Molecular and phenotypic analysis revealed that the strains are closely related, with 16S rRNA gene sequences showing 100 % identity and genomic average nucleotide identity (ANI) values ≥99 %. The closest related species based on type strain 16S rRNA gene sequences are Swingsia samuiensis, Acetobacter peroxydans, Bombella favorum and Bombella intestini (94.8-94.7% identity), whereas the closest related species based on type strain genome analysis are Saccharibacter floricola and Bombella intestini (ANI values of 68.8 and 68.2 %, respectively). The reconstruction of a phylogenomic tree based on 107 core proteins revealed that the branch leading to DDB2-T1T and DM15PD is localized between Oecophyllibacter and Saccharibacter-Bombella. Further genomic distance metrics such as ANI, percentage of conserved proteins and alignment fraction values were consistent with these strains belonging to a new genus. The key phenotypic characteristics were one MALDI-TOF-MS peak (m/z=4601.9±2.0) and the ability to produce acid from d-arabinose. Based on this polyphasic approach, including phylogenetics, phylogenomics, genome distance calculations, ecology and phenotypic characteristics, we propose to name the novel strains Aristophania vespae gen. nov., sp. nov., with the type strain DDB2-T1T (=KACC 21507T=LMG 31759T).

Genomic plasticity and adaptive capacity of the quaternary alkyl-ammonium compound and copper tolerant Acinetobacter bohemicus strain QAC-21b isolated from pig manure.

Here, we present the genomic characterization of an Acinetobacter bohemicus strain QAC-21b which was isolated in the presence of a quaternary alky-ammonium compound (QAAC) from manure of a conventional German pig farm. The genetic determinants for QAAC, heavy metal and antibiotic resistances are reported based of the whole genome shotgun sequence and physiological growth tests. A. bohemicus QAC-21b grew in a species typical manner well at environmental temperatures but not at 37 °C. The strain showed tolerance to QAACs and copper but was susceptible to antibiotics relevant for Acinetobacter treatments. The genome of QAC-21b contained several Acinetobacter typical QAAC and heavy metal transporting efflux pumps coding genes, but no key genes for acquired antimicrobial resistances. The high genomic content of transferable genetic elements indicates that this bacterium can be involved in the transmission of antimicrobial resistances, if it is released with manure as organic fertilizer on agricultural fields. The genetic content of the strain was compared to that of two other A. bohemicus strains, the type strain ANC 3994T, isolated from forest soil, and KCTC 42081, originally described as A. pakistanensis, a metal resistant strain isolated from a wastewater treatment pond. In contrast to the forest soil strain, both strains from anthropogenically impacted sources showed genetic features indicating their evolutionary adaptation to the anthropogenically impacted environments. Strain QAC-21b will be used as model strain to study the transmission of antimicrobial resistance to environmentally adapted Acinetobacter in agricultural environments receiving high content of pollutants with organic fertilizers from livestock husbandry.

The anti-infective potential of the endophytic fungi associated with Allium cepa supported by metabolomics analysis and docking studies.

Endophytic fungi are known to be a rich source of anti-infective drugs. In our study, Allium cepa was investigated for fungal diversity using different media to give 11 isolates which were identified morphologically. Out of the isolated fungal strains, Penicillium sp. (LCEF10) revealed potential anti-infective activity against the tested microbes (Fusarium solani ATTC 25922, Pseudomonas aeruginosa (ATTC 29231), Staphylococcus aureus ATTC 27853, Candida albicans ATTC 10231), besides, their MICs were measured by well diffusion method, therefore, it was subjected to molecular identification in addition to phylogenetic analysis. Moreover, the ITS sequence of strain LCEF10 showed a consistent assignment with the highest sequence similarity (99.81%) to Penicillium oxalicum NRRL 787. The crude ethyl acetate extract of Penicillium sp. LCEF10 was investigated for metabolomic analysis using LC-HR-ESI-MS. The metabolic profiling revealed the presence of polyketides, macrolides, phenolics and terpenoids. Furthermore, in silico molecular docking study was carried out to predict which compounds most likely responsible for the anti-infective activity.

High diversity of the emerging pathogen Acinetobacter baumannii and other Acinetobacter spp. in raw manure, biogas plants digestates, and rural and urban wastewater treatment plants with system specific antimicrobial resistance profiles.

