bluuebunny/crossref_metadata_2025 huggingface dataset:
Response to trisodium phosphate treatment ofSalmonellaChester attached to fresh-cut green pepper slices
Authors: Liao Ching-Hsing , Cooke Peter H
Published: April 2011

A laboratory model using green pepper disks was developed to investigate the attachment of Salmonella Chester on plant tissue and to evaluate the effectiveness of sanitizer agents in inactivating attached bacteria on fruits. Pepper disks (14 mm in diam, and 3-4 mm in thickness) were immersed in a bacterial suspension containing 1.5 × 107cfu·mL-1of S. Chester for 30 s and subsequently air-dried at room temperature for 10 min. Approximately 30% of the bacteria retained on the disk after immersion were firmly attached and could not be removed by two washes and agitation. A positive correlation was observed between the number of bacteria attached and the concentration of bacteria in the suspension. Population studies and scanning electron microscopic examinations revealed that attachment of S. Chester on pepper disks occurred mainly on the surfaces of injured (cut) tissue but rarely on the unbroken skin. When inoculated disks were treated with 3% to 12% (w/v) of trisodium phosphate (TSP) at pH 12.3 for 5 min, the population of bacteria on the disk was reduced by 10- to 100-fold. A small portion (0.7% to 7.1%) of bacteria attached to the disk were either resistant to or protected from the TSP treatment. When the pH of TSP solution was reduced from 12.3 to 4.5, the effectiveness of TSP in inactivating S. Chester on pepper disks was reduced by 26%. This study shows that surfaces of injured fruit tissue are the principal sites for bacterial attachment, and a small portion of the bacteria attached to the tissue are resistant to the sanitizer treatment. Avoiding mechanical injuries to fresh fruits during and after harvest would reduce the chance of pathogen attachment and contamination on green pepper and fruits of similar nature.Key words: Salmonella, attachment, detachment, plant tissue, sanitizer treatment.
Ability ofEscherichia coliO157:H7 isolates to colonize the intestinal tract of conventional adult CD1 mice is transient
Authors: Conlan J Wayne , Bardy Sonia L , KuoLee Rhonda , Webb Ann , Perry Malcolm B
Published: April 2011

In an attempt to improve upon a current mouse model of intestinal colonization by Escherichia coli O157:H7 used in this laboratory for vaccine development, nine clinical isolates of the pathogen were screened for their ability to persist in the intestinal tract of conventional adult CD-1 mice. None of the test isolates of E. coli O157:H7 were capable of colonizing these mice for a period of more than two weeks. Most of the isolates appeared to be benign for the experimental host, but one isolate was lethal. This virulence correlated with the ability of the latter isolate to produce large quantities of Shiga-like toxin 2 in vitro.
Purification, cloning, and DNA sequence analysis of a chitinase from an overproducing mutant of Streptomyces peucetius defective in daunorubicin biosynthesis
Authors: Vetrivel Kuzhandhaivel S , Pandian Shunmugiah K , Chaudhary Uma , Dharmalingam Kuppamuthu
Published: April 2011

Extracellular chitinases of Streptomyces peucetius and a chitinase overproducing mutant, SPVI, were purified to homogeneity by ion exchange and gel filtration chromatography. The purified enzyme has a molecular mass of 42 kDa on SDS-PAGE, and the N-terminal amino acid sequence of the protein from the wild type showed homology to catalytic domains (Domain IV) of several other Streptomyces chitinases such as S. lividans 66, S. coelicolor A3(2), S. plicatus, and S. thermoviolaceus OPC-520. Purified SPVI chitinase cross-reacted to anti-chitinase antibodies of wild-type S. peucetius chitinase. A genomic library of SPVI constructed in E. coli using λ DASH II was probed with chiC of S. lividans 66 to screen for the chitinase gene. A 2.7 kb fragment containing the chitinase gene was subcloned from a λ DASH II clone, and sequenced. The deduced protein had a molecular mass of 68 kDa, and showed domain organization similar to that of S. lividans 66 chiC. The N-terminal amino acid sequence of the purified S. peucetius chitinase matched with the N-terminus of the catalytic domain, indicating the proteolytic processing of 68 kDa chitinase precursor protein to 42 kDa mature chitinase containing the catalytic domain only. A putative chiR sequence of a two-component regulatory system was found upstream of the chiC sequence.Key words: chitinase, chitinase purification, Streptomyces peucetius, daunorubicin, chiC.
The panmictic nature ofNeisseria meningitidisserogroup B during a period of endemic disease in Canada
Authors: Ashton Fraser E , Caugant Dominique A
Published: April 2011

Three hundred and one (301) strains of Neisseria meningitidis serogroup B, isolated from patients with meningococcal disease during the years 19941996, were subjected to multilocus enzyme electrophoresis, serotyping, and serosubtyping. Based on the analyses of 14 enzyme loci, 177 electrophoretic types (ETs) were identified. Of these, 136 were represented by single isolates and 41 were represented by multiple isolates (range 231). The mean genetic diversity for isolates was 0.444 and for ETs was 0.440. The index of association (IA) between loci was 0.530 ± 0.08 for isolates and 0.256 ± 0.10 for ETs. Cluster analysis revealed the presence of 39 lineages each represented by a single ET or clusters of ETs. The most common serotypes were 4, 15, and 14 and accounted for 84 (28.0%), 53 (17.6%), and 32 (10.6%) of the isolates, respectively, and were dispersed amongst 46 ETs (1122), 35 ETs (3165), and 26 ETs (1876), respectively. The 109 (36.6%) nontypable (NT) isolates were amongst 74 ETs (6177). The mean genetic diversity for serotypes 4, 15, and 14 and NT isolates was 0.368, 0.371, 0.343, and 0.442, respectively, and for ETs was 0.363, 0.354, 0.397, and 0.440, respectively. Combinations of serotypes and serosubtypes (number of isolates) that occurred most frequently were 4:P1.14 (17), 14:P1.16 (16), NT:P1.16 (16), 15:P1.16 (13), and NT:P1.13 (13). The majority of group B disease in Canada during 19941996 was caused by meningococci of considerable genetic diversity, and reflects a situation of endemic disease. However, the results also indicate that organisms belonging to the ET-5 complex, which has been responsible for outbreaks of group B disease globally for several decades, have been introduced into the country.Key words: meningococcal, genotypes, serotypes, serosubtypes, Neisseria meningitidis.
Monitoring of bacteria in acid mine environments by reverse sample genome probing
Authors: Léveillé S A , Leduc L G , Ferroni G D , Telang A J , Voordouw G
Published: April 2011

