Whole-Genome Sequencing of Pseudomonas koreensis Isolated from Diseased Tor tambroides.

Unlike environmental P. koreensis isolated from soil, which has been studied extensively for its role in promoting plant growth, pathogenic P. koreensis isolated from fish has been rarely reported. Therefore, we investigated and isolated the possible pathogen that is responsible for the diseased state of Tor tambroides. Herein, we reported the morphological and biochemical characteristics, as well as whole-genome sequences of a newly identified P. koreensis strain. We assembled a high-quality draft genome of P. koreensis CM-01 with a contig N50 value of 233,601 bp and 99.5% BUSCO completeness. The genome assembly of P. koreensis CM-01 is consists of 6,171,880 bp with a G+C content of 60.5%. Annotation of the genome identified 5538 protein-coding genes, 3 rRNA genes, 54 tRNAs, and no plasmids were found. Besides these, 39 interspersed repeat and 141 tandem repeat sequences, 6 prophages, 51 genomic islands, 94 insertion sequences, 4 clustered regularly interspaced short palindromic repeats, 5 antibiotic-resistant genes, and 150 virulence genes were also predicted in the P. koreensis CM-01 genome. Culture-based approach showed that CM-01 strain exhibited resistance against ampicillin, aztreonam, clindamycin, and cefoxitin with a calculated multiple antibiotic resistance (MAR) index value of 0.4. In addition, the assembled CM-01 genome was successfully annotated against the Cluster of Orthologous Groups of proteins database, Gene Ontology database, and Kyoto Encyclopedia of Genes and Genome pathway database. A comparative analysis of CM-01 with three representative strains of P. koreensis revealed that 92% of orthologous clusters were conserved among these four genomes, and only the CM-01 strain possesses unique elements related to pathogenicity and virulence. This study provides fundamental phenotypic and genomic information for the newly identified P. koreensis strain.

Assessment of zinc solubilization potential of zinc-resistant Pseudomonas oleovorans strain ZSB13 isolated from contaminated soil / Avaliação do potencial de solubilização do zinco da cepa ZSB13 de Pseudomonas oleovorans resistente ao zinco isolada de solo contaminado

Zinc is an essential micronutrient that is required for optimum plant growth. It is present in soil in insoluble forms. Bacterial solubilization of soil unavailable form of Zn into available form, is an emerging approach to alleviate the Zn deficiency for plants and human beings. Zinc solubilizing bacteria (ZSB) could be a substitute for chemical Zn fertilizer. The present study aimed to isolate and characterize bacterial species from the contaminated soil and evaluate their Zn solubilizing potential. Zn resistant bacteria were isolated and evaluated for their MIC against Zn. Among the 13 isolated bacterial strains ZSB13 showed maximum MIC value upto 30mM/L. The bacterial strain with the highest resistance against Zn was selected for further analysis. Molecular characterization of ZSB13 was performed by 16S rRNA gene amplification which confirmed it as Pseudomonas oleovorans. Zn solubilization was determined through plate assay and broth medium. Four insoluble salts (zinc oxide (ZnO), zinc carbonate (ZnCO3), zinc sulphite (ZnS) and zinc phosphate (Zn3(PO4)2) were used for solubilization assay. Our results shows 11 mm clear halo zone on agar plates amended with ZnO. Likewise, ZSB13 showed significant release of Zn in broth amended with ZnCO3 (17 and 16.8 ppm) and ZnO (18.2 ppm). Furthermore, Zn resistance genes czcD was also enriched in ZSB13. In our study, bacterial strain comprising Zn solubilization potential has been isolated that could be further used for the growth enhancement of crops.

