bluuebunny/crossref_metadata_2025 huggingface dataset:
Full-scale studies of factors related to coliform regrowth in drinking water
Authors: LeChevallier M W , Welch N J , Smith D B
Published: January 2020

An 18-month survey of 31 water systems in North America was conducted to determine the factors that contribute to the occurrence of coliform bacteria in drinking water. The survey included analysis of assimilable organic carbon (AOC), coliforms, disinfectant residuals, and operational parameters. Coliform bacteria were detected in 27.8% of the 2-week sampling periods and were associated with the following factors: filtration, temperature, disinfectant type and disinfectant level, AOC level, corrosion control, and operational characteristics. Four systems in the study that used unfiltered surface water accounted for 26.6% of the total number of bacterial samples collected but 64.3% (1,013 of 1,576) of the positive coliform samples. The occurrence of coliform bacteria was significantly higher when water temperatures were > 15 degrees C. For filtered systems that used free chlorine, 0.97% of 33,196 samples contained coliform bacteria, while 0.51% of 35,159 samples from chloraminated systems contained coliform bacteria. The average density of coliform bacteria was 35 times higher in free-chlorinated systems than in chloraminated water (0.60 CFU/100 ml for free-chlorinated water compared with 0.017 CFU/100 ml for chloraminated water). Systems that maintained dead-end free chlorine levels of < 0.2 mg/liter or monochloramine levels of < 0.5 mg/liter had substantially more coliform occurrences than systems that maintained higher disinfectant residuals. Free-chlorinated systems with AOC levels greater than 100 micrograms/liter had 82% more coliform-positive samples and 19 times higher coliform levels than free-chlorinated systems with average AOC levels less than 99 micrograms/liter. Systems that maintained a phosphate-based corrosion inhibitor and limited the amount of unlined cast iron pipe had fewer coliform bacteria. Several operational characteristics of the treatment process or the distribution system were also associated with increased rates of coliform occurrence. The study concludes that the occurrence of coliform bacteria within a distribution system is dependent upon a complex interaction of chemical, physical, operational, and engineering parameters. No one factor could account for all of the coliform occurrences, and one must consider all of the parameters described above in devising a solution to the regrowth problem.
Oxidative stress detection with Escherichia coli harboring a katG'::lux fusion
Authors: Belkin S , Smulski D R , Vollmer A C , Van Dyk T K , LaRossa R A
Published: January 2020

A plasmid containing a transcriptional fusion of the Escherichia coli katG promoter to a truncated Vibrio fischeri lux operon (luxCDABE) was constructed. An E. coli strain bearing this plasmid (strain DPD2511) exhibited low basal levels of luminescence, which increased up to 1,000-fold in the presence of hydrogen peroxide, organic peroxides, redox-cycling agents (methyl viologen and menadione), a hydrogen peroxide-producing enzyme system (xanthine and xanthine oxidase), and cigarette smoke. An oxyR deletion abolished hydrogen peroxide-dependent induction, confirming that oxyR controlled katG'::lux luminescence. Light emission was also induced by ethanol by an unexplained mechanism. A marked synergistic response was observed when cells were exposed to both ethanol and hydrogen peroxide; the level of luminescence measured in the presence of both inducers was much higher than the sum of the level of luminescence observed with ethanol and the level of luminescence observed with hydrogen peroxide. It is suggested that this construction or similar constructions may be used as a tool for assaying oxidant and antioxidant properties of chemicals, as a biosensor for environmental monitoring and as a tool for studying cellular responses to oxidative hazards.
Distribution of Pseudomonas syringae pathovars into twenty-three O serogroups
Authors: Saunier M , Malandrin L , Samson R
Published: January 2020