A variety of microorganisms can exist in acid mine drainage (AMD) environments, although their contribution to AMD problems is unclear. Environmental strains of Thiobacillus ferrooxidans and Thiobacillus acidophilus were purified by repeated plating and single-colony isolation on iron salts and tetrathionate media, respectively. Thiobacillus thiooxidans was enriched on sulfur-containing media. For the isolation of Leptospirillum ferrooxidans, iron salts and pyrite media were inoculated with environmental samples. However, L. ferrooxidans was never recovered on solid media. Denatured chromosomal DNAs from type and (or) isolated strains of T. ferrooxidans, T. acidophilus, T. thiooxidans, and L. ferrooxidans were spotted on a master filter for their detection in a variety of samples by reverse sample genome probing (RSGP). Analysis of enrichments of environmental samples by RSGP indicated that ferrous sulfate medium enriched T. ferrooxidans strains, whereas all thiobacilli grew in sulfur medium, T. thiooxidans strains being dominant. Enrichment in glucose medium followed by transfer to tetrathionate medium resulted in the selection of T. acidophilus strains. DNA was also extracted directly (without enrichment) from cells recovered from AMD water or sediments, and was analyzed by RSGP to describe the communities present. Strains showing homology with T. ferrooxidans and T. acidophilus were found to be major community components. Strains showing homology with T. thiooxidans were a minor community component, whereas strains showing homology with L. ferrooxidans were not detected.Key words: AMD, bacteria, DNA probing.
Certaines légumineuses du genre Crotalaria sont spécifiquement nodulées par une nouvelle espèce de Methylobacterium
Authors: Sy Abdoulaye , Giraud Éric , Samba Ramatoulaye , Lajudie Philippe de , Gillis Monique , Dreyfus Bernard
Published: April 2011

We studied a collection of 126 rhizobial isolates from eight species of Crotalaria (C. comosa, C. glaucoides, C. goreensis, C. hyssopifolia, C. lathyroides, C. perrottetii, C. podocarpa, and C. retusa) growing in Senegal. Nodulation and nitrogen-fixation tests on nine Crotalaria species revealed two specificity groups within the genus Crotalaria. Group I consists of plants solely nodulated by very specific fast-growing strains. Group II plants are nodulated by slow-growing strains similar to promiscuous Bradyrhizobium spp. strains already reported to nodulate many tropical legumes. SDSPAGE studies showed that slow-growing strains grouped with Bradyrhizobium while fast-growing strains constituted a homogeneous group distinct from all known rhizobia. Amplified ribosomal DNA restriction analysis (ARDRA) of 10 representative strains of this group using four restriction enzymes showed a single pattern for each enzyme confirming the high homogeneity of group I. The 16S rDNA sequence analysis revealed that this specific group belonged to the genus Methylobacterium, thus constituting a new branch of nodulating bacteria.Key words: Crotalaria, Methylobacterium, rhizobium, symbiosis.
Purification, cloning, and sequencing of an enzyme mediating the reductive dechlorination of tetrachloroethylene (PCE) fromClostridium bifermentansDPH-1
Authors: Okeke Benedict C , Chang Young C , Hatsu Masahiro , Suzuki Tohru , Takamizawa Kazuhiro
Published: April 2011

An enzyme mediating the reductive dechlorination of tetrachloroethylene (PCE) from cell-free extracts of Clostridium bifermentans DPH-1 was purified, cloned, and sequenced. The enzyme catalyzed the reductive dechlorination of PCE to cis-1,2-dichloroethylene via trichloroethylene, at a Vmaxand Kmof 73 nmol/mg protein and 12 µM, respectively. Maximal activity was recorded at 35°C and pH 7.5. Enzymatic activity was independent of metal ions but was oxygen sensitive. A mixture of propyl iodide and titanium citrate caused a light-reversible inhibition of enzymatic activity suggesting the involvement of a corrinoid cofactor. The molecular mass of the native enzyme was estimated to be approximately 70 kDa. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and matrix-assisted laser desorption ionization-time of flight/mass spectrometry (MALDITOF/MS) revealed molecular masses of approximately 35 kDa and 35.7 kDa, respectively. A broad spectrum of chlorinated aliphatic compounds (PCE, trichloroethylene, cis-1,2-dichloroethylene, trans-1,2-dichloroethylene, 1,1-dichloroethylene, 1,2-dichloropropane, and 1,1,2-trichloroethane) was degraded. With degenerate primers designed from the N-terminal sequence (27 amino acid residues), a partial sequence (81 bp) of the encoding gene was amplified by polymerase chain reaction (PCR) and sequenced. Southern analysis of C. bifermentans genomic DNA using the PCR product as a probe revealed restriction fragment bands. A 5.0 kb ClaI fragment, harboring the relevant gene (designated pceC) was cloned (pDEHAL5) and the complete nucleotide sequence of pceC was determined. The gene showed homology mainly with microbial membrane proteins and no homology with any known dehalogenase, suggesting a distinct PCE dehalogenase.Key words: tetrachloroethylene, Clostridium bifermentans DPH-1, PCE dehalogenase, gene cloning.
Increased phospholipid transfer protein activity inAspergillus oryzaegrown on various industrial phospholipid sources
Authors: Asther Michèle , Record Eric , Antona Claudine , Asther Marcel
Published: April 2011