O zinco é um micronutriente essencial necessário para o crescimento ideal das plantas. Ele está presente no solo em formas insolúveis. A solubilização bacteriana da forma indisponível de Zn no solo para a forma disponível é uma abordagem emergente para aliviar a deficiência de Zn em plantas e seres humanos. Bactérias solubilizadoras de zinco (ZSB) podem ser um substituto para fertilizantes químicos de Zn. O presente estudo teve como objetivo isolar e caracterizar espécies bacterianas de solo contaminado e avaliar seu potencial de solubilização de Zn. Bactérias resistentes ao Zn foram isoladas e avaliadas quanto ao seu MIC contra o Zn. Entre as 13 cepas bacterianas isoladas, ZSB13 apresentou valor máximo de MIC de até 30 mM/L. A cepa bacteriana com maior resistência ao Zn foi selecionada para análise posterior. A caracterização molecular de ZSB13 foi realizada por amplificação do gene 16S rRNA que o confirmou como Pseudomonas oleovorans. A solubilização do Zn foi determinada através de ensaio em placa e meio caldo. Quatro sais insolúveis (óxido de zinco (ZnO), carbonato de zinco (ZnCO3), sulfito de zinco (ZnS) e fosfato de zinco (Zn3 (PO4) 2) foram usados para o ensaio de solubilização. Nossos resultados mostram uma zona de halo clara de 11 mm em placas de ágar corrigidas com ZnO. Da mesma forma, ZSB13 mostrou liberação significativa de Zn em caldo alterado com ZnCO3 (17 e 16,8 ppm) e ZnO (18,2 ppm). Além disso, os genes de resistência ao Zn czcD também foram enriquecidos em ZSB13. Em nosso estudo, a cepa bacteriana compreendendo potencial de solubilização de Zn foi isolada e poderia ser usada posteriormente para o aumento do crescimento de safras.

Community Structure of Nitrifying and Denitrifying Bacteria from Effluents Discharged into Lake Victoria, Kenya.

An active microbial community of nitrifying and denitrifying bacteria is needed for efficient utilization of nitrogenous compounds from wastewater. In this study, we explored the bacterial community diversity and structure within rivers, treated and untreated wastewater treatment plants (WWTPs) discharging into Lake Victoria. Water samples were collected from rivers and WWTPs that drain into Lake Victoria. Physicochemical analysis was done to determine the level of nutrients or pollutant loading in the samples. Total community DNA was extracted, followed by Illumina high throughput sequencing to determine the total microbial community and abundance. Enrichment and isolation were then done to recover potential nitrifiers and denitrifiers. Physicochemical analysis pointed to high levels total nitrogen and ammonia in both treated and untreated WWTPs as compared to the samples from the lake and rivers. A total of 1,763 operational taxonomic units (OTUs) spread across 26 bacterial phyla were observed with the most dominant phylum being Proteobacteria. We observed a decreasing trend in diversity from the lake, rivers to WWTPs. The genus Planktothrix constituted 19% of the sequence reads in sample J2 collected from the lagoon. All the isolates recovered in this study were affiliated to three genera: Pseudomonas, Klebsiella and Enterobacter in the phylum Proteobacteria. A combination of metagenomic analysis and a culture-dependent approach helped us understand the relative abundance as well as potential nitrifiers and denitrifiers present in different samples. The recovered isolates could be used for in situ removal of nitrogenous compounds from contaminated wastewater.

Specific Detection of Pseudomonas savastanoi Pathovars: From End-Point PCR to Real-Time PCR and HRMA.

Pseudomonas savastanoi is a phytopathogenic bacterium causing severe disease on olive, oleander, ash, and other Oleaceae. Three main pathovars belong to this species: P. savastanoi pv. savastanoi, pv. nerii, and pv. fraxini. Detection methods are mostly based on the visual inspection of the typical symptoms (i.e., knots and galls). However, this bacterium can survive on the host plant also as an epiphyte without giving any symptom. To avoid the spread of P. savastanoi to areas where it is absent, it is necessary to develop efficient and sensitive detection methods. Here, we reported three different PCR-based techniques, able to discriminate the three P. savastanoi pathovars attacking woody plants.

Pseudomonas keratitis complicated with spontaneous expulsive suprachoroidal hemorrhage: A case report.

INTRODUCTION: Spontaneous expulsive suprachoroidal hemorrhage (SESCH) is a rare condition. The correlation between SESCH and chronic glaucoma has been reported previously. However, few reports have indicated a correlation between infective keratitis and SESCHs. PATIENT CONCERNS: Here, we report the case of an 82-year-old woman with a corneal ulcer who presented with left eye pain for 6 days. DIAGNOSIS: We found that she has Pseudomonas keratitis and history of chronic glaucoma. INTERVENTIONS AND OUTCOMES: During admission, her left eye showed elevated intraocular pressure (IOP). Three days later, the eyeball began to bleed and became painful. She had high blood pressure on that day. Hours after complaints of eye pain, intraocular tissue exposure related to eyeball rupture, and SESCH. The patient underwent evisceration and insertion of a silicone ball for the socket reconstruction. Histopathological evaluation revealed acute inflammation of the cornea and the choroidal vessels. CONCLUSION: In elderly patients with infective keratitis and a history of glaucoma and hypertension, it is important to control intraocular pressure and blood pressure and pay attention to the risk of spontaneous expulsive suprachoroidal hemorrhage.

