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The misuse of antibiotics and the emergence of antimicrobial resistance (AMR) is a concern in the aquaculture industry because it contributes to global health risks and impacts the environment. This study analyzed the AMR of sentinel bacteria associated with striped catfish ( Pangasisanodon hypophthalmus ) and giant snakehead ( Channa micropeltes ), the two main fish species reared in the pond culture in Cambodia. Phenotypic and genotypic characterization of the recovered isolates from fish, water, and sediment samples revealed the presence of bacteria, such as 22 species belonging to families Aeromonadaceae , Enterobacteriaceae , and Pseudomonadaceae . Among 48 isolates, Aeromonas caviae (n = 2), Aeromonas hydrophila (n = 2), Aeromonas ichthiosmia (n = 1), Aeromonas salmonicida (n = 4) were detected. A. salmonicida and A. hydrophilla are known as fish pathogens that occur worldwide in both fresh and marine water aquaculture. Antibiotic susceptibility testing revealed antibiotic resistance patterns of 24 (50 %) isolates among 48 isolates with higher multiple antibiotic resistance index (> 0.2). All the isolates of Enterobacteriaceae were susceptible to ciprofloxacin. Ciprofloxacin is a frontline antibiotic that is not recommended to use in aquaculture. Therefore, its use has to be strictly controlled. This study expands our knowledge of the AMR status in aquaculture farms which is very limited in Cambodia., Competing Interests: The authors declare there is no conflict., (© 2024 The Authors This is an Open Access article distributed under the terms of the Creative Commons Attribution Licence (CC BY 4.0), which permits copying, adaptation and redistribution, provided the original work is properly cited (http://creativecommons.org/licenses/by/4.0/).)

The phyllosphere microbiome plays an important role in plant fitness. Recently, bacteriophages have been shown to play a role in shaping the bacterial community composition of the phyllosphere. However, no studies on the diversity and abundance of phyllosphere bacteriophage communities have been carried out until now. In this study, we extracted, sequenced, and characterized the dsDNA and ssDNA viral community from a phyllosphere for the first time. We sampled leaves from winter wheat ( Triticum aestivum ), where we identified a total of 876 virus operational taxonomic units (vOTUs), mostly predicted to be bacteriophages with a lytic lifestyle. Remarkably, 848 of these vOTUs corresponded to new viral species, and we estimated a minimum of 2.0 × 10 6 viral particles per leaf. These results suggest that the wheat phyllosphere harbors a large and active community of novel bacterial viruses. Phylloviruses have potential applications as biocontrol agents against phytopathogenic bacteria or as microbiome modulators to increase plant growth-promoting bacteria.

A group of 11 bacterial strains was isolated from streams and lakes located in a deglaciated northern part of James Ross Island, Antarctica. They were rod-shaped, Gram-stain-negative, motile, and catalase-positive and produced blue-violet-pigmented colonies on R2A agar. A polyphasic taxonomic approach based on 16S rRNA gene sequencing, whole-genome sequencing, automated ribotyping, repetitive element sequence-based PCR (rep-PCR), MALDI-TOF MS, fatty acid profile, chemotaxonomy analyses, and extensive biotyping was applied in order to clarify the taxonomic position of these isolates. Phylogenetic analysis based on the 16S rRNA gene indicated that all the isolates constituted a coherent group belonging to the genus Rugamonas . The closest relatives to the representative isolate P5900 T were Rugamonas rubra CCM 3730 T , Rugamonas rivuli FT103W T , and Rugamonas aquatica FT29W T , exhibiting 99.2%, 99.1%, and 98.6% 16S rRNA pairwise similarity, respectively. The average nucleotide identity and digital DNA-DNA hybridization values calculated from the whole-genome sequencing data clearly proved that P5900 T represents a distinct Rugamonas species. The G+C content of genomic DNAs was 66.1 mol%. The major components in fatty acid profiles were summed feature 3 (C 16:1 ω7c /C 16:1 ω6c ), C 16:0 , and C 12:0 . The cellular quinone content contained exclusively ubiquinone Q-8. The predominant polar lipids were diphosphatidylglycerol, phosphatidylglycerol, and phosphatidylethanolamine. The polyamine pattern was composed of putrescine, 2-hydroxputrescine, and spermidine. IMPORTANCE Our polyphasic approach provides a new understanding of the taxonomy of novel pigmented Rugamonas species isolated from freshwater samples in Antarctica. The isolates showed considerable extracellular bactericidal secretions. The antagonistic activity of studied isolates against selected pathogens was proved by this study and implied the importance of such compounds' production among aquatic bacteria. The psychrophilic and violacein-producing species Roseomonas violacea may play a role in the diverse consortium among pigmented bacteria in the Antarctic water environment. Based on all the obtained results, we propose a novel species for which the name Rugamonas violacea sp. nov. is suggested, with the type strain P5900 T (CCM 8940 T ; LMG 32105 T ). Isolates of R. violacea were obtained from different aquatic localities, and they represent the autochthonous part of the water microbiome in Antarctica.

Strain F2A T , isolated from the cricket Acheta domesticus , was subjected to a polyphasic taxonomic characterization. Cells of the strain were rod-shaped, Gram-stain-negative and catalase- and oxidase-positive. It did not assimilate any carbohydrates. The strain's 16S rRNA gene sequence showed highest similarity to Entomomonas moraniae QZS01 T (96.4 %). The next highest similarity values were found to representatives of related genera (<93 %). The genome size of strain F2A T was 3.2 Mbp and the G+C content was 36.4 mol%. Average nucleotide identity values based on blast and MUMmer and average amino acid identity values between strain F2A T and E. moraniae QZS01 T were 74.29/74.43, 83.88 and 74.70 %, respectively. The quinone system predominantly contained ubiquinone Q-8. In the polar lipid profile, diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and an unidentified phospholipid were detected. The polyamine pattern consisted of the major compounds putrescine and spermidine. Major fatty acids were C 18 : 1  ω7 c and C 16 : 0 and the hydroxyl acids were C 12 : 0 3-OH, C 14 : 0 2-OH and C 14 : 0 3-OH. The diagnostic diamino acid of the peptidoglycan was meso -diaminopimelic acid. Due to its association with the only species of the genus Entomomonas but its distinctness from E. moraniae we here propose the novel species Entomomonas asaccharolytica sp. nov. F2A T (=CCM 9136 T =LMG 32211 T ).

A Gram-stain-negative, aerobic, non-motile and rod-shaped bacterium, designated as IMCC34836 T , was isolated from a freshwater stream. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain IMCC34836 T was most closely related to Permianibacter aggregans HW001 T (of the family Pseudomonadaceae ) with 95.6 % sequence similarity and formed a robust clade with P. aggregans HW001 T . The draft genome sequence of strain IMCC34836 T was 4.4 Mbp in size with 59.1 mol% DNA G+C content. Average nucleotide identity and digital DNA-DNA hybridization values between strain IMCC34836 T and P. aggregans HW001 T were 71.2 and 22.0 %, respectively, indicating that the new strain represents a novel species. The strain contained iso-C 15 : 0 , summed feature 3 (C 16 : 1  ω 6 c and/or C 16 : 1  ω 7 c ) and summed feature 9 (iso-C 17 : 1  ω 9 c and/or C 16 : 1 10-methyl) as the major fatty acids and harboured phosphatidylethanolamine, two unidentified aminophospholipids and three unidentified lipids as major polar lipids. The isoprenoid quinone detected in the strain was ubiquinone-8. Based on the phylogenetic and phenotypic characteristics, strain IMCC34836 T is considered to represent a novel species of the genus Permianibacter , for which the name Permianibacter fluminis sp. nov. is proposed. The type strain is IMCC34836 T (=KACC 21755 T =NBRC 114416 T ).