Carbapenem-resistant Acinetobacter baumannii causing immense treatment problems in hospitals. There is still a knowledge gap on the abundance and stability of acquired resistances and the diversity of resistant Acinetobacter in the environment. The aim of the study was to investigate the diversity and antimicrobial resistances of Acinetobacter spp. released from livestock and human wastewater into the environment. Raw and digested manure of small scale on farm biogas plants as well as untreated and treated wastewater and sewage sludge of rural and urban wastewater treatment plants (WWTPs) were studied comparatively. A total of 132 Acinetobacter isolates were phylogenetically identified (16S rRNA gene and rpoB sequence analyses) and 14 different phylotypes were detected. Fiftytwo isolates represented A. baumannii which were cultured from raw and digested manure of different biogas plants, and most stages of the rural WWTP (no hospital wastewater receiving) and the two studied urban WWTPs receiving veterinarian and human hospital wastewater. Multi-locus sequence typing (Pasteur_MLST) identified 23 novel and 12 known STs of A. baumannii. Most novel STs (18/23) were cultured from livestock samples and the rural WWTP. A. baumannii isolates from livestock and the rural WWTP were susceptible to carbapenems, colistin, ciprofloxacin, ceftazidime, and piperacillin. In contrast, A. baumannii isolates from the two urban WWTPs showed clinical linkage with respect to MLST and were multi-drug resistant (MDR). The presence of viable A. baumannii in digested manure and sewage sludge confirmed the survival of the strict aerobic bacteria during anoxic conditions. The study indicated the spread of diverse Acinetobacter from anthropogenic sources into the environment with a strong linkage of clinial associated MDR A. baumannii strains to the inflow of hospital wastewater to WWTPs. A more frequent detection of Acinetobacter in sewage sludge than effluent waters indicated that particle-attachment of Acinetobacter must be considered by the risk assessment of these bacteria.

Metabolomic profiling and cytotoxic potential of three endophytic fungi of the genera Aspergillus, Penicillium and Fusarium isolated from Nigella sativa seeds assisted with docking studies.

The main aim of our study is to investigate the anticancer potential of our cultivated entophytic fungal strains from Nigella sativa seeds. The strains were identified by sequencing of the partial 18S rRNA gene and the internal transcribed spacer (ITS) region as Aspergillus sp. (SA4), Penicillium sp. (SA5), and Fusarium sp. (SA6). We carried out metabolic profiling for three fungal strains to investigate their metabolites diversity. Profiling of the different extracts revealed their richness in diverse metabolites and consequently fourteen compounds (1-14) were annotated. In addition, the obtained extracts were examined against three cell lines HepG2, MCF-7 and Caco-2 showed activity with IC50 values in the range of 1.95-39.7 µg/mL. Finally, molecular docking study was performed showing questinol as the lowest glide binding score value (-5.925 kcal/mol) among all identified compounds. Our results showed Nigella sativa-associated endophytes as a promising source for further studies to look for anticancer secondary metabolites.

Neobacillus rhizosphaerae sp. nov., isolated from the rhizosphere, and reclassification of Bacillus dielmonensis as Neobacillus dielmonensis comb. nov.

A Gram-stain-positive, facultative anaerobic endospore-forming bacterium, which originated from roots/rhizosphere of maize (Zea mays), was investigated for its taxonomic position. On the basis of 16S rRNA gene sequence similarities, strain JJ-3T was grouped together with Neobacillus species showing the highest similarities to Neobacillus bataviensis (98.8 %) and the three species Neobacillus dendrensis, Neobacillus soli and Neobacillus cucumis (all 98.6 %). The 16S rRNA gene sequence similarities to the sequences of the type strains of other Neobacillus species were lower than 98.5 %. The average nucleotide identity, average amino acid identity and digital DNA-DNA hybridization values between the JJ-3T genome assembly and those of the other Neobacillus type strains were <83, <85 and <27 %, respectively. Chemotaxonomic features supported the grouping of the strain to the genus Neobacillus, e.g. the major fatty acids were C15 : 0 anteiso and C15 : 0 iso, the polar lipid profile contained the major components diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine, and the major quinone was menaquinone MK-7. Physiological and biochemical test results were slightly different from those of the most closely related species. For this reason, JJ-3T represents a novel species of the genus Neobacillus, for which we propose the name Neobacillus rhizosphaerae sp. nov., with JJ-3T (= CIP 111895T=LMG 32087T=DSM 111784T=CCM 9084T) as the type strain. We also propose to reclassify Bacillus dielmonensis as Neobacillus dielmonensis comb. nov. based mainly on the results of phylogenomic and conserved signature indel analyses.

Sutcliffiella rhizosphaerae sp. nov. isolated from roots.