Pseudomonas canavaninivorans sp. nov., isolated from bean rhizosphere.

A novel canavanine-degrading bacterium, strain HB002T, was isolated from rhizosphere soil of a catch crop field collected from the island of Reichenau in Konstanz, Germany, and characterized by using polyphasic taxonomy. The facultative aerobe, rod-shaped, Gram-stain-negative bacterium was oxidase- and catalase-positive. The isolate was able to grow on canavanine as a sole carbon and nitrogen source. Results of phylogenetic analysis based on 16S rRNA gene sequences revealed highest similarities to Pseudomonas bijieensis (L22-9T, 99.93 %), Pseudomonas brassicacearum subsp. neoaurantiaca (ATCC 49054T, 99.76 %), Pseudomonas brassicacearum subsp. brassicacearum (DBK 11T, 99.63 %), Pseudomonas thivervalensis (DSM 13194T, 99.51 %), Pseudomonas kilonensis (DSM 13647T, 99.39 %) and Pseudomonas corrugata (ATCC29736T, 99.39 %). Marker gene analysis placed the strain in the intrageneric group of Pseudomonas fluorescens, subgroup P. corrugata. In silico DNA-DNA hybridization and average nucleotide identity values were both under the recommended thresholds for species delineation. The predominant fatty acids of strain HB002T were C16 : 0, C17 : 0 cyclo ω7c and C18 : 1 ω7c. The major respiratory quinone was Q9, followed by Q8 and minor components of Q7 and Q10. Results from the phenotypic characterization showd the strain's inability to hydrolyse gelatin and to assimilate N-acetyl glucosamide and a positive enzymatic activity of acid phosphatase and naphthol-AS-BI phosphohydrolase that distinguish this strain from closely related type strains. Taken together, these results show that strain HB002T represents a novel species in the genus Pseudomonas, for which the name Pseudomonas canavaninivorans sp. nov. is proposed. The type strain is HB002T (=DSM 112525T=LMG 32336T).

Multilocus sequence based identification and adaptational strategies of Pseudomonas sp. from the supraglacial site of Sikkim Himalaya.

Microorganisms inhabiting the supraglacial ice are biotechnologically significant as they are equipped with unique adaptive features in response to extreme environmental conditions of high ultraviolet radiations and frequent freeze-thaw. In the current study, we obtained eleven strains of Pseudomonas from the East Rathong supraglacial site in Sikkim Himalaya that showed taxonomic ambiguity in terms of species affiliation. Being one of the most complex and diverse genera, deciphering the correct taxonomy of Pseudomonas species has always been challenging. So, we conducted multilocus sequence analysis (MLSA) using five housekeeping genes, which concluded the taxonomic assignment of these strains to Pseudomonas antarctica. This was further supported by the lesser mean genetic distances with P. antarctica (0.73%) compared to P. fluorescens (3.65%), and highest ANI value of ~99 and dDDH value of 91.2 of the representative strains with P. antarctica PAMC 27494. We examined the multi-tolerance abilities of these eleven Pseudomonas strains. Indeed the studied strains displayed significant tolerance to freezing for 96 hours compared to the mesophilic control strain, while except for four strains, seven strains exhibited noteworthy tolerance to UV-C radiations. The genome-based findings revealed many cold and radiation resistance-associated genes that supported the physiological findings. Further, the bacterial strains produced two or more cold-active enzymes in plate-based assays. Owing to the polyadaptational attributes, the strains ERGC3:01 and ERGC3:05 could be most promising for bioprospection.

Biodegradation of petroleum by bacteria isolated from fishes of Indian Ocean / Biodegradação do petróleo por bactérias isoladas de peixes do oceano Índico

In this study, oil degrading bacteria discovered from fish living near the oil ports at Karachi in Pakistan were characterized. The bacteria isolated from skin, gills, and gut in fish could consume crude oil as a source of carbon and energy. Total 36 isolates were tested using Nutrient Agar (NA) and MSA media with different crude oil concentrations (0.2%, 0.5%, 0.7%, 1%, 2%, and 5%) and 4 out of 36 isolates (two Gram positive and two Gram negative bacteria) were selected for further identification. 16S rRNA gene sequencing revealed that the isolates are related to Bacillus velezensis, Bacillus flexus, Pseudomonas brenneri and Pseudomonas azotoforman. Oil degrading potential of these bacteria was characterized by GC-MS analysis of degradation of oil components in crude oil as well as engine oil. We found that one (2, 6, 10, 14-Tetramethylpentadecane) out of 42 components in the crude oil was fully eliminated and the other oil components were reduced. In addition, 26 out of 42 oil components in the engine oil, were fully eliminated and the rest were amended. Taken together, these studies identify that B. velezensis, B. flexus, P. brenneri and P. azotoforman have high oil degrading potential, which may be useful for degradation of oil pollutants and other commercial applications.