Pseudomonas protegens is a rhizosphere pseudomonad with a high agronomical potential (entomopathogenic and beneficial to plants) and bio-catalytic activities, but no selective medium has been described for its isolation. We developed a semi-selective minimum agar medium for the specific isolation and growth of P. protegens. We searched for both (i) a carbon source allowing the growth of P. protegens but potentially inhibiting the growth of other pseudomonads and (ii) an antimicrobial agent suppressing other members of the bacterial rhizosphere community. The M9-PP-agar medium consists of M9 base agar with adipic acid as the only carbon source and Irgasan® as an anti-bacterial agent. We tested the selectivity and sensitivity of M9-PP-agar by measuring the growth of 68 bacterial strains from 36 different species on this medium. Ten of the species tested were able to grow on M9-PP-agar medium: four species from the Pseudomonadaceae (Pseudomonas aeruginosa, Pseudomonas protegens, Pseudomonas putida, Stenotrophomonas maltophilia) as well as Achromobacter xylosoxidans, Agrobacterium tumefaciens, Brevundimonas sp., Serratia liquefaciens, Serratia marcescens and Variovorax paradoxus. All colonies were white, except for those of P. protegens (12 strains), which were typically brown. We demonstrated the efficiency of the M9-PP agar medium for P. protegens isolation, by inoculating two soils with the reference strain P. protegens CHAO T and then reisolating them. We also developed a fitF-PCR test targeting a regulator gene of the insecticidal P. protegens fit locus, for the rapid molecular detection of P. protegens colonies. We, therefore, developed a highly specific process for the routine isolation of new P. protegens strains from the soil environment, based on the use of a semi-selective medium and the specific color of colonies., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The honey bee gut microbiota contains many bacterial lineages that are specific to this ecosystem. Apis cerana , raised across the Asian continent, is of great significance to the maintenance and development of ecology and agriculture in Asia. Here, we report the isolation and characterization of strain QZS01 T from the gut of Apis cerana from Pingwu County, Sichuan Province, PR China. The results of phylogenetic analysis based on 16S rRNA sequences showed that strain QZS01 T forms a monophyletic group together with clone sequences derived from variable insect hosts, and it shows 92% sequence similarity to its closest relative, Pseudomonas knackmussii . Strain QZS01 T possesses a reduced genome (3.3 Mbp; G+C content, 38.05 mol%) compared to all other Pseudomonas species, and the whole-genome based phylogenetic reconstruction showed that strain QZS01 T represents a novel genus within the family Pseudomonadaceae . Strain QZS01 T is a Gram-stain-negative facultative anaerobe. It grows on brain heart infusion agar and the energy sources utilized for growth are very limited. Based on the results of genotypic and phenotypic analyses, we propose a novel genus and species, Entomomonas moraniae gen. nov., sp. nov., with the type strain QZS01 T (=CGMCC 1.13498 T =KCTC 62495 T ).

A Dyella-like bacterium was previously isolated from the planthopper Hyalesthes obsoletus (Hemiptera). Based on its 16S rRNA gene sequence, strain DHo T was assigned to the family Rhodanobacteraceae with Dyella and Frateuria as its closest relatives. The closest 16S rRNA gene sequences were Frateuria aurantia DSM 6220 T (98.2 %), Dyella thiooxydans ATSB10 T (98 %), Dyella terrae JS14-6 T (97.8 %) and Dyella marensis CS5-B2 T (97.8 %). Strain DHo T is a Gram-negative, aerobic, motile, yellow-pigmented, rod-shaped bacterium. Strain DHo T cells grew well at 28-30 °C and at pH 6.5-7.5 on a nutrient agar plate. DNA-DNA hybridization showed that the relatedness between strain DHo T and D. jiangningensis strain SBZ3-12 T , and F. aurantia DSM 6220 T was 42.7 and 42.6 %, respectively. Ubiquinone Q-8 was the predominant respiratory quinone, and the major fatty acids (>10 %) were iso-C15 : 0, iso-C16 : 0 and iso-C17 : 0. In silico analysis based on phylogenetics and sequence identity at the nucleotide and protein levels suggests that Frateuria is the closest known relative of strain DHo T . Based on the phenotypic, chemotaxonomic and phylogenetic data, strain DHo T was designated as a novel species of the genus Frateuria, for which the name Frateuria defendens sp. nov. is proposed. The type strain is DHo T (=NCCB 100648 T ; =DLB T =DSM 106169 T ).

The identification of microorganisms is very important in different fields and alternative methods are necessary for a rapid and simple identification. The use of fatty acids for bacterial identification is gaining attention as phenotypic characteristics are reflective of the genotype and are more easily analyzed. In this work, gas chromatography-vacuum ultraviolet spectroscopy (GC-VUV) was used to determine bacteria fatty acid methyl esters (FAMEs), to identify and discriminate different environmental bacteria based on their fatty acid profile. Microorganisms were grown in agar and their fatty acids extracted, saponified, and esterified before analysis. Unique FAME profiles were obtained for each microorganism mainly composed of branched, cyclopropane, hydroxy, saturated, and unsaturated fatty acid methyl esters. S. maltophilia showed a higher diversity of fatty acids while Bacillus species showed higher complexity in terms of branched-chain FAMEs, with several iso and anteiso forms. 12 different bacteria genera and 15 species were successfully differentiated based on their fatty acid profiles after performing PCA and cluster analysis. Some difficult to differentiate species, such as Bacillus sp., which are genetically very similar, were differentiated with the developed method., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Migratory birds are considered as vectors of infectious diseases, owing to their potential for transmitting pathogens over large distances. The populations of barn swallow (Hirundo rustica) migrate from Southeast Asia to the Japanese mainland during spring and migrate back to Southeast Asia during autumn. This migratory population is estimated to comprise approximately hundreds to thousands of individuals per year. However, to date, not much is known about the gastrointestinal microbiota of the barn swallow. In this study, we characterized the fecal bacterial community in barn swallow. Using 16S rRNA gene metagenomic sequencing analysis, we examined the presence and composition of potentially pathogenic bacteria in the fecal samples, which were collected during spring season from Osaka. The number (±S.D.) of total bacteria was approximately 2.1(±3.4)×10 8 per gram of feces. In most samples, the bacterial community composition was dominated by families, such as Enterobacteriaceae, Pseudomonadaceae, Mycoplasmataceae, Enterococcaceae, Streptococcaceae, and Alcaligenaceae. However, no relationship was found between the bacterial community composition and geographical area in the fecal samples. Potentially pathogenic bacteria were detected at the rate of >0.1%, which included Pseudomonas spp., Escherichia/Shigella spp., Enterobacter spp., Yersinia spp., Mycoplasma spp., Enterococcus spp., Achromobacter spp., and Serratia spp. Our results suggested that barn swallow is instrumental in the transmission of these genera over large distances.

This study evaluated cutaneous microbial distribution, and microbial co-occurrence at different body sites and skin environments in Chinese women (39.6 ± 11.9 years, N = 100) during the winter season. Microbial distribution (Propionibacterium acnes, Staphylococcus aureus, Staphylococcus epidermidis, Lactobacillus, Pseudomonadaceae, and Malassezia furfur), association with biomarkers (antimicrobial peptides: LL-37, β-defensins [HBD-2, HBD-3]), and claudin-1) and skin biophysical parameters (transepidermal water loss, pH, skin scaliness and roughness, sebum and hydration levels) were also determined. Skin sites (glabella [GL], hand-back [HB], interdigital web-space [IS], antecubital fossa [AF], volar forearm [VF], back [BA]) were classified as normal, oily or dry based on two-step cluster analysis and exposed or unexposed (uncovered or covered by clothes, respectively) based on seasonal apparel. Pseudomonadaceae and Staphylococcus aureus had the highest and lowest detection rate respectively at all sites. Cluster analysis identified skin sites as 'normal' (HB, BA, AF, VF), 'dry' (IS) and 'oily' (GL). Bacterial alpha diversity was higher in exposed (HB, IS, and GL) compared with unexposed sites (BA, AF and VF). Co-occurrence of Staphylococcus aureus with any of the other five microorganisms was lower in dry and oily skin versus normal skin. Skin exposure, biophysical/barrier profile and biomarkers were found to be associated with bacterial distribution and co-occurrence.