Serological reactions of Pseudomonas syringae and Pseudomonas viridiflava were studied by Ouchterlony double diffusion. A total of 55 polyclonal antisera, containing anti-lipopolysaccharide (anti-LPS) precipitating antibodies, were cross-tested against antigenic suspensions of 51 strains. Twenty-three O serogroups were defined, primarily on the reaction of the type strains. Two families of O serogroups showed antigenic crossreactivities (PHA, MOP1, MOP2, MOP3, HEL1, HEL2, and SYR1; PERSAVTOM1, PERSAVTOM2, DEL, POR, and SYR2). Ten O serogroups showed a clearcut specificity: APTPIS, TAB, VIR1, VIR2, VIR3, SYR3, SYR4, SYR5, HUS, and LAC. The last serogroup (RIB) contained strains with rough colony morphology and side chain-deficient LPSs, as evidenced by sodium dodecyl sulfate-polycrylamide gel electrophoresis. The LPS basis of the O serogroups was demonstrated by immunoblotting. Serological reference strains were designated for all of the O serogroups and correspondence was established between the O serogroups studied and seven previous serogroups (L. T. Pastushenko and I.D. Simonovich, Mikrobiol, Zh. 41:222-229 and 330-339, 1979). A total of 355 strains of P. syringae (sensu lato) belonging to 15 pathovars, not including pathovar syringae, were typed into the 23 described O serogroups. O serogroups were assigned after double-diffusion reactions, with each strain compared with serological references. The utility of O serogrouping to study P. syringae pathovar structure and diversity is discussed.
Strain typing with IS200 fingerprints in Salmonella abortusovis
Authors: Schiaffino A , Beuzón C R , Uzzau S , Leori G , Cappuccinelli P , Casadesús J , Rubino S
Published: January 2020

A collection of Salmonella abortusovis isolates was examined for the presence of insertion element IS200. All proved to contain three or four copies of the element. One IS200 hybridization band of approximately 9 kb was found in all isolates, indicating that all S. abortusovis strains carry an IS200 element in similar or identical locations; this band can be potentially useful for serovar identification. S. abortusovis collection isolates from distinct geographic areas were highly polymorphic, suggesting that IS200 fingerprints might provide information on the geographic origin of S. abortusovis strains. Isolates obtained from the same geographic area (the island of Sardinia, Italy) were less polymorphic: all shared three constant IS200 hybridization bands, indicating that they derive from a single ancestor. Most strains analyzed contained an additional copy of IS200 in the variable region of the virulence plasmid. Certain Sardinian flocks proved to be infected by only one S. abortusovis strain, while others harbored two strains. Strain typing with IS200 fingerprints proved to be more reliable than plasmid analysis, because the latter yielded a high degree of polymorphism, even among isolates from the same flock.
Dissolution of Barium from Barite in Sewage Sludges and Cultures of Desulfovibrio desulfuricans
Authors: Baldi F , Pepi M , Burrini D , Kniewald G , Scali D , Lanciotti E
Published: January 2020

High concentrations of total barium, ranging from 0.42 to 1.58 mg(middot)g(sup-1) (dry weight) were found in sludges of two sewage treatment plants near Florence, Italy. Barium concentrations in the suspended matter decreased as redox potential values changed from negative to positive. An anoxic sewage sludge sample was aerated, and 30% of the total barium was removed in 24 h. To demonstrate that barium was solubilized from barite by sulfate-reducing bacteria, a strain of Desulfovibrio desulfuricans was used to study the solubilization of barium from barite under laboratory conditions. During cell growth with different concentrations of barite from 0.01 to 0.3 g(middot)liter(sup-1) (the latter is the MIC) as the only source of sulfates in the cultures, the D. desulfuricans strain accumulated barium up to 0.58 (mu)g(middot)mg(sup-1) (dry weight). Three times the quantity of barium was dissolved by bacteria than in the uninoculated medium (control). The unexpectedly low concentration of soluble barium (1.2 mg of Ba(middot)liter(sup-1)) with respect to the quantity expected (109 mg of Ba(middot)liter(sup-1)), calculated on the basis of the free H(inf2)S evolved from the dissimilatory reduction of sulfate from barite, was probably due to the formation of other barium compounds, such as witherite (BaCO(inf3)) and the transient species barium sulfide (BaS). The D. desulfuricans strain, growing on barite, formed visible aggregates. Confocal microscopy analysis showed that aggregates consisted of bacteria and barite. After 3 days of incubation, several autofluorescent crystals surrounded by a dissolution halo were observed. The crystals were identified as BaS by comparison with the commercial compound.
A computer-simulated restriction fragment length polymorphism analysis of bacterial small-subunit rRNA genes: efficacy of selected tetrameric restriction enzymes for studies of microbial diversity in nature
Authors: Moyer C L , Tiedje J M , Dobbs F C , Karl D M
Published: January 2020