Neste estudo, bactérias degradadoras de óleo descobertas em peixes que vivem perto dos portos de petróleo em Karachi, no Paquistão, foram caracterizadas. As bactérias isoladas da pele, guelras e intestinos dos peixes podem consumir petróleo bruto como fonte de carbono e energia. No total, 36 isolados foram testados usando Agar Nutriente (NA) e meio MSA com diferentes concentrações de óleo bruto (0,2%, 0,5%, 0,7%, 1%, 2% e 5%) e 4 de 36 isolados (dois Gram positivos e duas bactérias Gram negativas) foram selecionadas para posterior identificação. O sequenciamento do gene 16S rRNA revelou que os isolados estão relacionados a Bacillus velezensis, Bacillus flexus, Pseudomonas brenneri e Pseudomonas azotoforman. O potencial de degradação do óleo dessas bactérias foi caracterizado pela análise de GC-MS da degradação dos componentes do óleo no óleo cru, bem como no óleo do motor. Descobrimos que um (2, 6, 10, 14-tetrametilpentadecano) de 42 componentes do óleo cru foi totalmente eliminado e os outros componentes do óleo foram reduzidos. Além disso, 26 dos 42 componentes do óleo do motor foram totalmente eliminados e o restante corrigido. Juntos, esses estudos identificam que B. velezensis, B. flexus, P. brenneri e P. azotoforman têm alto potencial de degradação de óleo, o que pode ser útil para a degradação de poluentes de óleo e outras aplicações comerciais.

Pseudomonas guryensis sp. nov. and Pseudomonas ullengensis sp. nov., isolated from soil.

Two Gram-staining-negative, aerobic, rod-shaped bacteria designated strains SR9T and UL070T, were isolated from soil and subjected to taxonomic characterization. Strain SR9T grew at 10-37 °C (optimum 30 °C), at pH 4.0-10.0 (optimum pH 8.0) and in the presence of 0-1 % NaCl (optimum 0 %), and UL070T at 4-33 °C (optimum 30 °C), at pH 4.0-10.0 (optimum pH 7.0) and in the presence of 0-2 % NaCl (optimum 0 %), respectively. Strain UL070T was motile with flagella. Analysis of 16S rRNA gene sequences indicated that the two strains fell into phylogenetic clusters belonging to the genus Pseudomonas. Both strains SR9T and UL070T were mostly related to Pseudomonas campi S1-A32-2T with 99.70 and 99.01% sequence similarities, and the similarity between the two isolates was 98.90 %. The genome-based in silico analyses indicated that each of the strains SR9T and UL070T was clearly separated from other species of Pseudomonas, as the orthologous average nucleotide identity (OrthoANI) and the digital DNA-DNA hybridization (dDDH) values were no higher than 93.09 and 50.03% respectively with any related species, which were clearly below the cutoff for species distinction. The fatty acid profiles of the two strains mainly consisting of unsaturated components, the presence of ubiquinone 9 (Q-9) as the major respiratory quinone, and phosphatidylethanolamine (PE) and diphosphatidylglycerol (DPG) as the diagnostic polar lipids were consistent with their classification into Pseudomonas. The DNA G+C contents of strains SR9T and UL070T were 63.2 mol% and 63.6 mol% respectively. On the basis of both phenotypic and phylogenetic evidences, each of the isolated strains should be classified as a novel species, for which the names Pseudomonas guryensis sp. nov. (type strain=SR9T=KCTC 82228T=JCM 34509T) and Pseudomonas ullengensis sp. nov. (type strain=UL070T=KCTC 82229T=JCM 34510T) are proposed.

Pseudomonas tohonis sp. nov., isolated from the skin of a patient with burn wounds in Japan.