Background: Altered composition of airway microbiota has been reported in subjects suffering from asthma but its relation to eosinophilic phenotype is unclear., Objective: To examine the relationship between sputum microbiota, asthma severity and inflammatory type in asthmatic subjects from Guangzhou, China., Methods: Induced sputum samples were obtained from 49 non-smoking asthma patients, 25 severe and 24 non-severe, and 15 healthy subjects. Total DNA was amplified using primers specific for the V3-V5 hypervariable region of bacterial 16s rRNA and sequenced using the 454 GS FLX sequencer. Sequences were assigned to bacterial taxa by comparing them with 16s rRNA sequences in the Ribosomal Database Project., Results: Sputum eosinophil counts were higher and FEV 1 (% predicted) was lower in severe compared to non-severe asthmatics. There were no significant differences in operational taxonomic unit (OTU) numbers at the phylum level and in diversity scores between non-severe asthmatics and severe asthmatics, and healthy subjects. At the family level, Porphyromonadaceae was most abundant in healthy subjects whereas Pseudomonadaceae and Enterobacteriaceae were higher in severe asthmatics compared to non-severe asthmatics (p < 0.05). Actinomycetaceae was particularly abundant in eosinophilic asthma patients compared to non-eosinophilic asthma (p = 0.011). Bacteroidaceae was positively correlated with FEV 1 in all subjects (r = 0.335, p < 0.01), whereas body mass index was negatively associated with the number of species observed (r = -0.3, p < 0.05). Principal component analysis confirmed the positive association of Actinomycetaceae and Enterobacteriaceae abundance with eosinophilic asthma., Conclusion: Patients with asthma have an altered airway microbiota, with specific bacteria associated with severe asthma and the eosinophilic inflammatory phenotype., (Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Acute Oak Decline is a syndrome within the Oak Decline complex in Britain. Profuse stem bleeding and larval galleries of the native buprestid, Agrilus biguttatus characterize the disease. A systematic study comparing healthy with diseased trees was undertaken. This work reports the result of isolations from healthy trees, diseased and non-symptomatic tissue within AOD affected trees, at five sites in England. Bacteria and fungi were identified using the DNA gyrase B gene, or ITS 1 sequencing. A significantly higher proportion of diseased tissues (82%) yielded more bacteria than either healthy (18%) or non-symptomatic tissue in diseased trees (33%). Overall bacterial community compositions varied at each site, but significant similarities were evident in diseased tissues at all sites. Enterobacteriaceae dominated in diseased trees whereas Pseudomonadaceae dominated healthy trees. Significant associations between diseased tissues and certain bacterial species occurred, implying that the cause of tissue necrosis was not due to random microbiota. Brenneria goodwinii and Gibbsiella quercinecans were key species consistently isolated from diseased tissue; Rahnella victoriana and an un-named Pseudomonas taxon were also frequently isolated from both healthy and diseased trees. Most fungi isolated were from the outer bark and had no significant association with tree health status. It was concluded that there was a shift in the cultivatable bacterial microbiome of diseased trees, with Enterobacteriaceae strongly represented in symptomatic but not healthy tissues. No single species dominated the isolations from diseased tissues and the tissue degradation in AOD is therefore likely to have a polymicrobial cause., (Crown Copyright © 2016. Published by Elsevier GmbH. All rights reserved.)

Eight Gram-stain-negative bacteria (B4199T, C6819, C6918, D2441, D3318, E1086, E1148 and E5571) were identified during a retrospective study of unidentified strains from a historical collection held in the Special Bacteriology Reference Laboratory at the Centers for Disease Control and Prevention. The strains were isolated from eight patients: five female, two male and one not specified. No ages were indicated for the patients. The sources were urine (3), leg tissue (2), foot wound, lung tissue and deep liver. The strains originated from seven different states across the USA [Colorado, Connecticut (2), Indiana, North Carolina, Oregon and Pennsylvania]. The strains grew at 10-42 °C, were non-motile, alkalitolerant, slightly halophilic, microaerophilic, and catalase- and oxidase-positive. The DNA G+C content was 47.3-47.6 mol%. The major cellular fatty acids were tetradecanoic acid (C14 : 0), hexadecanoic acid (C16 : 0) and 11-octadecenoic acid (C18 : 1ω7c). Polar lipids detected were phosphatidylglycerol, phosphatidylethanolamine, diphosphatidylglycerol and unknown phospholipids; the only respiratory quinone detected was the ubiquinone Q-9 (100 %). 16S rRNA gene sequence analysis produced results with 95.6 % similarity to Pseudomonas caeni DSM 24390T and 95.2 % similarity to Thiopseudomonas denitrificans X2T. The results of the biochemical, chemotaxonomic and phylogenetic analyses between the study strains and some related type strains indicated that these strains represent a novel species of a new genus within the family Pseudomonadaceae, for which the name Oblitimonas alkaliphila gen. nov., sp. nov. is proposed. The type strain is B4199T (=DSM 100830T=CCUG 67636T).

Removal of 3-nitro-1,2,4-triazol-5-one (NTO) was investigated in conjunction with heterotrophic and autotrophic denitrifying growth conditions by a microbial consortium from a wastewater treatment plant. Microcosms were supplemented with molasses, methanol, or thiosulfate. Cultures were passaged twice by transferring 10 % of the culture volume to fresh media on days 11 and 21. Rates of NTO removal were 18.71 ± 0.65, 9.04 ± 2.61, and 4.34 ± 2.72 mg/L/day while rates of nitrate removal were 20.08 ± 1.13, 21.58 ± 1.20, and 24.84 ± 1.26 mg/L/day, respectively, for molasses, methanol, or thiosulfate. Metagenomic analysis showed that Proteobacteria and Firmicutes were the major phyla in the microbial communities. In molasses supplemented cultures, the community profile at the family level changed over time with Pseudomonadaceae the most abundant (67.4 %) at day 11, Clostridiaceae (65.7 %) at day 21, and Sporolactobacillaceae (35.4 %) and Clostridiaceae (41.0 %) at day 29. Pseudomonadaceae was the dominant family in methanol and thiosulfate supplemented cultures from day 21 to 29 with 76.6 and 81.6 % relative abundance, respectively.

Three bacterial strains, designated DS48-6-5T, DS48-6-7 and DS48-6-9, were isolated from a sediment sample taken from Daechung Reservoir (Republic of Korea) at a water depth of 48 m. Cells of the strains were Gram-stain-negative, aerobic, rod-shaped and motile with a single polar flagellum. Comparative 16S rRNA gene sequence studies showed that the three isolates had clear affiliation with Betaproteobacteria and the closest relatives were Rhizobacter bergeniae KCTC 32299T, Rhizobacter dauci DSM 11587T and Rhizobacter fulvus KCTC 12591T with 97.2-97.9 % 16S rRNA gene sequence similarities; the 16S rRNA gene sequence similarities between the three strains were 99.5-100 %. The only isoprenoid quinone of the three strains was ubiquinone-8, and the major fatty acids were summed feature 3 (C16 : 1ω6c and/or C16 : 1ω7c) and C16 : 0. The G+C content of the genomic DNA of strains DS48-6-5T, DS48-6-7 and DS48-6-9 was 66.7, 67.0 and 66.8 mol%, respectively. DNA-DNA hybridization values of the novel strains with R. bergeniae KCTC 32299T, R. dauci DSM 11587T and R. fulvus KCTC 12591T were 19.3-48.5 %. Based on the evidence from this taxonomic study using a polyphasic approach, it is proposed that strains, DS48-6-5T, DS48-6-7 and DS48-6-9, represent a novel species of the genus Rhizobacter, for which the name Rhizobacter profundi sp. nov. is proposed. The type strain is DS48-6-5T ( = KCTC 42645T = NBRC 111169T).