An assessment of 10 tetrameric restriction enzymes (TREs) was conducted by using a computer-simulated restriction fragment length polymorphism (RFLP) analysis for over 100 proximally and distally related bacterial small-subunit (SSU) rRNA gene sequences. Screening SSU rDNA clone libraries with TREs has become an effective strategy because of logistic simplicity, commercial availability, and economy. However, the rationale for selecting the type and number of TREs has not been systematically evaluated. Our objective was to identify the optimal combination of TREs for RFLP screening of cloned SSU rRNA genes from undefined bacterial clone libraries. After computer-simulated TRE digestion, the resultant fragments were categorized on the basis of the frequency of different restriction fragment size classes. Three groups of distribution patterns for the TREs were determined and further examined via graphical exploratory data analysis. The RFLP size-frequency distribution data for each group of enzymes were then used to infer phylogenetic relationships via the neighbor-joining method. The resulting bootstrap values and the correct placement of node bifurcations were used as additional criteria to evaluate the efficacy of the selected TREs. These RFLP data were compared with known phylogenetic relationships based on SSU rRNA sequence analysis as defined by the Ribosomal Database Project. A heuristic approach testing random combinations of TREs showed that three or more TRE combinations detected > 99% of the operational taxonomic units (OTUs) within the model data set. OTUs that remained undetected after three TRE treatments had a median sequence similarity of 96.1%. Of the 10 restriction enzymes examined, HhaI, RsaI, and BstUI (group 3) were the most efficacious at detecting and differentiating bacterial SSU rRNA genes on the basis of their ability to correctly classify OTUs. Group 3 TREs are therefore recommended for screening in studies using bacterial SSU rRNA genes as descriptors of in situ microbial diversity.
Fate of enterohemorrhagic Escherichia coli O157:H7 in bovine feces
Authors: Wang G , Zhao T , Doyle M P
Published: January 2020

Dairy cattle have been identified as a principal reservoir of Escherichia coli O157:H7. The fate of this pathogen in bovine feces at 5, 22, and 37 degrees C was determined. Two levels of inocula (10(3) and 10(5) CFU/g) of a mixture of five nalidixic acid-resistant E. coli O157:H7 strains were used. E. coli O157:H7 survived at 37 degrees C for 42 and 49 days with low and high inocula, respectively, and at 22 degrees C for 49 and 56 days with low and high inocula, respectively. Fecal samples at both temperatures had low moisture contents (about 10%) and water activities ( < 0.5) near the end of the study. E. coli O157:H7 at 5 degrees C survived for 63 to 70 days, with the moisture content (74%) of feces remaining high through the study. Chromosomal DNA fingerprinting of E. coli O157:H7 isolates surviving near the completion of the study revealed that the human isolate strain 932 was the only surviving strain at 22 or 37 degrees C. All five strains were isolated near the end of incubation from feces held at 5 degrees C. Isolates at each temperature were still capable of producing both verotoxin 1 and verotoxin 2. Results indicate that E. coli O157:H7 can survive in feces for a long period of time and retain its ability to produce verotoxins. Hence, bovine feces are a potential vehicle for transmitting E. coli O157:H7 to cattle, food, and the environment. Appropriate handling of bovine feces is important to control the spread of this pathogen.
Linkage among genes responsible for fumonisin biosynthesis in Gibberella fujikuroi mating population A
Authors: Desjardins A E , Plattner R D , Proctor R H
Published: January 2020

Most naturally occurring strains of the fungus Gibberella fujikuroi mating population A produce high levels of the mycotoxin fumonisin B1 (FB1), which is oxygenated at both carbons C-5 and C-10. Some strains, however, produce only FB2 or FB3, suggesting that they lack the ability to hydroxylate position C-10 or C-5, respectively. Genetic analysis indicates that these different phenotypes are due to single gene defects at closely linked loci designated fum2 and fum3. Further allellism tests indicate that both fum2 and fum3 are closely linked to fum1, a previously identified gene that regulates fumonisin production. The recovery frequency of FB1-producing progency from cross 510 between fum1 and fum2 mutations suggests a map distance of approximately 6.2 cM between these two loci. Amplified fragment length polymorphism analysis of parents and progeny of cross 510 was employed to confirm that the FB1-producing strains are recombinant progeny. We conclude that fum1, fum2, and fum3 constitute a fumonisin biosynthetic gene cluster on chromosome 1 of the restriction fragment length-map of G. fujikuroi.
Bacterial enrichment at the gas-water interface of a laboratory apparatus
Authors: Powelson D K , Mills A L
Published: January 2020