Strain TUM18999T was isolated from the skin of a patient with burn wounds in Japan. The strain was successfully cultured at 20-42 °C (optimum, 30-35 °C) in 1.0-4.0% NaCl (w/v) and at pH 5.5-9.5, optimum pH 5.5-8.5. The phylogenetic tree reconstructed using 16S rRNA, gyrB, rpoB and rpoD gene sequences indicated that strain TUM18999T is closely related to Pseudomonas otitidis MCC10330T. Although the partial 16S rRNA gene sequence (1412 bp) of TUM18999T exhibits high similarity to those of Pseudomonas alcaligenes NBRC 14159T (99.08 %) and Pseudomonas otitidis MCC10330T (98.51 %), multi-locus sequence analysis using 16S rRNA, gyrB, rpoB and rpoD genes reveals a clear distinction between TUM18999T and other Pseudomonas species. In addition, an average nucleotide identity >90 % was not observed in the P. aeruginosa group. Moreover, TUM18999T and P. otitidis can be distinguished based on the minimum inhibitory concentration for carbapenem. Meanwhile, the cellular fatty acids are enriched with C18 : 1 ω7c/C18 : 1 ω6c (34.35 %), C16 : 1 ω7c/C16 : 1 ω6c (24.22 %), C16 : 0 (19.79 %) and C12 : 0 (8.25 %). Based on this evidence, strain TUM18999T can be defined as representing a novel Pseudomonas species, with the proposed name Pseudomonas tohonis sp. nov. The type strain is TUM18999T (GTC 22698T=NCTC 14580T).

Pseudomonas schmalbachii sp. nov., isolated from the gut of a millipede (Trigoniulus corallinus) from a coconut tree.

During an investigation of microbes associated with arthropods living in decaying coconut trees, a Pseudomonas isolate, Milli4T, was cultured from the digestive tract of the common Asian millipede, Trigoniulus corallinus. Sequence analysis of 16S rRNA and rpoB genes found that Milli4T was closely related but not identical to Pseudomonas panipatensis Esp-1T, Pseudomonas knackmussi B13T and Pseudomonas humi CCA1T. Whole genome sequencing suggested that this isolate represents a new species, with average nucleotide identity (OrthoANIu) values of around 83.9-87.7% with its closest relatives. Genome-to-genome distance calculations between Milli4T and its closest relatives also suggested they are distinct species. The genomic DNA G+C content of Milli4T was approximately 65.0 mol%. Phenotypic and chemotaxonomic characterization and fatty acid methyl ester analysis was performed on Milli4T and its related type strains. Based on these data, the new species Pseudomonas schmalbachii sp. nov. is proposed, and the type strain is Milli4T (=BCRC 81294T=JCM 34414T=CIP 111980T).

Multifarious effect of ACC deaminase and EPS producing Pseudomonas sp. and Serratia marcescens to augment drought stress tolerance and nutrient status of wheat.

Drought is the prime abiotic stress that rigorously influences plant growth, yield and quality of crops. The current investigation illustrated the bio-protective characters of Serratia marcescens and Pseudomonas sp. to ameliorate drought stress tolerance, plant growth and nutrient status of wheat. The present study aimed for search of potential drought tolerant plant growth-promoting rhizobacteria (PGPR). All screened bacterial isolates exhibited potential plant growth promoting (PGP) attributes such as production of ACC deaminase, exo-polysaccharide, siderophore, ammonia, IAA, and efficiently solubilized zinc and phosphate under in vitro conditions. To assess the in situ plant growth promotion potential of PGPR, a greenhouse experiment was conducted by priming wheat seeds with screened plant PGPR. Improved water status, reactive oxygen species, osmolyte accumulation, chlorophyll and carotenoids content in plant leaves confirmed the excellent drought tolerance conferring ability of RRN II 2 and RRC I 5. Among all PGPR, RRN II 2 and RRC I 5 inoculated plants not only demonstrated greater harvest index but also exhibited more micronutrient (zinc and iron) content in wheat grains. Further, RRN II 2 and RRC I 5 were identified through 16S rDNA sequencing as S. marcescens and Pseudomonas sp., respectively. Furthermore, amplification of acdS gene (Amplified band size of acdS gene was ~ 1.8 Kb) also confirmed ACC deaminase enzyme producing ability of Pseudomonas sp. Moreover, correlation coefficient, principal component analysis and cluster analysis also demonstrated that nutrient status and values of agronomical parameters of wheat primed with S. marcescens and Pseudomonas sp. were at par with the positive control. Thus, the outcome of this comparative investigation indicates that Pseudomonas sp. and S. marcescens could be utilized as bioinoculant in wheat since they can improve the physiological status, productivity and nutrient status in wheat crop under drought.

Pseudomonas capsici sp. nov., a plant-pathogenic bacterium isolated from pepper leaf in Georgia, USA.