An aerobic, Gram-staining-positive, endospore-forming bacterium, isolated from the rhizosphere of roots of maize (Zea mays), was taxonomically studied. On the basis of 16S rRNA gene sequence similarity comparisons, strain JJ-125T clustered together with species of the genus Sutcliffiella and showed the highest similarities with Sutcliffiella zhanjiangensis (98.7 %). The 16S rRNA gene sequence similarities to the sequences of the type strains of other species of the genus Sutcliffiella were <98.4 %. The genome sequence of JJ-125T was 4 516 360 bp long and had a DNA G+C content of 37.3 %. A DNA-DNA hybridization with the type strain of S. zhanjiangensis DSM 23010T resulted in values of 42.3 and 43.9 % (reciprocal). The average nucleotide identity, average amino acid identity and digital DNA-DNA hybridization values between the JJ-125T genome assembly and those of the other type strains of species of the genus Sutcliffiella were <75%, <80 % and <21 %, respectively. Chemotaxonomic features supported the grouping of the strain with the genus Sutcliffiella, e.g. the major fatty acids included iso-C15 : 0, iso-C17 : 1 ω10c and iso-C17 : 0, the polar lipid profile contained the major components diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine, the only quinone was menaquinone MK-7 and the characteristic diamino acid was meso-diaminopimelic acid. Physiological and biochemical test results were also different from those of the most closely related species. As a consequence, JJ-125T represents a novel species of the genus Sutcliffiella, for which we propose the name Sutcliffiella rhizosphaerae sp. nov., with JJ-125T (= CIP 111883T = LMG 32156T = CCM 9046T) as the type strain.

Insights into the impact of manure on the environmental antibiotic residues and resistance pool.

The intensive use of antibiotics in the veterinary sector, linked to the application of manure-derived amendments in agriculture, translates into increased environmental levels of chemical residues, AR bacteria (ARB) and antibiotic resistance genes (ARG). The aim of this review was to evaluate the current evidence regarding the impact of animal farming and manure application on the antibiotic resistance pool in the environment. Several studies reported correlations between the prevalence of clinically relevant ARB and the amount and classes of antibiotics used in animal farming (high resistance rates being reported for medically important antibiotics such as penicillins, tetracyclines, sulfonamides and fluoroquinolones). However, the results are difficult to compare, due to the diversity of the used antimicrobials quantification techniques and to the different amounts and types of antibiotics, exhibiting various degradation times, given in animal feed in different countries. The soils fertilized with manure-derived products harbor a higher and chronic abundance of ARB, multiple ARG and an enriched associated mobilome, which is also sometimes seen in the crops grown on the amended soils. Different manure processing techniques have various efficiencies in the removal of antibiotic residues, ARB and ARGs, but there is only a small amount of data from commercial farms. The efficiency of sludge anaerobic digestion appears to be dependent on the microbial communities composition, the ARB/ARG and operating temperature (mesophilic vs. thermophilic conditions). Composting seems to reduce or eliminate most of antibiotics residues, enteric bacteria, ARB and different representative ARG in manure more rapidly and effectively than lagoon storage. Our review highlights that despite the body of research accumulated in the last years, there are still important knowledge gaps regarding the contribution of manure to the AMR emergence, accumulation, spread and risk of human exposure in countries with high clinical resistance rates. Land microbiome before and after manure application, efficiency of different manure treatment techniques in decreasing the AMR levels in the natural environments and along the food chain must be investigated in depth, covering different geographical regions and countries and using harmonized methodologies. The support of stakeholders is required for the development of specific best practices for prudent - cautious use of antibiotics on farm animals. The use of human reserve antibiotics in veterinary medicine and of unprescribed animal antimicrobials should be stopped and the use of antibiotics on farms must be limited. This integrated approach is needed to determine the optimal conditions for the removal of antibiotic residues, ARB and ARG, to formulate specific recommendations for livestock manure treatment, storage and handling procedures and to translate them into practical on-farm management decisions, to ultimately prevent exposure of human population.

Paenibacillus allorhizoplanae sp. nov. from the rhizoplane of a Zea mays root.