A denitrifying bacteria consortium was enriched from LiHe River's sediment, the dynamics of total nitrogen (TN), nitrate (NO3- -N), nitrite (NO2- -N), ammonium (NH4+ -N) and COD at different enrichment cultivation stages were studied, and the total volume, the releasing rates and the composition of gas released during the denitrification process were analyzed. The full-length 16S rDNA clone library was constructed, enclosing the diversity of the denitrifying bacteria consortium. The results showed, in the enrichment phase 4, under the load of TN 330 mg x L(-1), the best nitrogen removal effect was obtained, which the TN and NO3- -N removal rates reached 90.9% and 100% within 9 hours, respectively. The accumulation amounts of NO2- -N and NH4+ -N were merely 3.39 mg x L(-1) and 16.64 mg x L(-1). And the COD removal rate was 85%. The process released 260 mL of the compound gas, in which the main ingredient was N2 associated with a small quantity of CH4 and CO2. The denitrifying bacteria consortium consisted of the family Pseudomonadaceae and the family Rhodocyclaceae, belonging to Proteobacteria phylum, in which the OUT abundances were 57.8% and 31.6%, respectively. The family Pseudomonadaceae was the predominant group.

A Gram-staining-negative, rod-shaped, motile and facultatively anaerobic bacterial strain, designated X2(T), was isolated from the sludge of an anaerobic, denitrifying, sulfide-removal bioreactor, and found to oxidize sulfide anaerobically with nitrate as electron acceptor. The strain grew at salinities of 0-3% (w/v) NaCl (optimum, 0-1%). Growth occurred at pH 6.0-10.0 (optimum, pH 8.0) and 10-37 °C (optimum, 30 °C). The genomic DNA G+C content was 59 mol%. Q-8 and Q-9 were detected as the respiratory quinones. The major fatty acids (>10 %) were C16:1ω7c and/or C16: 1ω6c, C18: 1ω7c and C16:0. The polar lipids consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol and one unidentified phospholipid. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain X2(T) formed a novel clade within the family Pseudomonadaceae, with the highest sequence similarity to Pseudomonas caeni KCTC 22292(T) (93.5%). On the basis of phenotypic, chemotaxonomic and phylogenetic characteristics, it is proposed that this strain represents novel genus and species within the family Pseudomonadaceae, for which the name Thiopseudomonas denitrificans gen. nov., sp. nov. is proposed. The type strain is X2(T) ( =CCTCC M 2013362(T) =DSM 28679(T) = KCTC 42076(T))., (© 2015 IUMS.)

This paper was the 1st research focusing on the design of a halophilic lactic starter for the production of fermented fish products using a quantitative approach, based on the evaluation of the growth index and acidification score, as well as on the use of a multivariate approach to select the most promising strains. Fifty-nine strains were randomly selected from salted fish and phenotypically characterized through Gram staining, catalase activity, glucose metabolism, H2 S and indole production, nitrate reduction, citrate utilization, and hydrolysis of arginine, esculin, casein, gelatin, starch, Tween 80, and urea. Then the Gram positive isolates (44 out of 59) were studied for their growth at different temperatures (10, 25, 40, and 55 °C), salt (0%, 20%, and 30%), pHs (4.5 and 9.5), and acidification score in lab medium. Data were modelled through growth index and used as input to run a preliminary cluster analysis and a principal component analysis. Three promising strains were selected, identified as members of the genus Pediococcus and used for the validation at laboratory level through the assessment of their performances in the production of a fermented fish sauce. The results were really promising as their use not only reduced the fermentation time (2 d) but also improved the microbiological quality of the final product. This paper represents a 1st report on the use of a simple step-by-step methodology to select promising halophilic strains for the optimization of a starter for fish-fermented products., (© 2014 Institute of Food Technologists®)

The purpose of this review is to discuss the scientific literature on waterborne healthcare-associated infections (HCAIs) published from 1990 to 2012. The review focuses on aquatic bacteria and describes both outbreaks and single cases in relation to patient characteristics, the settings and contaminated sources. An overview of diagnostic methods and environmental investigations is summarized in order to provide guidance for future case investigations. Lastly, on the basis of the prevention and control measures adopted, information and recommendations are given. A total of 125 reports were included, 41 describing hospitalized children. All cases were sustained by opportunistic pathogens, mainly Legionellaceae, Pseudomonadaceae and Burkholderiaceae. Hot-water distribution systems were the primary source of legionnaires' disease, bottled water was mainly colonized by Pseudomonaceae, and Burkholderiaceae were the leading cause of distilled and sterile water contamination. The intensive care unit was the most frequently involved setting, but patient characteristics were the main risk factor, independent of the ward. As it is difficult to avoid water contamination by microbes and disinfection treatments may be insufficient to control the risk of infection, a proactive preventive plan should be put in place. Nursing staff should pay special attention to children and immunosuppressed patients in terms of tap-water exposure and also their personal hygiene, and should regularly use sterile water for rinsing/cleaning devices., (© 2014 The Authors.)

A novel bacterial strain, capable of aggregating potential biofuel-producing microalgae, was isolated from the phycosphere of an algal culture and designated HW001(T). The novel bacterial strain was identified on the basis of its phylogenetic, genotypic, chemotaxonomic and phenotypic characteristics in this study. Cells were aerobic, Gram-negative rods. 16S rRNA gene-based phylogenetic analysis revealed that strain HW001(T) is affiliated with the family Pseudomonadaceae in the phylum Proteobacteria, but forms a distinct clade within this family. The DNA G+C content of strain HW001(T) was 55.4 mol%. The predominant cellular fatty acids were iso-C15:0, summed feature 9 (iso-C17:1ω9c), C16:0 and summed feature 3 (C16:1ω7c/C16:1ω6c). Q-8 was the main respiratory quinone. The polar lipid profile contained phosphatidylethanolamine, an unidentified aminophospholipid and some unidentified lipids. Based on the extensive polyphasic analysis, strain HW001(T) represents a novel species of a new genus in the family Pseudomonadaceae, for which the name Permianibacter aggregans gen. nov., sp. nov., is proposed. The type strain of the type species is HW001(T) ( = CICC 10856(T) = KCTC 32485(T))., (IUMS.)