Three phytopathogenic bacterial strains (Pc19-1T, Pc19-2 and Pc19-3) were isolated from seedlings displaying water-soaked, dark brown-to-black, necrotic lesions on pepper (Capsicum annuum) leaves in Georgia, USA. Upon isolation on King's medium B, light cream-coloured colonies were observed and a diffusible fluorescent pigment was visible under ultraviolet light. Analysis of their 16S rRNA gene sequences showed that they belonged to the genus Pseudomonas, with the highest similarity to Pseudomonas cichorii ATCC 10857T (99.7 %). The fatty acid analysis revealed that the majority of the fatty acids were summed feature 3 (C16  :  1 ω7c/C16  :  1 ω6c), C16  :  0 and summed feature 8 (C18  :  1 ω7c/C18  :  1 ω6c). Phylogenomic analyses based on whole genome sequences demonstrated that the pepper strains belonged to the Pseudomonas syringae complex with P. cichorii as their closest neighbour, and formed a separate monophyletic clade from other species. Between the pepper strains and P. cichorii, the average nucleotide identity values were 91.3 %. Furthermore, the digital DNA-DNA hybridization values of the pepper strains when compared to their closest relatives, including P. cichorii, were 45.2 % or less. In addition, biochemical and physiological features were examined in this study and the results indicate that the pepper strains represent a novel Pseudomonas species. Therefore, we propose a new species Pseudomonas capsici sp. nov., with Pc19-1T (=CFBP 8884T=LMG 32209T) as the type strain. The DNA G+C content of the strain Pc19-1T is 58.4 mol%.

The Impacts of Different Biological Treatments on the Transformation of Explosives Waste Contaminated Sludge.

The dinitrotoluene isomers 2,4 and 2,6-dinitrotoluene (DNT) represent highly toxic, mutagenic, and carcinogenic compounds used in explosive manufacturing and in commercial production of polyurethane foam. Bioremediation, the use of microbes to degrade residual DNT in industry wastewaters, represents a promising, low cost and environmentally friendly alternative technology to landfilling. In the present study, the effect of different bioremediation strategies on the degradation of DNT in a microcosm-based study was evaluated. Biostimulation of the indigenous microbial community with sulphur phosphate (2.3 g/kg sludge) enhanced DNT transformation (82% transformation, from 300 g/L at Day 0 to 55 g/L in week 6) compared to natural attenuation over the same period at 25 °C. The indigenous microbial activity was found to be capable of transforming the contaminant, with around 70% transformation of DNT occurring over the microcosm study. 16S rDNA sequence analysis revealed that while the original bacterial community was dominated by Gammaproteobacteria (30%), the addition of sulphur phosphate significantly increased the abundance of Betaproteobacteria by the end of the biostimulation treatment, with the bacterial community dominated by Burkholderia (46%) followed by Rhodanobacter, Acidovorax and Pseudomonas. In summary, the results suggest biostimulation as a treatment choice for the remediation of dinitrotoluenes and explosives waste.

Identification of pathogens isolated in acute external otitis cases and evaluation of aminoglycoside and quinolone sensitivity.

OBJECTIVE: This study aimed to identify pathogens isolated in acute external otitis cases and determine their distribution according to ages and seasons as well as investigate the susceptibility or resistance to the aminoglycoside and quinolone group antibiotics of which topical forms are available. METHOD: A total of 168 patients diagnosed with acute external otitis were evaluated retrospectively. Growing bacteria were identified according to the species by conventional methods. Antibiotic susceptibility status was determined for the growing bacteria. RESULTS: The most common bacteria detected were pseudomonas group bacteria (38.7 per cent). Resistance to the amikacin group of antibiotics was found to be the lowest and resistance to the ciprofloxacin group of antibiotics was the highest. CONCLUSION: External auditory canal cultures should be taken simultaneously with empirical treatment. Seasonal effect and age group should be taken into consideration in the choice of treatment and after questioning about chronic exposure to water. Empirical treatment should then be started.

Pseudomonas lactucae sp. nov., a pathogen causing bacterial rot of lettuce in Japan.