A Gram-positive staining, aerobic, endospore-forming bacterial strain, isolated from the rhizosphere of Zea mays was studied for its detailed taxonomic allocation. Based on the 16S rRNA gene sequence similarity comparisons, strain JJ-42 T was shown to be a member of the genus Paenibacillus, most closely related to the type strain of Paenibacillus pectinilyticus (98.8%). The 16S rRNA gene sequence similarity to all other Paenibacillus species was below 98.5%. The pairwise average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values of the JJ-42 T genome assembly against publicly available Paenibacillus type strain genomes were below 92% and 47%, respectively. The quinone system of strain JJ-42 T consisted exclusively of menaquinone MK-7. The polar lipid profile consisted of the major components diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, three aminophospholipids (APL), and one unidentified lipid. The major fatty acids were iso- and anteiso-branched with the major compound anteiso C15:0. Physiological and biochemical characteristics allowed a further phenotypic differentiation of strain JJ-42 T from the most closely related species. Thus, JJ-42 T represents a novel species of the genus Paenibacillus, for which the name Paenibacillus allorhizoplanae sp. nov. is proposed, with JJ-42 T (= LMG 32089 T = CCM 9085 T = DSM 111786 T = CIP 111891 T) as the type strain.

Pseudocitrobacter corydidari sp. nov., isolated from the Asian emerald cockroach Corydidarum magnifica.

A Gram-negative bacterial strain, G163CMT, was isolated from the gut of the Asian emerald cockroach Corydidarum magnifica. The 16S rRNA gene sequence (1416 bp) of strain G163CMT showed 99.22% similarity to Pseudocitrobacter faecalis CCM 8479T and Pseudocitrobacter vendiensis CPO20170097T. The average nucleotide identity, digital DNA-DNA hybridization and average amino acid identity values of strain G163CMT were 92.4, 48.8 and 95.7% to P. faecalis CCM 8479T, and 93.3, 52.4 and 95.7% to P. vendiensis CPO20170097T. This strongly supports the designation of G163CMT as representing a new species in the genus Pseudocitrobacter. Phylogenetic trees based on the alignment of 16S rRNA, multilocus sequence analysis of six single-copy genes (fusA, pyrG, leuS, rpoB, recN and mnmE) and 107 core protein sequences consistently showed G163CMT to be a member of the genus Pseudocitrobacter, closely related to P. vendiensis CPO20170097T. In contrast to P. faecalis CCM 8479T and P. vendiensis CPO20170097T, the genome of G163CMT did not encode for proteins conferring resistance to antibiotics. However, all three genomes encoded a similar number of virulence factors and specialized metabolite biosynthetic proteins. The major fatty acids of strain G163CMT were C16:0 (31.5 %), C18:1 ω7c (22.6 %), C17:0 cyclo (15.3 %) and C14:0 (6.5 %). Based on the polyphasic results, we conclude that strain G163CMT represents a novel species of the genus Pseudocitrobacter and we propose the name Pseudocitrobacter corydidari sp. nov. with the type strain G163CMT (=DSM 112648T=CCM 9160T).

Erysipelothrix anatis sp. nov., Erysipelothrix aquatica sp. nov. and Erysipelothrix urinaevulpis sp. nov., three novel species of the genus, and emended description of Erysipelothrix.

Seven genotypically distinct strains assigned to the genus Erysipelothrix were isolated in different laboratories from several animal sources. Strain D17_0559-3-2-1T and three further strains were isolated from samples of duck, pig and goose. The strains had >99 % 16S rRNA gene sequence similarity to each other and to strain VA92-K48T and two further strains isolated from samples of medical leech and a turtle. The closest related type strains to the seven strains were those of Erysipelothrix inopinata (96.74 %) and Erysipelothrix rhusiopathiae (95.93 %). Average nucleotide identity, amino acid identity and in silico DNA-DNA hybridization results showed that the strains represented two separate novel species. One further phylogenetically distinct strain (165301687T) was isolated from fox urine. The strain had highest 16S rRNA gene sequence similarity to the type strains of Erysipelothrix tonsillarum (95.67 %), followed by Erysipelothrix piscisicarius (95.58 %) and Erysipelothrix larvae (94.22 %) and represented a further novel species. Chemotaxonomic and physiological data of the novel strains were assessed, but failed to unequivocally differentiate the novel species from existing members of the genus. MALDI-TOF MS data proved the discrimination of at least strain 165301687T from all currently described species. Based on the presented phylogenomic and physiological data, we propose three novel species, Erysipelothrix anatis sp. nov. with strain D17_0559-3-2-1T (=DSM 111258T= CIP 111884T=CCM 9044T) as type strain, Erysipelothrix aquatica sp. nov. with strain VA92-K48T (=DSM 106012T=LMG 30351T=CIP 111492T) as type strain and Erysipelothrix urinaevulpis sp. nov. with strain 165301687T (=DSM 106013T= LMG 30352T= CIP 111494T) as type strain.