The gas-water interface (GWI) is likely to have important effects on bacterial adsorption and transport in unsaturated porous media. A glass apparatus that isolated GWIs in ports above a flowthrough suspension of a groundwater bacterial isolate was used to represent unsaturated porous media. The surface microlayer was collected by placing a polycarbonate filter on the GWI. The filter was stained, and the bacteria were enumerated by direct count. The significance of five independent variables on the surface density of cells (s, in cells per square millimeter) was determined by nonlinear multiple regression. Three of the variables were shown to be significant: surfactant concentration (d), time (t), and bulk bacterial concentration (B). The surface density decreased with increasing d and increased with increasing t and B. When surfactant was absent, the GWI became highly enriched in bacteria. For example, when d = 0, 48 h < t < 72 h, and 5,000 cells mm(sup-3) < B < 10,000 cells mm(sup-3), s averaged 3.0 x 10(sup4) cells mm(sup-2). This surface density occupied about 6.0% of the GWI, and the surface microlayer concentration of cells was 190 times the bulk concentration. The other two variables: pH (p) and ionic strength (I) were shown to be insignificant. The strong effect of d and the lack of effect of I and p support the hypothesis that hydrophobic interaction dominates bacterial adsorption to the GWI.
Comamonas testosteroni colony phenotype influences exopolysaccharide production and coaggregation with yeast cells
Authors: Bossier P , Verstraete W
Published: January 2020

A Comamonas testosteroni strain was isolated from activated sludge on the basis of its ability to coaggregate with yeast cells. On agar plates the following two types of colonies were formed: colonies with a mucoid appearance and colonies with a nonmucoid appearance. On plates this strain alternated between the two forms, making sectored colonies. In liquid medium with constant agitation no such change was observed. In the absence of agitation and in contact with a glass surface a culture with predominantly nonmucoid-colony-forming cells very rapidly shifted to a culture dominated by mucoid-colony-forming cells. In liquid medium the reverse was observed under stress conditions imposed by hydrogen peroxide, sodium dodecyl sulfate, or starvation. Nonmucoid cells formed very rapidly settling flocs with yeast cells, while coaggregation of mucoid cells with yeast cells did not occur. These findings may be relevant to the behavior of activated sludge microbial communities.
Four unnamed species of nonsymbiotic rhizobia isolated from the rhizosphere of Lotus corniculatus
Authors: Sullivan J T , Eardly B D , van Berkum P , Ronson C W
Published: January 2020

Previously, we found that genetically diverse rhizobia nodulating Lotus corniculatus at a field site devoid of naturalized rhizobia had symbiotic DNA regions identical to those of ICMP3153, the inoculant strain used at the site (J. T. Sullivan, H. N. Patrick, W. L. Lowther, D. B. Scott, and C. W. Ronson, Proc. Natl. Acad. Sci. USA 92:8985-8989, 1995). In this study, we characterized seven nonsymbiotic rhizobial isolates from the rhizosphere of L. corniculatus. These included two from plants at the field site sampled by Sullivan et al. and five from plants at a new field plot adjacent to that site. The isolates did not nodulate Lotus species or hybridize to symbiotic gene probes but did hybridize to genomic DNA probes from Rhizobium loti. Their genetic relationships with symbiotic isolates obtained from the same sites, with inoculant strain ICMP3153, and with R. loti NZP2213T were determined by three methods. Genetic distance estimates based on genomic DNA-DNA hybridization and multilocus enzyme electrophoresis were correlated but were not consistently reflected by 16S rRNA nucleotide sequence divergence. The nonsymbiotic isolates represented four genomic species that were related to R. loti; the diverse symbiotic isolates from the site belonged to one of these species. The inoculant strain ICMP3153 belonged to a fifth genomic species that was more closely related to Rhizobium huakuii. These results support the proposal that nonsymbiotic rhizobia persist in soils in the absence of legumes and acquire symbiotic genes from inoculant strains upon introduction of host legumes.
The Bacillus thuringiensis insecticidal toxin binds biotin-containing proteins
Authors: Du C , Nickerson K W
Published: January 2020

Brush border membrane vesicles from larvae of the tobacco hornworm, Manduca sexta, contain protein bands of 85 and 120 kDa which react directly with streptavidin conjugated to alkaline phosphatase. The binding could be prevented either by including 10 microM biotin in the reaction mixture or by prior incubation of the brush border membrane vesicles with an activated 60- to 65-kDa toxin from Bacillus thuringiensis HD-73. The ability of B. thuringiensis toxins to recognize biotin-containing proteins was confirmed by their binding to pyruvate carboxylase, a biotin-containing enzyme, as well as to biotinylated ovalbumin and biotinylated bovine serum albumin but not to their nonbiotinylated counterparts. Activated HD-73 toxin also inhibited the enzymatic activity of pyruvate carboxylase. The biotin binding site is likely contained in domain III of the toxin. Two highly conserved regions within domain III are similar in sequence to the biotin binding sites of avidin, streptavidin, and a biotin-specific monoclonal antibody. In particular, block 4 of the B. thuringiensis toxin contains the YAS biotin-specific motif. On the basis of its N-terminal amino acid sequence, the 120-kDa biotin-containing protein is totally distinct from the 120-kDa aminopeptidase N reported to be a receptor for Cry1Ac toxin.
Distribution of class II transposase and resolvase genes in soil bacteria and their association with mer genes
Authors: Pearson A J , Bruce K D , Osborn A M , Ritchie D A , Strike P
Published: January 2020