Two phytopathogenic bacteria, MAFF 301380T and MAFF 301381, isolated from rot lesions of lettuce (Lactuca sativa L. var. capitata L.) in Japan, were characterized using a polyphasic approach. The cells were Gram-reaction-negative, aerobic, non-spore-forming, rod-shaped and motile with one to three polar flagella. Analysis of the 16S rRNA gene sequences showed that the strains belong to the genus Pseudomonas and are closely related to Pseudomonas cedrina subsp. cedrina CFML 96-198T (99.72 %), Pseudomonas cedrina subsp. fulgida P515/12T (99.65 %), Pseudomonas gessardii DSM 17152T (99.51 %), Pseudomonas synxantha DSM 18928T (99.44 %), Pseudomonas libanensis CIP 105460T (99.44 %) and Pseudomonas lactis DSM 29167T (99.44 %). The genomic DNA G+C content was 60.4 mol% and the major fatty acids consisted of summed feature 3 (C16 : 1 ω7c/C16 : 1 ω6c), C16 : 0 and summed feature 8 (C18 : 1 ω7c/C18 : 1 ω6c). Phylogenetic analysis using the rpoD gene sequences and phylogenomic analyses based on the whole genome sequences demonstrated that the strains are members of the Pseudomonas fluorescens subgroup but formed a monophyletic and robust clade separated from their closest relatives. The average nucleotide identity and digital DNA-DNA hybridization values between the strains and their closely related species were 88.65 % or less and 36.3 % or less, respectively. The strains could be distinguished from their closest relatives by phenotypic characteristics, pathogenicity towards lettuce and whole-cell MALDI-TOF MS profiles. The evidence presented in this study supports the classification of the strains as representing a novel Pseudomonas species, for which we propose the name Pseudomonas lactucae sp. nov., with the type strain MAFF 301380T (=ICMP 23838T).

Pseudomonas arcuscaelestis sp. nov., isolated from rainbow trout and water.

Cells of strains P66T, V1 and W15Feb18 are Gram-stain-negative short rods and motile by one polar flagellum. Strain P66T was isolated from rainbow trout (Oncorhynchus mykiss) cultivated at a fish farm in Turkey. Strain V1 was isolated from sand of an intertidal shore on the Galicia coast in Spain and strain W15Feb18 was isolated from water collected at the Woluwe River in Belgium. Based on 16S rRNA sequence similarity values, the strains were grouped under the genus Pseudomonas and the Pseudomonas putida phylogenetic group of species. The DNA G+C content ranged from 58.5 to 58.9 mol%. The strains were characterized phenotypically by the API 20NE and Biolog GEN III tests, and chemotaxonomically by their whole-cell MALDI-TOF MS protein profiles and fatty acid contents. The absence of the hydrolysis of gelatin and the assimilation of arabinose, mannose and mannitol differentiated these strains from the closest species, Pseudomonas alkylphenolica. The major fatty acid components were C16:0 (29.91-31.68 %) and summed feature 3 (36.44-37.55 %). Multilocus sequence analysis with four and 83 housekeeping gene sequences and a core proteome analysis showed that these strains formed a phylogenetic cluster in the P. putida group of species. Genome comparisons by the average nucleotide identity based on blast and the Genome-to-Genome Distance Calculator demonstrated that the three strains belonged to the same genomic species and were distant from any known species, with similarity values lower than the thresholds established for species in the genus Pseudomonas. These data permitted us to conclude that strains P66T, V1 and W15Feb18 belong to a novel species in the genus Pseudomonas, for which the name Pseudomonas arcuscaelestis sp. nov. is proposed. The type strain is P66T (=CECT 30176T=CCUG 74872T). The other strains have been deposited in the CECT with the corresponding collection numbers: V1 (=CECT 30356) and W15Feb18 (=CECT 30355).

Pseudomonas paraversuta sp. nov. isolated from refrigerated dry-aged beef.