The aim of this study was to isolate, quantify, identify, and compare opportunistic microorganisms (Candida and Staphylococcus genera and Enterobacteriaceae/Pseudomonadaceae families) from prosthesis-fitting surfaces, the hard palate, and mouth rinses of individuals wearing removable maxillary prosthesis with (50) and without (50) lesions of denture stomatitis (DS). The strains were collected and identified using phenotypic, biochemical and molecular tests. The counts of microorganisms were significantly higher in the group of individuals with DS (P < 0.05). C. albicans was the most frequently isolated yeast species in both groups, following by C. tropicalis and C. glabrata. Six isolates were identified as C. dubliniensis. S. aureus and S. epidermidis were the most frequent Staphylococcus species in both groups. Klebsiella pneumoniae was the predominant species in both groups. The association between Candida spp. and bacteria isolated in this study with DS suggests that these microorganisms may play important roles in the establishment and persistence of this disease., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Aims: To examine for psychrophilic or psychrotolerant micro-organisms in pharmaceutical cold rooms (in relation to numbers, incidents and species) and to determine, where such micro-organisms are present, whether standard microbiological environmental monitoring regimes require modification. This is presented as a case study., Methods and Results: Comparative environmental monitoring within different pharmaceutical facility cold rooms (using standard mesophilic and low temperature incubation). Data were collected over two periods, 5 years apart. The results indicated that psychrophilic micro-organisms were not present and that those micro-organisms deemed psychrotolerant, primarily pseudomonads, could be grown on standard media under mesophilic conditions., Conclusions: Psychrophilic micro-organisms were not detected and those considered to be psychrotolerant were only found in low numbers. Pyschrotolerant organisms were recovered under both low temperature incubation conditions and under standard conditions (between 20 and 35°C). Further evaluation may be required, using alternative agar, and microbiologists should regularly review the species recovered to note differences between different environments., Significance and Impact of the Study: The study came about from requests made by US and UK regulators concerning the risk of any extremophiles present in pharmaceutical manufacturing facilities upon product safety. Regulators expressed concerns about whether standard, and accepted, environmental monitoring regimes were capable of detecting such micro-organisms. The data provide a benchmark to support pharmaceutical manufacturers in relation to their existing monitoring programmes or as a case study with which to undertake a similar study., (© 2012 The Society for Applied Microbiology.)

A rubber-degrading bacterium, the strain NS21, that was isolated from a soil sample in a botanical garden in Japan (Imai et al., 2011) was examined by phenotypic, phylogenetic and chemotaxonomic approaches to determine its taxonomic position. The strain NS21 was motile, possessing a single polar flagellum and a facultatively anaerobic straight rod. Analysis of the 16S rRNA and gyrB gene sequences of NS21 revealed a close relationship to the genus Rhizobacter. The predominant quinone type was Q-8. The G+C content of the NS21 genomic DNA was 70.8 mol%. The major fatty acids were C16:0, C17:0 cyclo, C18:1ω7c and C16:1ω7c and/or iso-C15:0 2-OH. C12:0 2-OH was present. The DNA-DNA hybridization experiments indicated that the DNA relatedness values of the strain NS21 to R. dauci H6(T) and R. fulvus Gsoil 322(T) were lower than 24%. The phenotypic characteristics showed obvious dissimilarities when compared to closely related species. On the basis of these taxonomic properties, a novel species is proposed as Rhizobacter gummiphilus sp. nov., with the strain NS21(T) (NBRC 109400(T), BCC 58006(T)) as the type strain. The emended description of the genus Rhizobacter was also presented.

The epiphytic bacterial communities colonising roots and leaves have been described for many plant species. In contrast, microbiologists have rarely considered flowers of naturally growing plants. We identified bacteria isolated from the surface of petals and leaves of two plant species, Saponaria officinalis (Caryophyllaceae) and Lotus corniculatus (Fabaceae). The bacterial diversity was much lower on petals than on leaves of the same plants. Moreover, the bacterial communities differed strongly in composition: while Pseudomonadaceae and Microbacteriaceae were the most abundant families on leaves, Enterobacteriaceae dominated the floral communities. We hypothesise that antibacterial floral volatiles trigger the low diversity on petals, which is supported by agar diffusion assays using substances emitted by flowers and leaves of S. officinalis. These results suggest that bacteria should be included in the interpretation of floral traits, and possible effects of bacteria on pollination are proposed and discussed., (© 2011 German Botanical Society and The Royal Botanical Society of the Netherlands.)

Urmia Lake is one of the most permanent hypersaline lakes in the world which is threatened by hypersalinity and serious dryness. In spite of its importance no paper has been published regarding bacterial community of this lake. Accordingly, the present study aimed to investigate the halophilic bacteria in the aforementioned lake. In so doing, thirty seven strains were isolated on six different culture media. The isolated strains were characterized using phenotypic and genotypic methods. Growth of the strains occurred at 2535 degrees C, pH 6-9 and 7 to 20% (w/v) NaCl indicating that most of the isolates were moderately halophiles. Catalase, oxidase and urease activities were found to be positive for the majority of the isolates. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the isolated bacteria belonged to two major taxa: Gammaproteobacteria (92%, including Salicola [46%], Pseudomonas [13.5%], Marinobacter [ 11%], Idiomarina [11%], and Halomonas [8%]) and Firmicutes (8%, including Bacillus [5%] and Halobacillus [3%]). In addition, a novel bacterium whose 16S rRNA gene sequence showed almost 98% sequence identity with the taxonomically troubled DSM 3050T, Halovibrio denitrificans HGD 3T and Halospina denitrificans HGD 1-3T, each, was isolated. 16S rRNA gene similarity levels along with phenotypic characteristics suggest that some of the isolated strains could be regarded as potential type strain for novel species, on which further studies are recommended.

The aim was to evaluate the presence of Staphylococcus spp., Enterobacteriaceae and Pseudomonadaceae in the oral cavities of HIV-positive patients. Forty-five individuals diagnosed as HIV-positive by ELISA and Western-blot, and under anti-retroviral therapy for at least 1 year, were included in the study. The control group constituted 45 systemically healthy individuals matched to the HIV patients to gender, age and oral conditions. Oral rinses were collected and isolates were identified by API system. Counts of microorganisms from HIV and control groups were compared statistically by a Mann-Whitney test (α=5%). The percentages of individuals positive for staphylococci were similar between the groups (p=0.764), whereas for Gram-negative rods, a higher percentage was observed amongst HIV-positive (p=0.001). There was no difference in Staphylococcus counts between HIV and control groups (p=0.1008). Counts were lower in the oral cavities of patients with low viral load (p=0.021), and no difference was observed in relation to CD4 counts (p=0.929). Staphylococcus aureus was the most frequently isolated species in HIV group, and Staphylococcus epidermidis was the prevalent species in the control group. Significantly higher numbers of enteric bacteria and pseudomonas were detected in the oral cavities of the HIV group than in the control (p=0.0001). Enterobacter cloacae was the most frequently isolated species in both groups. Counts of enteric bacteria and pseudomonas were significantly lower in patients with low CD4 counts (p=0.011); however, there was no difference relating to viral load. It may be concluded that HIV group showed greater species diversity and a higher prevalence of Enterobacteriaceae/Pseudomonadaceae., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Disease-suppressive soils are exceptional ecosystems in which crop plants suffer less from specific soil-borne pathogens than expected owing to the activities of other soil microorganisms. For most disease-suppressive soils, the microbes and mechanisms involved in pathogen control are unknown. By coupling PhyloChip-based metagenomics of the rhizosphere microbiome with culture-dependent functional analyses, we identified key bacterial taxa and genes involved in suppression of a fungal root pathogen. More than 33,000 bacterial and archaeal species were detected, with Proteobacteria, Firmicutes, and Actinobacteria consistently associated with disease suppression. Members of the γ-Proteobacteria were shown to have disease-suppressive activity governed by nonribosomal peptide synthetases. Our data indicate that upon attack by a fungal root pathogen, plants can exploit microbial consortia from soil for protection against infections.