Corallococcus soli sp. Nov., a Soil Myxobacterium Isolated from Subtropical Climate, Chalus County, Iran, and Its Potential to Produce Secondary Metabolites.

A novel myxobacterial strain ZKHCc1 1396T was isolated in 2017 from a soil sample collected along Chalus Road connecting Tehran and Mazandaran, Iran. It was a Gram-negative, rod-shaped bacterial strain that displayed the general features of Corallococcus, including gliding and fruiting body formation on agar and microbial lytic activity. Strain ZKHCc1 1396T was characterized as an aerobic, mesophilic, and chemoheterotrophic bacterium resistant to many antibiotics. The major cellular fatty acids were branched-chain iso-C17:0 2-OH, iso-C15:0, iso-C17:1, and iso-C17:0. The strain showed the highest 16S rRNA gene sequence similarity to Corallococcusterminator CA054AT (99.67%) and C. praedator CA031BT (99.17%), and formed a novel branch both in the 16S rRNA gene sequence and phylogenomic tree. The genome size was 9,437,609 bp, with a DNA G + C content of 69.8 mol%. The strain had an average nucleotide identity (ANI) value lower than the species cut-off (95%), and with the digital DNA-DNA hybridization (dDDH) below the 70% threshold compared to the closest type strains. Secondary metabolite and biosynthetic gene cluster analyses revealed the strain's potential to produce novel compounds. Based on polyphasic taxonomic characterization, we propose that strain ZKHCc1 1396T represents a novel species, Corallococcus soli sp. nov. (NCCB 100659T = CIP 111634T).

Bacillus rhizoplanae sp. nov. from maize roots.

A Gram-stain-positive, aerobic and endospore-forming bacterial strain, isolated from the root surface of maize (Zea mays) was taxonomically studied. It could be clearly shown that, based on 16S rRNA gene sequence similarity comparisons, strain JJ-63T is a member of the genus Bacillus, most closely related to the type strain of Bacillus pseudomycoides (98.61%), followed by Bacillus cereus (98.47 %). Detailed phylogenetic analysis based on the 16S rRNA gene and the 87 proteins conserved within the phylum Firmicutes placed the strain into the Cereus clade. The average nucleotide identity, average amino acid identity and digital DNA-DNA hybridization values against the type strain of B. pseudomycoides were 80.97, 81.45 and 26.30 %, respectively. The quinone system of strain JJ-63T consisted exclusively of menaquinone MK-7. The polar lipid profile consisted of the major components diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and an unidentified glycolipid. Major fatty acids were iso- and anteiso-branched with the major compounds iso-C15 : 0 and iso-C17 : 0. Also, the characteristic compounds C13 : 0 iso and C16 : 1 cis10 were found. Physiological and biochemical characteristics allowed a further phenotypic differentiation of strain JJ-63T from the most closely related species. For this reason, JJ-63T represents a novel species of the genus Bacillus, for which the name Bacillus rhizoplanae sp. nov. is proposed, with JJ-63T (=LMG 32091T=CCM 9090T=DSM 111827T= CIP 111899T) as the type strain.

Rhodococcus pseudokoreensis sp. nov. isolated from the rhizosphere of young M26 apple rootstocks.

The Gram-positive strain R79T, isolated from the rhizosphere of young M26 apple rootstocks, was investigated by a polyphasic taxonomic approach. Phylogenetic identification based on the full-length 16S rRNA gene sequence revealed highest 16S rRNA gene sequence similarity to the type strains of Rhodococcus wratislaviensis (99.6%) and Rhodococcus opacus (99.2%) followed by Rhodococcus imtechensis (98.9%). All other 16S rRNA gene sequence similarities were below 98.65%. A phylogenomic tree calculated based on a whole-genome sequence also showed a distinct clustering with the type strain of Rhodococcus koreensis. Average nucleotide identity (ANI) values between whole-genome sequences of R79T and the closest related type strains were below 95% supported the novel species status. The DNA G + C content of R79T was 67.24% mol. Predominant fatty acids were C16:0, C15:0 and C17:1ω8c. The strain contained MK8-H2 as the major respiratory quinone. The polar lipid profile consists of diphosphatidylglycerol and phosphatidylethanolamine, as well as of some unidentified lipids. The peptidoglycan type of the strain is A1γ meso-diaminopimelic acid. Based on the obtained genotypic and phenotypic, including chemotaxonomic data, we conclude that R79T represents a novel species of the genus Rhodococcus, for which the name Rhodococcus pseudokoreensis sp. nov. is proposed. The type strain is R79T (= DSM 113102T = LMG 32444T = CCM 9183T).