The effect of industrial carbon sources on phospholipid transfer protein production was investigated. Phospholipid fractions of different composition were prepared from various plant oils (i.e., soybean, rapeseed, and sunflower) according to the Lucas Meyer extraction and purification process. The effect of these fractions on phospholipid transfer protein activity of cell extracts from Aspergillus oryzae grown on medium containing these phospholipids as sole carbon source was studied. It was shown that phospholipid transfer activity was markedly increased by extracts containing a particular phospholipid composition. However, this stimulation depends mainly upon the phospholipid composition of the fraction used as fermentation substrate. Fractions enriched mainly in phosphatidylinositol (Epikuron 110), at the expense of phosphatidylcholine, were the most efficient sources for phospholipid transfer protein production by A. oryzae. Maximal phospholipid transfer activity, as well as biomass production, were increased 4.1- and 9.7-fold, respectively, when cultures were supplemented with Epikuron 110 prepared from sunflower lecithin, as compared to glucose-control cultures.Key words: Aspergillus oryzae, industrial phospholipid sources, phospholipid transfer protein.
Adequacy of in situ glass slides and direct sand extractions to assess the microbiota within sand columns used for drinking water treatment
Authors: Långmark Jonas , Ashbolt Nicholas J , Szewzyk Ulrich , Stenström Thor Axel
Published: April 2011

Historically, Cholodny-Rossi buried glass slide techniques have been used to study the microbiota of subsurface environments, yet the bias of such a technique has not been compared against direct sand extraction using modern in situ probing. Over a period of 34 wk, four separate 4-m-deep sand columns receiving raw lake water were examined to compare direct extraction of sand filter biofilm material against in situ glass slide biofilms. Significantly different DAPI direct counts and fluorescent in situ hybridization signals for major phylogenetic groups were observed. Not only were lower proportions (P < 0.001) of EUB338-probed DAPI cells observed on in situ glass slides, but also fewer γ-Proteobacteria (12%21%) and more α-Proteobacteria (16%33%) when compared to direct sand extracts. Hence, investigators of the microbial ecology of even simple sand biofilms must consider the inherent biases from "accepted" methods and seek further independent methods to identify those which may be most accurate.Key words: sand filter, biofilms, in situ hybridization, groundwater recharge.
Bacteriophage T4 multiplication in a glucose-limited Escherichia coli biofilm
Authors: Corbin Brian D , McLean Robert JC , Aron Gary M
Published: April 2011

An Escherichia coli K-12 biofilm was grown at a dilution rate of 0.028 h-1 for 48 h in a glucose-limited chemostat coupled to a modified Robbins' device to determine its susceptibility to infection by bacteriophage T4. Bacteriophage T4 at a multiplicity of infection (MOI) of 10 caused a log reduction in biofilm density (expressed as colony forming units (CFU) per cm2) at 90 min postinfection. After 6 h, a net decrease and equilibrium in viral titer was seen. When biofilms were exposed to T4 phage at a MOI of 100, viral titer doubled after 90 min. After 6 h, viral titers (expressed as plaque forming units (PFU) per cm2) stabilized at levels approximately one order of magnitude higher than seen at a MOI of 10. Scanning confocal laser microscopy images also indicated disruption of biofilm morphology following T4 infection with the effects being more pronounced at a MOI of 100 than at a MOI of 10. These results imply that biofilms under carbon limitation can act as natural reservoirs for bacteriophage and that bacteriophage can have some influence on biofilm morphology.Key words: bacteriophage T4, biofilm, biofilm morphology, bacteriophage ecology, carbon limitation.
Characterization of plant growth promoting rhizobacteria isolated from polluted soils and containing 1-aminocyclopropane-1-carboxylate deaminase
Authors: Belimov Andrei A , Safronova Vera I , Sergeyeva Tatyana A , Egorova Tatyana N , Matveyeva Victoria A , Tsyganov Viktor E , Borisov Alexey Y , Tikhonovich Igor A , Kluge Christoph , Preisfeld Angelika , Dietz Karl-Josef , Stepanok Vitaley V
Published: April 2011

Fifteen bacterial strains containing 1-aminocyclopropane-1-carboxylate (ACC) deaminase were isolated from the rhizoplane of pea (Pisum sativum L.) and Indian mustard (Brassica juncea L.) grown in different soils and a long-standing sewage sludge contaminated with heavy metals. The isolated strains were characterized and assigned to various genera and species, such as Pseudomonas brassicacearum, Pseudomonas marginalis, Pseudomonas oryzihabitans, Pseudomonas putida, Pseudomonas sp., Alcaligenes xylosoxidans, Alcaligenes sp., Variovorax paradoxus, Bacillus pumilus, and Rhodococcus sp. by determination of 16S rRNA gene sequences. The root elongation of Indian mustard and rape (Brassica napus var. oleifera L.) germinating seedlings was stimulated by inoculation with 8 and 13 isolated strains, respectively. The bacteria were tolerant to cadmium toxicity and stimulated root elongation of rape seedlings in the presence of 300 µM CdCl2 in the nutrient solution. The effect of ACC-utilising bacteria on root elongation correlated with the impact of aminoethoxyvinylglycine and silver ions, chemical inhibitors of ethylene biosynthesis. A significant improvement in the growth of rape caused by inoculation with certain selected strains was also observed in pot experiments, when the plants were cultivated in cadmium-supplemented soil. The biomass of pea cv. Sparkle and its ethylene sensitive mutant E2 (sym5), in particular, was increased through inoculation with certain strains of ACC-utilising bacteria in pot experiments in quartz sand culture. The beneficial effect of the bacteria on plant growth varied significantly depending on individual bacterial strains, plant genotype, and growth conditions. The results suggest that plant growth promoting rhizobacteria containing ACC deaminase are present in various soils and offer promise as a bacterial inoculum for improvement of plant growth, particularly under unfavourable environmental conditions.Key words: ACC deaminase, cadmium, ethylene, Indian mustard, pea, phytoremediation, rape, rhizobacteria.
Sterol composition of itraconazole-resistant and itraconazole-susceptible isolates of Aspergillus fumigatus
Authors: Dannaoui Eric , Persat Florence , Borel Elisabeth , Piens Marie-Antoinette , Picot Stéphane
Published: April 2011