The aim of this study was to study the psychrotrophic microbiota developing during milk creaming of Grana Trentino cheese-making. 138 isolates from raw whole milk, cream and skim milk samples were screened by Randomly amplified polymorphic DNA PCR biotyping and representative strains of each biotype were characterised by partial 16S rRNA gene sequencing and enzymatic activity. Pseudomonadaceae were commonly isolated in cream samples while Streptococcaceae and Enterobacteriaceae in milk samples. Moraxellaceae and Flavobacteriaceae were found in both cream and milk samples. More than 80% of psychrotrophic isolates could grow at 37°C. All Flavobacteriaceae and half of Pseudomonadaceae biotypes displayed proteolytic activity on milk agar even at low temperatures such as 10°C. All Streptococcaceae and some of Enterobacteriaceae displayed acidifying activity and almost all Acinetobacter spp. (Moraxellaceae) displayed lipolytic activity towards tributyrin. Even if psychrotrophic bacteria is not the dominant microbial group in raw milk, their total number increases during creaming and becomes one of the most present group together with Lactic Acid Bacteria. Their enzymatic activities may be key players in determining milk quality for cheese making., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Unlabelled: The purpose of this study was to evaluate the use of Lactobacillus plantarum, isolated from table olives "Bella di Cerignola," a traditional variety of Apulian region (Southern Italy), as a starter for this kind of food. We focused on the interaction of the starter with the natural occurring microflora, the quantitative/qualitative composition of yeast population, the decrease of pH, and the content of organic acids. After a preliminary characterization, 3 strains of Lb. plantarum, selected for their probiotic and technological performances, were used as a multiple-strain starter and inoculated (approximately 2%) in olives, processed according to Spanish style, brined at 8% and 10% of NaCl and added with 0.5% of glucose. The combination of the starter and glucose assured a correct fermentation course, decreasing the pH up to a safe value (4.3 to 4.5) and controlled the growth of yeasts. The concentrations of both L- and D-lactic acids increased throughout the fermentation, while citric and malic acids (both the isomers D and L) remained at low levels (0.2 to 0.4 g/L). Concerning yeast species, Candida guilliermondii was mainly isolated at the beginning (7 to 14 d), while C. famata prevailed at the end of fermentation., Practical Applications: To the question "How to standardize and maintain quality of "Bella di Cerignola" olives (Southern Italy)" we can suggest the following answer: use Lb. plantarum and a low amount of glucose (0.5%). The result is a decrease of the pH below the safety break point.

A total of 364 bacterial isolates, obtained from spinach leaves, were assayed in a decarboxylase broth containing histidine, lysine, and ornithine to check their ability to produce biogenic amines, and then quantified by high-performance liquid chromatography. Among these isolates, 240 formed cadaverine, 208 formed putrescine, and 196 formed histamine, in widely varying amounts. They frequently produced more than one biogenic amine. Klebsiella pneumoniae subsp. pneumoniae and Morganella morganii were the main histamine producers, with mean values of 1,600 and 2,440 mg/liter, respectively, followed by Pantoea spp. 3 (1,710 mg/liter) and Hafnia alvei (2,500 mg/liter). Enterobacter amnigenus and Enterobacter cloacae produced particularly high amounts of putrescine, with mean values of 2,340 and 2,890 mg/liter, respectively. The strongest cadaverine formation was shown by Serratia liquefaciens (3,300 mg/liter), Serratia marcescens (3,280 mg/liter), and Stenotrophomonas maltophilia (1,000 mg/liter).

The decline of take-all disease (Gaeumannomyces graminis var. tritici), which may take place during wheat monocropping, involves plant-protecting, root-colonizing microorganisms. So far, however, most work has focused on antagonistic fluorescent pseudomonads. Our objective was to assess the changes in rhizobacterial community composition during take-all decline of field-grown wheat. The study was based on the development and utilization of a taxonomic 16S rRNA-based microarray of 575 probes, coupled with cloning-sequencing and quantitative PCR. Plots from one experimental field grown with wheat for 1 yr (low level of disease), 5 yr (high level of disease) or 10 yr (low level of disease, suppressiveness reached) were used. Microarray data discriminated between the three stages. The outbreak stage (5 yr) was mainly characterized by the prevalence of Proteobacteria, notably Pseudomonas (Gammaproteobacteria), Nitrosospira (Betaproteobacteria), Rhizobacteriaceae, Sphingomonadaceae, Phyllobacteriaceae (Alphaproteobacteria), as well as Bacteroidetes and Verrucomicrobia. By contrast, suppressiveness (10 yr) correlated with the prevalence of a broader range of taxa, which belonged mainly to Acidobacteria, Planctomycetes, Nitrospira, Chloroflexi, Alphaproteobacteria (notably Azospirillum) and Firmicutes (notably Thermoanaerobacter). In conclusion, take-all decline correlated with multiple changes in rhizobacterial community composition, far beyond the sole case of pseudomonads.

16S rRNA gene sequenced-based phylogeny indicates that Rhizobacter dauci ATCC 43778(T) branches within the radiation of Methylibium type strains. A comparative chemotaxonomic study including fatty acid methyl esters, polar lipids and polyamines reveals significant differences that, in combination with the topology of phylogenetic trees, support a dissection of the genus Methylibium. The proposals of this study include the transfer of Methylibium fulvum to the genus Rhizobacter as Rhizobacter fulvus comb. nov. (type strain Gsoil 322(T) =KCTC 12591(T) =DSM 19916(T)) and the reclassification of Methylibium aquaticum as Piscinibacter aquaticus gen. nov., comb. nov. (the type strain of Piscinibacter aquaticus is IMCC1728(T) =KCCM 42364(T) =NBRC 102349(T) =DSM 19915(T)) and of Methylibium subsaxonicum as Rivibacter subsaxonicus gen. nov., comb. nov. (the type strain of Rivibacter subsaxonicus is BF49(T) =DSM 19570(T) =CIP 109700(T)). As a consequence of these reclassifications, emended descriptions of the genera Methylibium and Rhizobacter are provided.

Objective: The aim of this study was to correlate the presence of Enterobacteriaceae, Pseudomonadaceae, Moraxellaceae and Xanthomonadaceae on the posterior dorsum of the human tongue with the presence of tongue coating, gender, age, smoking habit and denture use., Material and Methods: Bacteria were isolated from the posterior tongue dorsum of 100 individuals in MacConkey agar medium and were identified by the API 20E system (Biolab-Mérieux)., Results: 43% of the individuals, presented the target microorganisms on the tongue dorsum, with greater prevalence among individuals between 40 and 50 years of age (p = 0.001) and non-smokers (p=0.0485)., Conclusions: A higher prevalence of Enterobacteriaceae and Pseudomonadaceae was observed on the tongue dorsum of the individuals evaluated. There was no correlation between these species and the presence and thickness of tongue coating, gender and presence of dentures.

The effect of short-term frozen storage prior to thawing on the quality of freeze-chilled chicken fillets was investigated, as was the effect of modified atmosphere packaging (MAP). Four process treatments were used: (1) fresh chicken chilled at 4 degrees C without previous freezing, (2) freeze-chilled for 7 days and thawed at 4 degrees C, (3) chilled at 4 degrees C packaged under MAP (70% N(2)-30%CO(2)), and (4) packaged under MAP, freeze-chilled for 7 days and thawed at 4 degrees C. Microbiological, chemical and sensory analyses were conducted on samples for a period up to 15 days. Freeze-chilled fillets gave a lower total viable count (TVC) at a given sampling day than chilled fillets. MAP, as expected, delayed microbial growth. The Pseudomonads were the dominant microbial species in fillets under aerobic conditions. MAP reduced the populations of Pseudomonads by 2-4 log cfu/g. Lactic acid bacteria (LAB) and Enterobacteriaceae increased progressively for all treatments throughout storage. Yeasts and molds were inhibited by MAP and by freeze chilling. Total volatile basic nitrogen (TVB-N) values increased rapidly for the chilled fillets but remained significantly lower for the freeze-chilled and the MA-packaged samples. MAP and especially freeze chilling enhanced drip loss. MAP did not affect redness or yellowness of product while freeze chilling decreased product redness. Lightness was not affected by either MAP or freeze chilling. Based on taste, which proved to be the most sensitive sensory attribute, shelf life of product ranged from 6 to 7 days for all treatments leading to the conclusion that freeze chilling is a suitable technology for fresh chicken fillets enabling their distribution as a frozen product and upon subsequent thawing at their final destination, their retail display as chilled products. MAP in combination with freeze chilling had a negligible effect on product quality.