A polyphasic approach was applied to investigate the diversity of microbiota that evolved during cold storage beef ripening. Isolate V4/DAB/S4/2aT with a unique BOX-rep-PCR fingerprint profile revealed more than 99 % nucleotide identities upon pairwise comparisons of 16S rDNA sequences from the type strains Pseudomonas versuta DSM 101070T, Pseudomonas saxonica DSM 108989T, Pseudomonas deceptionensis DSM 26521T and Pseudomonas weihenstephanensis DSM 29166T, placing it within the Pseudomonas fragi / lundensis branch of the genus Pseudomonas. Additional rpoB based comparison revealed P. versuta DSM 101070T as the nearest relative, with 98.5 % nucleotide identity. Calculation of ANIb values of the V4/DAB/S4/2aT draft genome identified P. versuta DSM 101070T with 90.1 %, P. deceptionensis DSM 26521T with 85.1 %, P. fragi DSM 3456T with 84.4 %, Pseudomonas psychrophila DSM 17535T and Pseudomonas bubulae DSM 107389T with 84.2 % similarities each. Pairwise genome-to-genome distance calculations [digital DNA-DNA hybridization (dDDH)] resulted in values of 47.1, 35.1, 34.8, 34.2 and 34.1 %, respectively. A second isolate was detected years later in ground beef and showed ANIb values of 99.3 % and dDDH of 96.1 % relatedness to V4/DAB/S4/2aT. The DNA G+C content was 58.6 mol% for both isolates. The predominant cellular fatty acids of V4/DAB/S4/2aT were C16 : 0, C18 : 1ω7c, C17 : 0 cyclo and a summed feature containing C16 : 1ω7c and/or C15 : 0 iso 2-OH. The major polar lipids were phosphatidylethanolamine, phosphatidylglycerol and diphosphatidylglycerol, the major respiratory quinone was Q9, with a small portion of Q8. The combined data on genotypic and phenotypic features support the proposal of a novel species, for which the name Pseudomonas paraversuta sp. nov. is proposed. The type strain is V4/DAB/S4/2aT (=DSM 111361T=LMG 31844T) and a second isolate is UBT376 (=DSM 111360=LMG 31845).

Pseudomonas lalucatii sp. nov. isolated from Vallgornera, a karstic cave in Mallorca, Western Mediterranean.

Caves are extreme underground environments colonized by oligotrophic bacterial communities that influence mineral transformations. The identification at the species level is important and this study aims to the taxonomic characterisation of four bacterial strains previously isolated from rock surfaces and water samples from a karstic cave located on Mallorca (Spain) that were assigned to the genus Pseudomonas according to 16S rRNA nucleotide sequence analysis. Sequence analysis of the RNA polymerase sigma factor gene (rpoD) allocated these strains to the P. fluorescens lineage within the P. anguilliseptica phylogenetic group, close to the P. benzenivorans type strain. A polyphasic taxonomic approach included phenotypic characterization, fatty acid composition analysis, and whole-cell protein profiling, together with phylogenomic data. The results supported the proposal of a new species in the Pseudomonas genus. Characteristic fatty acid methyl esters of members of the Pseudomonas genus were present (C16:0, C10:0 3-OH, C12:0 2-OH and C12:0 3-OH) and the C12:1 3OH content differentiated these strains from P. benzenivorans. The genomic G + C mol% content of the four sequenced genomes was 66.9%. The average nucleotide indices based on BLAST analysis and the calculation of genome-to-genome distance with respect to their closest relative were lower than 88% and 30%, respectively. These data confirm that the four isolates, R1b-4, R1b-52A, A2bC-1 and R1b-54T, represent a new species, for which the name Pseudomonas lalucatii is proposed, with strain R1b-54T as the type strain (=CCUG 74754T = CECT 30179T). This is the first species in the P. anguilliseptica group isolated from this extreme habitat.

Pseudomonas quercus sp. nov, associated with leaf spot disease of Quercus mongolica.

Two Gram-negative, aerobic, motile bacteria strains were isolated from leaf spot disease of Quercus mongolica. Strain hsmgli-8T has 99.86 % 16S rRNA gene sequence similarity to LY10J, and the highest 16S rRNA gene sequence similarity to Pseudomonas cerasi 58T (97.2 %), then Pseudomonas ficuserectae JCM 2400T (97.18 %), Pseudomonas meliae CFBP 3225T, Pseudomonas tremae CFBP 6111T and Pseudomonas congelans DSM 14939T (all 97.12 %), and less than 97.1 % similarity to other recognized species. In phylogenetic trees based on 16S rRNA gene and multilocus sequence data, the two novel strains form a separate branch, indicating that they do not belong to any Pseudomonas group and subgroup, and should belong to a novel species within the genus Pseudomonas. This assertion is also supported by the results of genome average nucleotide identity analysis. The major fatty acids are C16 : 0, C18 : 1 ω7c and/or C18 : 1 ω6c, C16 : 1 ω7c and/or C16 : 1 ω6c. Polar lipids include phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, aminolipid and seven uncharacterized phospholipids. The predominant respiratory quinone is Q-9. The DNA G+C content is 59.45-59.50 mol%. Based on these data, we propose that the two novel strains should be assigned as a novel species within the genus Pseudomonas. We propose that the novel strains be named Pseudomonas quercus sp. nov. The type strain is hsmgli-8T (=CFCC 15739T=LMG 31544T).