Sterol composition of four clinical isolates of Aspergillus fumigatus resistant to itraconazole was determined by gas chromatography  mass spectrometry and compared with that of four susceptible strains. For all strains, the major sterol was ergosterol. Sterol compositions were qualitatively and quantitatively similar for the resistant and susceptible strains. These results suggest that itraconazole resistance is not related, for the strains studied, to alterations in the ergosterol synthesis pathway.Key words: Aspergillus fumigatus, itraconazole resistance, sterol composition.
Evaluation of bacteria isolated from rice for plant growth promotion and biological control of seedling disease of rice
Authors: Adhikari Tika B , Joseph C M , Yang Guoping , Phillips Donald A , Nelson Louise M
Published: April 2011

Of 102 rhizoplane and endophytic bacteria isolated from rice roots and stems in California, 37% significantly (P [Formula: see text] 0.05) inhibited the growth in vitro of two pathogens, Achlya klebsiana and Pythium spinosum, causing seedling disease of rice. Four endophytic strains were highly effective against seedling disease in growth pouch assays, and these were identified as Pseudomonas fluorescens (S3), Pseudomonas tolaasii (S20), Pseudomonas veronii (S21), and Sphingomonas trueperi (S12) by sequencing of amplified 16S rRNA genes. Strains S12, S20, and S21 contained the nitrogen fixation gene, nifD, but only S12 was able to reduce acetylene in pure culture. The four strains significantly enhanced plant growth in the absence of pathogens, as evidenced by increases in plant height and dry weight of inoculated rice seedlings relative to noninoculated rice. Three bacterial strains (S3, S20, and S21) were evaluated in pot bioassays and reduced disease incidence by 50%73%. Strain S3 was as effective at suppressing disease at the lowest inoculum density (106 CFU/mL) as at higher density (108 CFU/mL or undiluted suspension). This study indicates that selected endophytic bacterial strains have potential for control of seedling disease of rice and for plant growth promotion.Key words: biological control, plant growth promotion, endophytes, rice, seedling disease.
Development of a method for mRNA differential display in filamentous fungi: comparison of mRNA differential display reverse transcription polymerase chain reaction and cDNA amplified fragment length polymorphism in Leptosphaeria maculans
Authors: Gellatly Kevin S , Ash Gavin J , Taylor Janet L
Published: April 2011

We modified a technique, cDNA-AFLP, for identifying differentially expressed genes in plants to work in the filamentous fungus Leptosphaeria maculans (Desmaz.) Ces. & De Not. The cDNA fragments generated by our method ranged in size from approximately 100 to 400 bps. On average, twice as many cDNA fragments were amplified per primer set with cDNA amplified fragment length polymorphism in comparison with mRNA differential display reverse transcription polymerase chain reaction. The DNA fragments of interest were excised from gels and analyzed by single-stranded conformation polymorphism to eliminate nondifferentially expressed cDNA contamination. The method was used to examine gene expression differences between cultures grown in the presence or absence of an analog of the Brassica phytoalexin brassinin. Eleven of the fourteen fragments examined were determined by reverse Northern blot to be differentially expressed. In examining gene expression differences between young cultures not producing sirodesmins and older cultures that were producing these phytotoxins, we found 17 of 25 fragments were differentially expressed. Northern blots with these fragments confirmed the results.Key words: Phoma lingam, ascomycete, blackleg, Brassica.
Efficient and predictable recovery of viruses from water by small scale ultrafiltration systems
Authors: Winona L J , Ommani A W , Olszewski J , Nuzzo J B , Oshima K H
Published: April 2011

Current methods to concentrate viruses from large volumes of water are prone to inconsistent results and are costly and complex procedurally. Ultrafiltration can utilize size exclusion rather than adsorption and (or) elution to concentrate viruses and, therefore, may offer greater flexibility in developing methods that can provide more consistent recoveries among different viruses and widely varying water conditions. Two small scale ultrafiltration systems (hollow fiber and tangential flow) were tested with a virus suspended in 2 L of reagent grade, tap, ground, or surface water. Three model viruses were used (bacteriophages PP7 and T1 and poliovirus) to compare and characterize the recovery of viruses with the two ultrafiltration systems. Pretreatment of the ultrafilters with blocking agents and the use of elution agents can serve to prevent viral adsorption to the filter surface or to elute bound virus and keep viral agents suspended in the retentate. The use of a blocking and elution step concentrated viruses (>60% recovery) from widely varying water qualities, including surface water, such that a single method can be used to efficiently concentrate viruses from all of the water types tested. Both ultrafiltration systems appear to be able to efficiently recover viruses; however, the hollow fiber systems provided slightly better results in the 2-L volumes tested.Key words: ultrafiltration, waterborne virus, poliovirus, enterovirus.
Directed evolution to produce an alkalophilic variant from aNeocallimastix patriciarumxylanase
Authors: Chen Yew-Loom , Tang Tsung-Yin , Cheng Kuo-Joan
Published: April 2011