The use of beneficial soil microorganisms as agricultural inputs for improved crop production requires selection of rhizosphere-competent microorganisms with plant growth-promoting attributes. A collection of 563 bacteria originating from the roots of pea, lentil, and chickpea grown in Saskatchewan was screened for several plant growth-promoting traits, for suppression of legume fungal pathogens, and for plant growth promotion. Siderophore production was detected in 427 isolates (76%), amino-cyclopropane-1-carboxylic acid (ACC) deaminase activity in 29 isolates (5%), and indole production in 38 isolates (7%). Twenty-six isolates (5%) suppressed the growth of Pythium sp. strain p88-p3, 40 isolates (7%) suppressed the growth of Fusarium avenaceum, and 53 isolates (9%) suppressed the growth of Rhizoctonia solani CKP7. Seventeen isolates (3%) promoted canola root elongation in a growth pouch assay, and of these, 4 isolates promoted the growth of lentil and one isolate promoted the growth of pea. Fatty acid profile analysis and 16S rRNA sequencing of smaller subsets of the isolates that were positive for the plant growth-promotion traits tested showed that 39%-42% were members of the Pseudomonadaceae and 36%-42% of the Enterobacteriaceae families. Several of these isolates may have potential for development as biofertilizers or biopesticides for western Canadian legume crops.

Biogenic amines and polyamines were studied in 18 market samples of spinach. Histamine and spermidine were detected in relatively high amounts in all samples within the ranges of 9.5-69.7 and 15.6-53.0 mg/kg, respectively. Other biologically active amines were either detected at low levels or not found at all. Changes in amine content during storage at 6 degrees C were studied. The content of most of the amines remained constant during storage, with the exception of spermidine and histamine. Spermidine showed a clear decreasing trend, whereas histamine significantly increased in all trials, but decreased at the end of the storage in two of the trials. Trials showing a decrease in histamine content also showed the highest spermidine decrease and recorded the highest pH values. Microbial loads throughout storage were also followed, with Pseudomonadaceae and Enterobacteriaceae being the predominant bacterial groups. Trials with higher microbial loads in initial samples also showed the highest histamine content in these samples. Potential explanations for both the formation and the degradation of histamine during storage are discussed.

The genetic structures of total bacterial and pseudomonad communities were characterized in rhizosphere soil and rhizoplane+root tissues of tobacco wild type and a ferritin overexpressor transgenic line (P6) by a cultivation-independent method using directly extracted DNA at the end of three consecutive plant cultures. The structure of total bacterial communities was characterized by automated ribosomal intergenic spacer analysis (A-RISA), and that of pseudomonad communities was characterized by PCR-restriction fragment length polymorphism (PCR-RFLP) from DNA amplified with specific primers. The structure of total bacterial communities was significantly modified in the rhizosphere soil by the overaccumulation of iron in the tobacco transgenic P6 line at the first culture, to a lesser extent at the second culture, and not at all at the third culture. No significant difference was recorded between the total communities associated with the roots (rhizoplane+root tissues) of the two plant genotypes in any of the cultures. In contrast, the difference in pseudomonad structure between the two plant genotypes increased with successive culture at the root level, but was not detected at a significant level in the rhizosphere soil. The impact of iron overaccumulation by the tobacco transgenic P6 line on pseudomonads supports previous findings on the importance of iron competition among fluorescent pseudomonads.

Bacterial colony counts in reusable water reservoirs in a humidified oxygen supply system maintained only by the addition of nonsterilized water, with or without washing (conventional procedure), were examined. Bacteria were detected in all the reservoirs even before the operation of the oxygen suppliers, and colony counts of more than 1 x 10(2)/ml of bacteria were detected in some reservoirs without washing. Many bacteria were also detected in many reservoirs, after the operation of the oxygen suppliers, indicating that bacteria had contaminated the water reservoirs and propagated during this operation. These results suggest that it is not recommended to maintain these water reservoirs by the conventional procedure.

The genus Pseudomonas (sensu stricto) represents a group of microorganisms directly involved in functions conferring plant health. We performed a study in the DOK long-term agricultural field experiment on the basis of previously published Pseudomonas-selective PCR primers in order to investigate the community structure of the microbial groups defined by the target range of these primers. Three different agricultural management systems, i.e., conventional, biodynamic, and bio-organic, along with mineral and unfertilized controls were investigated, with each system planted with either winter wheat or a grass-clover ley. Amplified small-subunit rRNA gene fragments were analyzed using the genetic profiling techniques restriction fragment length polymorphism (RFLP) and denaturing gradient gel electrophoresis (DGGE), revealing distinct differences between soils planted with winter wheat and grass clover but only minor differences between the management systems. Phylogenetic analyses of 59 clone sequences retrieved from bio-organic and unfertilized systems identified sequences related to Pseudomonas fluorescens and a novel cluster termed Cellvibrio-related Pseudomonadaceae (CRP). The CRP clones were exclusively isolated from winter wheat soil samples and were responsible for the crop-specific differences observed in RFLP and DGGE profiles. New primers were designed for the amplification of CRP targets directly from soil DNA, yielding strong signals exclusively for winter wheat soils. We concluded that crop-associated CRP exist in agricultural soils and that genetic profiling followed by specific probe design represents a valuable approach for identification as well as sensitive and rapid monitoring of novel microbial groups in the environment.

The key enzyme catalyzing the second step in the phenol degradation meta-cleavage pathway (C230) has been purified to homogeneity from a new bacterial strain, which belongs to genus Pseudomonas. The species was growing on phenol as carbon source. The C230 was detected and identified by absorption spectroscopy. The protein was isolated using sucrose density centrifugation and anion exchange chromatography. The purified protein showed a molecular mass of 32 kDa to sodium dodecyl sulfate polyacrylamid gel electrophoresis and an isoelectric point of 5 estimated by analytical isoelectrical focusing. Matrix-assisted laser desorption ionization-time of flight mass spectrometry and peptide mapping was attempted for the identification of the isolated protein and proteins involved in the metabolic pathway.

During the past ten years, bacterial soft rot and midrib rot of glasshouse-grown butterhead lettuce (Lactuca sativa L. var. capitata) and field-grown endive (Cichorium endivia L.) has become increasingly common in the region of Flanders, Belgium. Severe losses and reduced market quality caused by bacterial rot represent an important economical threat for the production sector. Symptoms of midrib rot are a brownish rot along the midrib of one or more inner leaves, often accompanied by soft rot of the leaf blade. Twenty-five symptomatic lettuce and endive samples were collected from commercial growers at different locations in Flanders. Isolations of dominant bacterial colony types on dilution plates from macerated diseased tissue extracts yielded 282 isolates. All isolates were characterized by colony morphology and fluorescence on pseudomonas agar F medium, oxidase reaction, and soft rot ability on detached chicory leaves. Whole-cell fatty acid methyl esters profile analyses identified the majority of isolates (85%) as belonging to the Gammaproteobacteria, which included members of the family Enterobacteriaceae (14%) and of the genera Pseudomonas (73%), Stenotrophomonas (9%), and Acinetobacter (3%). Predominant bacteria were a diverse group of fluorescent Pseudomonas species. They were further differentiated based on the non-host hypersensitive reaction on tobacco and the ability to rot potato slices into 4 phenotypic groups: HR-/P- (57 isolates), HR-/P+ (54 isolates), HR+/P (16 isolates) and HR+/P+ (35 isolates). Artificial inoculation of suspensions of HR-, pectolytic fluorescent pseudomonads in the leaf midrib of lettuce plants produced various symptoms of soft rot, but they did not readily cause symptoms upon spray inoculation. Fluorescent pseudomonads with phenotype HR+ were consistently isolated from typical dark midrib rot symptoms, and selected isolates reproduced the typical midrib rot symptoms when spray-inoculated onto healthy lettuce plants.