Southern hybridization was performed on 30 gram-negative, mercury-resistant soil bacteria isolated from three terrestrail sites in Great Britain; two of these sites were mercury polluted (SO and SE), and one was pristine (SB). Most of the isolates (20 of 30) hybridized to probes encoding regions of the transposase (tnpA) and resolvase (tnpR) genes from Tn501 and Tn21. Isolates SE9 and SB3 hybridized to the Tn21 but not the Tn501 tnpA probe; however, they differed in that SB3 hybridized to both Tn501 and Tn21 tnpR probes while SE9 did not hybridize to either tnpR probe. The remaining isolates (7 of 30) did not hybridize to any of the transposon gene probes under the conditions used. tnpA and tnpR regions were PCR amplified from most of the hybridizing isolates and from Tn501 and Tn21, and variation was assessed by restriction fragment length polymorphism analysis. On the basis of these data, tnpA regions were divided into eight restriction fragment length polymorphism classes and tnpR regions were divided into five classes. Similarity coefficients were calculated between classes and used to construct dendrograms showing percent similarity. A compilation of the data from this study on tnpA and tnpR regions and a previous study on merRT delta P regions (A. M. Osborn, K. D. Bruce, P. Strike, and D. A. Ritchie, Appl. Environ. Microbiol. 59:4024-4030, 1993) indicates the presence of hybrid transposons and provides evidence for extensive recombination, both between transposon genes and between transposon and mer genes, within these natural populations of bacteria.
Influence of aroclor 1242 concentration on polychlorinated biphenyl biotransformations in hudson river test tube microcosms
Authors: Fish K M
Published: January 2020

When 93.3 to 933 (mu)mol of Aroclor 1242 per kg was added to Hudson River sediment test tube microcosms, the rates of polychlorinated biphenyl biotransformations increased with increasing Aroclor 1242 concentration after a 4- to 8-week acclimation period. In contrast, when 37.3 (mu)mol of Aroclor 1242 per kg was added, polychlorinated biphenyl biotransformations occurred at slow constant rates.
Polymorphic restriction patterns of ribosomal internal transcribed spacers in the biocontrol fungus Puccinia carduorum correlate with weed host origin
Authors: Berthier Y T , Bruckart W L , Chaboudez P , Luster D G
Published: January 2020

The internal transcribed spacer (ITS) regions of ribosomal DNA were amplified by PCR and used to develop genetic markers for isolates of Puccinia carduorum being evaluated for biological control of Carduus thoermeri (musk thistle). Unique patterns were produced upon restriction of ITS DNA amplified from four separate Puccinia spp. Restriction patterns of ITS DNA of isolates of P. carduorum from Carduus acanthoides and C. thoermeri were distinct from those of P. carduorum from Carduus tenuiflorus and Carduus pycnocephalus. By this technique, isolates of P. carduorum from four different weed hosts can be differentiated from other Puccinia spp. and separated into two host groups.
Kinetic Analyses of Desulfurization of Dibenzothiophene by Rhodococcus erythropolis in Continuous Cultures
Authors: Wang P , Humphrey A E , Krawiec S
Published: January 2020

Rhodococcus erythropolis N1-36, a desulfurization strain, was grown in continuous culture at 10 different dilution rates with 50 (mu)M dibenzothiophene sulfone (DBTO(inf2)) as the growth-limiting nutrient. The steady-state biomass, concentrations of substrate (DBTO(inf2)) and product (monohydroxybiphenyl), saturation constant (0.39 (mu)M DBTO(inf2)), and cell yield coefficient (9 mg of biomass(middot)(mu)M(sup-1) DBTO(inf2)) were measured. Continuous cultures at five temperatures allowed calculation of activation energy (0.84 kcal(middot)mol(sup-1) [ca. 3.5 kJ(middot)mol(sup-1)]) near the optimal temperature (30(deg)C) for growth. A washout technique was used to calculate the maximum specific growth rate (0.235 h(sup-1)), a value equivalent to a minimum generation time of 2.95 h.
Isolation and characterization of isopimaric acid-degrading bacteria from a sequencing batch reactor
Authors: Wilson A E , Moore E R , Mohn W W
Published: January 2020