The catalytic domain of a xylanase from the anaerobic fungus Neocallimastix patriciarum was made more alkalophilic through directed evolution using error-prone PCR. Transformants expressing the alkalophilic variant xylanases produced larger clear zones when overlaid with high pH, xylan-containing agar. Eight amino acid substitutions were identified in six selected mutant xylanases. Whereas the wild-type xylanase exhibited no activity at pH 8.5, the relative and specific activities of the six mutants were higher at pH 8.5 than at pH 6.0. Seven of the eight amino acid substitutions were assembled in one enzyme (xyn-CDBFV) by site-directed mutagenesis. Some or all of the seven mutations exerted positive and possibly synergistic effects on the alkalophilicity of the enzyme. The resulting composite mutant xylanase retained a greater proportion of its activity than did the wild type at pH above 7.0, maintaining 25% of its activity at pH 9.0, and its retention of activity at acid pH was no lower than that of the wild type. The composite xylanase (xyn-CDBFV) had a relatively high specific activity of 10 128 µmol glucose·min1·(mg protein)1at pH 6.0. It was more thermostable at 60°C and alkaline tolerant at pH 10.0 than the wild-type xylanase. These properties suggest that the composite mutant xylanase is a promising and suitable candidate for paper pulp bio-bleaching.Key words: xylanase, Neocallimastix patriciarum, alkalophilicity, random mutagenesis, directed evolution.
Elemental analysis of uncultured magnetotactic bacteria exposed to heavy metals
Authors: Keim Carolina N , Lins Ulysses , Farina Marcos
Published: April 2011

Natural enrichments of magnetotactic bacteria were used to study the sites where heavy metals accumulate in uncultured bacteria. Most bacteria obtained by magnetic concentration from these enrichments contained, in addition to the magnetosomes, large phosphorus-rich granules in the cytoplasm. Metal (Zn, Mn, Sr, Cd, Al, Cr, and Pb) chlorides were added independently to the enrichments, and after 24 h, the elemental composition of the phosphorus-rich granules, magnetosomes, and "soft parts" (cytoplasm plus cell envelope) of whole bacteria was analyzed by energy-dispersive X-ray analysis on a transmission electron microscope. All bacteria contained Mn and Sr in the phosphorus-rich granules; some of them presented Mn peaks also in the soft parts. Zinc accumulation was variable and was found mainly in the phosphorus-rich granules, but also in the soft part of some bacteria. Some analyzed bacteria presented Zn peaks only in the soft parts, and some of them did not present Zn in any structure. Cadmium and Al were found only in the granules of some bacteria. Chromium was found in the soft parts of some bacteria. Lead was not detected in any bacteria. We concluded that the phosphorus-rich granules are major sites for metal accumulation by these bacteria. No conclusive results for magnetosomes were obtained because of the limitations of the analytical techniques particularly when used for whole cell analysis.Key words: magnetotactic bacteria, polyphosphate granules, heavy metals, uncultured bacteria, biomineralization, X-ray microanalysis.
Isolation and characterization of an antifreeze protein with ice nucleation activity from the plant growth promoting rhizobacteriumPseudomonas putidaGR12-2
Authors: Xu Hao , Griffith Marilyn , Patten Cheryl L , Glick Bernard R
Published: April 2011

An antifreeze protein secreted to the growth medium by the plant growth promoting rhizobacterium Pseudomonas putida GR12-2 was purified to apparent homogeneity. The purified protein has a molecular mass of 164 ± 15 kDa and an isoelectric point of 5.3, contains both carbohydrate and lipid moieties, and is relatively rich in glycine and alanine. The properties of the purified antifreeze protein are similar to the properties previously reported for bacterial ice-nucleation proteins. In fact, the purified antifreeze protein also displays a low level of ice-nucleation activity. Removal of approximately 92 kDa of carbohydrate from the 164-kDa antifreeze glycoprotein did not noticeably alter the antifreeze activity of the molecule, although it did diminish the ice-nucleation activity. This is the first report of an antifreeze protein that also is active as an ice-nucleation protein.Key words: antifreeze protein, plant growth promoting rhizobacteria, freezing tolerance, ice-nucleation protein.
Phenotypic identification ofAeromonasgenomospecies from clinical and environmental sources
Authors: Borrell N , Figueras M J , Guarro J
Published: April 2011

A collection of 983 Aeromonas isolates from environmental and clinical sources have been identified to the genomospecies level. A phenotypic method identified 93% of the strains. The use of citrate and the production of acid from sorbitol enabled the members of the Aeromonas hydrophila complex to be separated. The most common genomospecies from intestinal sources were Aeromonas veronii biotype sobria and Aeromonas caviae. The former, together with A. hydrophila,was the most frequently isolated species of extraintestinal origin. Most pathogenic species were very prevalent in environmental samples, with A. veronii biotype sobria being the most common in lakes and reservoirs (41.5%) and in treated drinking water (25.0%), and A. caviae was the most common in sea water (26.0%) and milk products (35.5%). Aeromonas hydrophila (18.1%) was the second most prevalent species isolated in untreated drinking water. Since Aeromonas infections are generally regarded as a water- and food-borne diseases, the high environmental prevalence of these pathogenic genomospecies should be regarded as an important threat to public health.Key words: Aeromonas, food, water, clinical, Spain.
Distribution of denitrification and nitrogen fixation genes inHyphomicrobiumspp. and other budding bacteria
Authors: Fesefeldt A , Kloos K , Bothe H , Lemmer H , Gliesche C G
Published: April 2011