Endophytic and epiphytic bacteria were isolated from two soybean cultivars (Foscarin and Cristalina). Significant differences were observed in bacterial population densities in relation to season of isolation, soybean growth phase and the tissues from which the isolates were obtained. The isolates were identified by partial 16S rDNA sequence analysis, with most of the isolates belonging to the Pseudomonaceae, Burkholderiacea and Enterobacteriaceae groups. The potential of the isolates for plant growth promotion was evaluated by screening for indoleacetic acid (IAA) production and mineral phosphate solubilization; 34% of endophytic bacteria produced IAA and 49% were able to solubilize mineral phosphate whereas only 21% of epiphytic bacteria produced IAA although 52% were able to solubilize mineral phosphate. A high frequency of IAA producing isolates occurred in the early ripening Foscarin cultivar whereas a high percentage of phosphate solubilizing isolates were obtained from plants in the initial development stage (V6). We also found that 60% of endophytic and 69% of epiphytic isolates that produced IAA and solubilized mineral phosphate were also able to fix nitrogen in vitro. The soybean-associated bacteria showing characteristics related to plant growth promotion were identified as belonging to the genera Pseudomonas, Ralstonia, Enterobacter, Pantoea and Acinetobacter.

Three bacterial strains, designated as Wphe1, Sphe1, and Ophe1, were isolated from Greek soils contaminated with polycyclic aromatic hydrocarbon (PAH)-containing waste from the wood processing, steel, and oil refinery industries. Wphe1, Sphe1, and Ophe1 were characterized and identified as species of Pseudomonas, Microbacterium, and Paracoccus, respectively, based on Gram staining, biochemical tests, phospholipid analysis, FAME analysis, G+C content and 16S rRNA gene sequence analysis. The results of gas chromatography showed that strain Wphe1 degraded naphthalene, phenanthrene, and m-cresol over a wide temperature range; strain Sphe1 was a degrader of phenanthrene and n-alkanes; most interestingly, strain Ophe1 degraded anthracene, phenanthrene, fluorene, fluoranthene, chrysene, and pyrene, as well as cresol compounds and n-alkanes as sole carbon source. This is the first report of a representative of the genus Paracoccus capable of degrading PAHs with such versatility. These three strains may be useful for bioremediation applications.

A 16S rRNA gene-based fingerprinting method was developed for the identification of Azotobacteraceae and tested onto 48 soil isolates and 28 reference strains belonging to the free-living nitrogen-fixing bacterial group and to the most common species found in soil samples. According to this method, the 16S rRNA gene was amplified using universal primers for Eubacteria and PCR products were subsequently digested with RsaI, HhaI, HpaII, FnuDII, and AluI. The analysis of the restriction profiles obtained showed that the method is able to define a unique species-specific phylotype (SSP) for each of the eight Azotobacteraceae species tested. Cluster analysis was successfully employed for the identification of members of the family Azotobacteraceae, being assignation into species of the isolates confirmed by means of partial 16S rRNA gene sequencing.

Recent studies have indicated possible changes in the incidence of acute mastoiditis. A retrospective review of children discharged with a diagnosis of acute mastoiditis was undertaken to describe the epidemiology, clinical presentation, microbiology, and treatment of acute mastoiditis over past years. Demographic historic, clinical, and laboratory data were collected. Eighty-six children (88 episodes of acute mastoiditis) were identified (1 month-16 years) (median 3.3 years). Almost half had a history of middle ear disease; 8% recurrent episodes and 68.2% received antibiotics preadmission, 91.2% for acute otitis media. Bacterial etiology was established in 43 patients (68.2% isolation rate). Pseudomonas aeruginosa and Streptococcus pneumoniae were the most frequently isolated agents. This review showed a significant increase (150%) in the number of patients with acute mastoiditis.

Genus-specific real-time PCR assays were developed and optimized for the direct culture-independent detection and quantification of Mycobacteria and Pseudomonads in contaminated metalworking fluids (MWF) and the results were compared with conventional culturing using selective media. It included optimization of the direct DNA isolation from the fluid matrix and the amplification conditions using genus-specific primers. Mycobacterium-specific primers based on 65-kDa heat shock protein (hsp) gene, and Pseudomonas-specific primers based on 16S rRNA gene were used. A standard curve was developed each for the two model bacterial species Mycobacterium immunogenum and Pseudomonas fluorescens, representing two important genera frequently isolated from MWF. A minimum quantification limit of 10 cells/ml was achieved although as low as 1 cell/ml yielded a detectable amplicon signal. Of the twenty MWF field samples contaminated with mixed microflora, only two samples yielded putative colonies of Mycobacteria and Pseudomonads by culturing method, while seven samples responded to the genus-specific real-time PCR detection and quantification for each genus. In contrast to the low culturable counts, the real-time PCR based cell counts ranged from 1.3 x 10(2) to 5.5 x 10(5)cells/ml and 5.2 x 10(2) to 7.0 x 10(5)cells/ml for Mycobacteria and Pseudomonads, respectively, indicating a significant non-culturable fraction in the fluids, for the two genera. This is the first application of real-time PCR protocol to MWF samples for detection and quantification of total (culturable and non-culturable) Mycobacteria and Pseudomonads without culturing.

Production of indole-3-acetic acid (IAA), a key physiological feature of culturable, O2-tolerant bacteria associated with the freshwater macrophyte Juncus effusus L., was examined over a period of 2 years. Up to 74% of rhizobacteria identified and tested produced IAA. The number of indoleacetic acid producers decreased in winter. IAA was produced even when L-tryptophan, a precursor of IAA, was not added to the medium. Most of the IAA-producing strains were dominated by strains that were not identifiable to species level on the basis of API testing. Based on 16S rRNA gene sequencing and fatty acid analysis, it was found that IAA-producing rhizosphere bacteria associated with the freshwater wetland plant Juncus effusus L. are representatives of several families, including the Enterobacteriaceae, Pseudomonadaceae, Aeromonadaceae, Burkholderiaceae, and Bacillaceae. This study identifies numerous potentially important bacterial physiological groups of freshwater wetlands. Additionally, the study provides a baseline for monitoring and assessing the mutualistic relationships of wetland plants with rhizosphere bacteria in freshwater wetlands.

Mycobacteria and pseudomonads occurring in modern metalworking fluids (MWF) have been implicated in occupational health hazards as causal agents for hypersensitivity pneumonitis (HP) and other respiratory illnesses in machine workers exposed to these fluids and their aerosols. Unlike the conventional cultural and biochemical methods, which are often slow and ambiguous and detect only culturable cells, DNA-based methods offer a time-saving alternative for reliable detection and identification of both culturable and nonculturable bacteria in MWF and for selective quantification of individual genera of pathogens of interest in these fluids. This is the first report on DNA-based direct detection of mycobacteria and pseudomonads in MWF without culturing. Genus-specific PCR approach was successfully applied for screening of field MWF samples originating from different industrial users for detection of mycobacteria or pseudomonads including both culturable and nonculturable cells. PCR in combination with amplicon DNA sequencing led to the identification of Mycobacterium chelonae, Pseudomonas nitroreducens, and an undefined Pseudomonas species from these fluids. Genome fingerprinting by pulsed-field gel electrophoresis (PFGE) on Mycobacterium isolates further showed that the isolates represented three strains of M. chelonae although the possibility of one of the strains being clonal with M. immunogenum cannot be excluded. In parallel efforts, a quantitative competitive PCR method developed based on the Pseudomonas-specific PCR was applied to quantify total P. fluorescens cells in contaminated metalworking fluid and MWF aerosol without culturing. The DNA-based protocols developed in this study will allow rapid screening of field MWF samples for the presence of both culturable and nonculturable cells and thus facilitate effective fluid management and timely exposure assessment.