Genomic DNA of Hyphomicrobium spp., Hirschia baltica ATCC 49814T, Hyphomonas oceanitis ATCC 33897T, and Pedomicrobium ferrugineum S-1290Twas investigated with gene probes specific for nitrate reductase (narG), cytochrome cd1containing nitrite reductase (nirS), Cu-containing nitrite reductase (nirK), nitrous oxide reductase (nosZ), ammonia monooxygenase (amoA), and nitrogenase reductase (nifH) by Southern or dot blot hybridization. The presence of denitrification genes could be demonstrated for Hyphomicrobium denitrificans 1869T, Hyphomicrobium aestuarii IFAM NQ-521GrT, Hyphomicrobium zavarzinii IFAM ZV-580, Hyphomicrobium zavarzinii subsp. chengduense, in the Hyphomicrobium DNA-DNA hybridization groups 3, 12, 13, 18, 26a1, 26a2, 26c, 26d, 26e, 26f, 26g2, and 29, and in three isolates from a denitrifying sand filter in a municipal wastewater treatment plant. The Cu-containing nitrite reductase appeared to be more frequent than the cytochrome cd1containing nitrite reductase in hyphomicrobia. Resulting positive DNA-DNA hybridization signals correlated with physiological activity measurements of intact cells in all cases determined. The nifH-like gene fragment was found in Hyphomicrobium aestuarii, Hyphomicrobium zavarzinii, Hyphomicrobium zavarzinii subsp. chengduense, Hyphomicrobium facilis and eight additional DNA-DNA hybridization groups. Ammonia monooxygenase was not genetically detected in any of the strains investigated. The results significantly extended the previous findings that genetically different hyphomicrobia of a sewage treatment plant and its adjacent receiving lake could occupy different ecological niches. Denitrification genes or the nifH-like gene fragment were not found in the other budding bacteria investigated.Key words: Hyphomicrobium, denitrification, nitrification, nitrogen fixation, activated sludge, biofilm.
Molecular characterization ofGluconobacter oxydans recAgene and its inhibitory effect on the function of the host wild-typerecAgene
Authors: Liu Yu-Tien , Chao Der-Chiang , Lee Fan , Chen Chia-Geun , Ji Dar-Der
Published: April 2011

A DNA fragment containing the recA gene of Gluconobacter oxydans was isolated and further characterized for its nucleotide sequence and ability to functionally complement various recA mutations. When expressed in an Escherichia coli recA host, the G. oxydans recA protein could efficiently function in homologous recombination and DNA damage repair. The recA gene's nucleotide sequence analysis revealed a protein of 344 amino acids with a molecular mass of 38 kDa. We observed an E. coli-like LexA repressor-binding site in the G. oxydans recA gene promoter region, suggesting that a LexA-like mediated response system may exist in G. oxydans. The expression of G. oxydans recA in E. coli RR1, a recA+strain, surprisingly caused a remarkable reduction of the host wild-type recA gene function, whereas the expression of both Serratia marcescens recA and Pseudomonas aeruginosa recA gene caused only a slight inhibitory effect on function of the host wild-typerecA gene product. Compared with the E. coli RecA protein, the identity of the amino acid sequence of G. oxydans RecA protein is much lower than those RecA proteins of both S. marcescens and Pseudomonas aeruginosa. This result suggests that the expression of another wild-type RecA could interfere with host wild-type recA gene's function, and the extent of such an interference is possibly correlated to the identity of the amino acid sequence between the two classes of RecA protein.Key words: Gluconobacter oxydans, recA gene, recombination, SOS function, interference.
Production of new extracellular glycolipids by a strain ofCellulomonas cellulans(Oerskovia xanthineolytica) and their structural characterization
Authors: Arino S , Marchal R , Vandecasteele J -P
Published: April 2011

Glycolipid-producing bacteria were isolated from soil samples. One of the strains, identified as Cellulomonas cellulans (Oerskovia xanthineolytica), was found to produce significant amounts of unusual extracellular glycolipids, which were shown to be composed of at least 11 individual compounds. Hydrolysis of the glycolipid mixture and gas chromatography - mass spectrometry analysis revealed the presence of fatty acids and hydroxy fatty acids ranging from C10to C18, 16 of which were identified. The glycidic moiety consisted of glucose, rhamnose, and ribose. The same sugars were found to be present in the cell wall of Cellulomonas cellulans, which also contained polar lipids including glycolipids. During strain cultivation, glycolipid excretion was stimulated when nitrogen was exhausted from the culture medium. In these conditions, the production in fermenters on glycerol, expressed in glucose equivalents, reached 8.9 g/L. Cell hydrophobicity, which rose to 95% during the growth phase, decreased to 50% during the production phase. The overall results show that the bacterial cell wall is involved in the synthesis of these new extracellular glycolipids.Key words: glycolipid, excretion, Cellulomonas cellulans, Oerskovia xanthineolytica, cell wall.
Evidence of self-inhibition by filamentous fungi accounts for unidirectional hyphal growth in colonies
Authors: Bottone Edward J , Nagarsheth Nimesh , Chiu Kenneth
Published: April 2011

Hyphal growth in filamentous fungi proceeds in an unidirectional radial pattern from a point inoculation. An inhibitor produced, secreted, and absorbed by the advancing hyphae has been speculated to account for directional growth. Working with Mucor and Aspergillus, laboratory evidence is provided for the production of an inhibitor by the advancing hyphae of these filmentous fungi that precludes back growth. Strains of Mucor and Aspergillus inoculated in agar on a 0.45-mm membrane filter placed on agar surfaces grow radially beyond the filter perimeter and onto the agar surface. Removal of the filter pad does not result in hyphal extension back into the clear zone underlying the membrane filter. Inoculation of the clear zone with a fresh Mucor or Aspergillus spore suspension does not result in hyphal growth. Instead, germinating spores show aberrant and aborted hyphae. The fungal inhibitor shows cross-reactivity between Mucor and Aspergillus, is resistant to pronase (10 mg/mL) but not chloroform inactivation, and passes through dialysis tubing with a molecular weight cutoff of 3500.Key words: fungal inhibitor, unidirectional hyphal growth, radial hyphal growth, Mucor, Aspergillus.
Phage display: applications, innovations, and issues in phage and host biology
Authors: Wilson Dan R , Finlay B Brett
Published: April 2011

In the 7 years since the first publications describing phage-displayed peptide libraries, phage display has been successfully employed in a variety of research. Innovations in vector design and methods to identify target clones account for much of this success. At the same time, not all ventures have been entirely successful and it appears that phage and host biology play important roles in this. A key issue concerns the role played by a displayed peptide or protein in its successful expression and incorporation into virions. While few studies have examined these issues specifically in context of phage display, the literature as a whole provides insight. Accordingly, we review phage biology, relevant aspects of host biology, and phage display applications with the goals of illustrating (i) relevant aspects of the interplay between phage-host biology and successful phage display and (ii) the limitations and considerable potential of this important technology.Key words: bacteriophage M13, phage display, pIII, pVIII, expression libraries.
The nonrandom binding distribution ofStreptococcus pneumoniaeto type II pneumocytes in culture is dependent on the relative distribution of cells among the phases of the cell cycle
Authors: Bérubé Luc R , Jouishomme Hervé , Jarrell Harold C
Published: April 2011

The adherence of Streptococcus pneumoniae to epithelial (A549) lung cells was studied and the bacterial binding distribution was found to be nonrandom (non-Gaussian). Analysis of the dependency of bacterial binding on the cell cycle of A549 cells revealed that approximately 1.8 times more bacteria bind to G2cells than to G0-G1phase cells. Furthermore, bacterial binding curves exhibited a plateau of binding to G2cells at a normalized bacteria to cell ratio approximately 1.8 times larger than that at which the plateau of binding to G0-G1cell was observed. Since G2cells are on average 1.4-1.8 times larger than G0-G1cells, the results indicate that bacterial binding is proportional to cell size and not to the preferential binding (higher affinity) of bacteria to A549 cells in the G2phase. Finally, the non-Gaussian distribution of bacterial binding could be mathematically modeled by a linear combination of three Gaussian distributions each representing bacterial binding to cells in a particular phase of the cell cycle (G0-G1, S, and G2-M). Because the Gaussian function contains a term that takes into account the relative number of cells in each of the phases, this last result implies that the overall (non-Gaussian) binding distribution (and hence the median of bacterial binding) can be highly sensitive to the relative proportion of cells in the various phases of the cell cycle.Key words: bacterial adherence, Streptococcus pneumoniae, flow cytometry, cell cycle.
Characterization of the bacterial community of a zinc-polluted soil
Authors: Brim H , Heuer H , Krögerrecklenfort E , Mergeay M , Smalla K
Published: April 2011

The bacterial community of a zinc-contaminated soil (Maatheide soil in Lommel, Belgium) was studied using cultivation as well as cultivation-independent techniques. Colony-forming units (CFU) were determined by plating on media with or without metals. Dominant isolates were characterized by fatty acid methyl ester analysis (FAME analysis) and PCR fingerprinting using repetitive extragenic palindromic sequences as primers. DNA was directly extracted from soil samples and used as a template for the PCR amplification of the 16S rDNA (8-1511) or a 16S rDNA fragment (968-1401). Clones resulting from cloning the 16S rDNA from soil DNA were sequenced. Temperature gradient gel electrophoresis (TGGE analysis) was performed for 16S rDNA fragments (968-1401) amplified from the dominant isolates, the clones, and the total soil DNA extracted according to two protocols differing in strength of lysis. Total CFU ranged from 104to 105/g soil. The majority of the isolates were identified by FAME analysis as Arthrobacter spp. (18 out of 23). None of the isolates were identified as a Ralstonia eutropha like strain (formerly Alcaligenes eutrophus). MetalloresistantRastomia eutropha like strains were previously shown to be dominant in the analyzed biotope. Most of the isolates were zinc tolerant but only seven could be considered zinc resistant. Sequences of the 16S rDNA clones obtained from total soil DNA were affiliated with genes of different bacteria such as alpha-proteobacteria, beta-proteobacteria, and the Cytophaga-Flexibacter-Bacteroides group. None of the sequenced clones aligned with the Ralstonia eutropha 16S rRNA gene. TGGE analysis of the 16S rDNA fragments (968-1401) amplified from the dominant strains, the clones, and the total soil DNA showed that isolates and clones represented only a part of the bands present in the TGGE pattern from total DNA. The 968-1401 fragment amplified from all Arthrobacter strains had a similar electrophoretic mobility. This band was seen as a major band in the pattern of DNA extracted from soil using a harsh cell lysis, whereas it did not appear, or appeared only as a weak band, in patterns obtained from soil DNA extracted using gentle lysis. The previously reported predominance of a Ralstonia eutropha like strain in this soil was no longer observed. This may suggest a population replacement by less resistant bacteria, concomitant with a progressive decrease of the zinc toxicity in the Maatheide soil.Key words: microbial community analysis, cultivation, 16S rDNA analysis, TGGE, sequencing, Zn-polluted soil.
Biodegradation of benzothiophene sulfones by a filamentous bacterium
Authors: Bressler David C , Leskiw Brenda K , Fedorak Phillip M
Published: April 2011

Previous studies showed that benzothiophene and 3- and 5-methylbenzothiophenes are oxidized by some bacteria to yield their corresponding sulfones, which were not subsequently degraded. In this study, a filamentous bacterium was isolated, which grew on each of these three sulfones as its sole carbon, sulfur, and energy source. Based on 16S rRNA gene sequencing and scanning electron microscopy, the isolate was found to belong to the genus Pseudonocardia and assigned the strain designation DB1. Benzothiophene sulfone and 3-methylbenzothiophene sulfone were more readily biodegraded than 5-methylbenzothiophene sulfone, and growth on these three compounds resulted in the release of 57, 62, and 28% of the substrate carbon as CO2, respectively. The thiophene ring was also cleaved, and between 44 and 88% of the sulfur from the consumed substrate was found as sulfate and (or) sulfite. Strain DB1 grew on benzoate, dibenzothiophene sulfone, and hexadecanoic acid, but it could not grow on benzofuran, dibenzothiophene, dibenzothiophene sulfoxide, hexadecane, indole, naphthalene, phenol, 2-sulfobenzoic acid, sulfolane, benzothiophene, or 3- or 5-methylbenzothiophenes. In addition, it did not oxidize the latter three compounds to their sulfones.Key words: benzothiophene sulfone, biodegradation, mineralization, sulfur heterocycles, Pseudonocardia.
Initiation of root growth stimulation by Azospirillum lipoferum CRT1 during maize seed germination
Authors: Jacoud Colette , Job Dominique , Wadoux Patrick , Bally René
Published: April 2011

Maize seeds were inoculated with a commercial inoculant containing 1.3 × 107 Azospirillum lipoferum CRT1 cells. After 24 or 48 h, bacteria were washed from the seed surface. Washed and unwashed seeds were then planted in pots containing perlite and grown for 28 days under greenhouse conditions. Whatever the density of Azospirillum at planting, the number of these bacteria at the end of the experiment was similar (1.9-8.0 × 107 bacteria·plant-1). However, comparison of root surface areas of the plants were different depending on the period of contact between seeds and the density of the inoculum. Twenty-four hours of contact was not sufficient to increase root growth surface areas. Contact for 48 h permitted us to obtain root surface areas comparable with those measured after a continuous contact. These results showed that in order to promote maize root surface areas, an optimal density of Azospirillum is not required during the whole cultural cycle. This optimal density is indispensable only up to the emergence of the radicle.Key words: Azospirillum, maize, inoculation, PGPR.
Stress tolerance in a yeast lipid mutant: membrane lipids influence tolerance to heat and ethanol independently of heat shock proteins and trehalose
Authors: Swan Tracey M , Watson Kenneth
Published: April 2011

The response of a yeast unsaturated fatty acid auxotroph, defective in Δ9-desaturase activity, to heat and ethanol stresses was examined. The most heat- and ethanol-tolerant cells had membranes enriched with oleic acid (C18:1), followed in order by cells enriched with linoleic (C18:2) and linolenic (C18:3) acids. Cells subjected to a heat shock (25-37°C for 30 min) accumulated trehalose and synthesized typical heat shock proteins. Although there were no obvious differences in protein profiles attributable to lipid supplementation of the mutant, relative protein synthesis as determined by densitometric analysis of autoradiograms suggested that hsp expression was different. However, there was no consistent relationship between the synthesis of heat shock proteins and the acquisition of thermotolerance in the lipid supplemented auxotroph or related wild type. Furthermore, trehalose accumulation was also not closely related to stress tolerance. On the other hand, the data presented indicated a more consistent role for membrane lipid composition in stress tolerance than trehalose, heat shock proteins, or ergosterol. We suggest that the sensitivity of C18:3-enriched cells to heat and ethanol may be attributable to membrane damage associated with increases in membrane fluidity and oxygen-derived free radical attack of membrane lipids.Key words: stress tolerance, yeast lipid mutant, membrane lipid unsaturation, trehalose, heat shock proteins.
Genetic and biochemical characterization of an exopolygalacturonase and a pectate lyase fromYersinia enterocolitica
Authors: Liao Ching-Hsing , Revear Larry , Hotchkiss Arland , Savary Brett
Published: April 2011

Yersinia enterocolitica, an invasive foodborne human pathogen, degrades polypectate by producing two depolymerizing enzymes, pectate lyase (PL) and polygalacturonase (PG). The gene encoding the PG activity, designated pehY, was located in a 3-kb genomic fragment of Y. enterocolitica ATCC 49397. The complete nucleotide sequence of this 3-kb fragment was determined and an open reading frame consisting of 1803 bp was predicted to encode a PG protein with an estimated Mrof 66 kDa and pI of 6.3. The amino acid sequence of prePG showed 59 and 43% identity to that of the exopolygalacturonase (exoPG) of Erwinia chrysanthemi and Ralstonia solanacearum, respectively. The Y. enterocolitica PG overproduced in Escherichia coli was purified to near homogeneity using perfusion cation exchange chromatography. Analysis of the PG depolymerization products by high performance anion-exchange chromatography and pulsed amperometric detection (HPAEC-PAD) revealed the exolytic nature of this enzyme. The Y. enterocolitica PL overproduced in E. coli was also partially purified and the Mrand pI were estimated to be 55 kDa and 5.2, respectively. HPAEC-PAD analysis of the PL depolymerization products indicated the endolytic nature of this enzyme. Southern hybridization analyses revealed that pehY and pel genes of Y. enterocolitica are possibly encoded in the chromosome rather than in the plasmid. Purified exopolygalacturonase (over 10 activity units) was unable to macerate plant tissues.Key words: pectinase activities, human pathogen, HPLC analysis, pehY gene.