We isolated two aerobic, gram-negative bacteria which grew on the diterpene resin acid isopimaric acid (IpA) as the sole carbon source and electron donor. The source of the isolates was a sequencing batch reactor treating a high-strength process stream from a paper mill. The isolates, IpA-1 and IpA-2, also grew on pimaric and dehydroabietic acids, and IpA-1 grew on abietic acid. Both strains used fatty acids, but neither strain used camphor, sitosterol, or betulin. Strain IpA-1 grew anaerobically with nitrate as an electron acceptor. Strains IpA-1 and IpA-2 had growth yields of 0.19 and 0.23 g of protein per g of IpA, respectively. During growth, both strains transformed IpA carbon to approximately equal amounts of biomass, carbon dioxide, and dissolved organic carbon. In both strains, growth on IpA induced an enzymatic system which caused cell suspensions to transform all four of the above resin acids. Cell suspensions of IpA-1 and IpA-2 removed IpA at rates of 0.56 and 0.13 mumol mg of protein-1 h-1, respectively. Cultures and cell suspensions of both strains failed to completely consume pimaric acid and yielded small amounts of an apparent metabolite from this acid. Cultures and cell suspensions of both strains yielded large amounts of three apparent metabolites from dehydroabietic acid. Analysis of 16S rDNA sequences indicated that the isolates are distinct members of the genus Pseudomonas sensu stricto.
Purification and partial characterization of a novel antibacterial agent (Bac1829) Produced by Staphylococcus aureus KSI1829
Authors: Crupper S S , Iandolo J J
Published: January 2020

A novel antimicrobial agent from Staphylococcus aureus KSI1829, designated Bac1829, was purified by sequential steps of ammonium sulfate precipitation, Sephadex G-50 gel filtration chromatography, and hydrophobic interaction chromatography. Purified Bac1829 has a molecular mass of 6,418 +/- 2 Da. The peptide in heat stable, since full biological activity is retained after heating at 95 degrees C for 15 min, and it is destroyed by digestion with proteases. Amino acid sequence analysis revealed a high concentration of Ala and Gly residues, which respectively comprised 24 and 19% of the total amino acid content. Additionally, high levels of hydrophobic amino acids were present, accounting for the hydrophobic nature of Bac1829. Purified Bac1829 killed exponentially growing Corynebacterium renale in a dose-dependent manner by a bactericidal mode of action. A partial inhibitory spectrum analysis revealed that the following organisms were sensitive to the inhibitory activity of Bac1829: S. aureus RN4220, Streptococcus suis, Corynebacterium pseudotuberculosis, C. renale, Corynebacterium diptheriae, Haemophilus parasuis, Bordetella pertussis, Bordetella bronchoseptica, Moraxella bovis, and Pasteurella multocida.
Evaluation of colicins for inhibitory activity against diarrheagenic Escherichia coli strains, including serotype O157:H7
Authors: Murinda S E , Roberts R F , Wilson R A
Published: January 2020

Twenty-four Escherichia coli strains producing standard colicins were evaluated for inhibitory activity against 27 diarrheagenic E. coli strains of serotypes O15:H-, O26:(H11, H-), and O111:(H8, H11, H-), including O157:H7, representing diarrheagenic E. coli clones, 3, 4, 8, 9, and 10. Overlay techniques were used to assess inhibition on Luria agar and Luria agar supplemented with 0.25 micrograms of mitomycin C per ml to induce colicin production. As a group, the A colicins (Col) E1 to E8, K, and N inhibited 23 to 25 (85.2 to 92.6%) of the 27 diarrheagenic strains on mitomycin C-containing agar, whereas the most active group B colicins, Col D and Ia, inhibited 9 and 12 (33.3 and 44.4%), of the diarrheagenic strains, respectively. Col G and H and Mcc B17 inhibited 22 to 27 (81.5 to 100%) of the diarrheagenic strains on Luria agar but were suppressed on mitomycin C-containing agar medium. All O157:H7 strains evaluated were sensitive to Col E1 to E8, K, and N on mitomycin C-containing agar and to Col G and H and Mcc B17 on Luria agar. Sensitivity to colicins of the selected set of diarrheagenic strains was in the order diarrheagenic E. coli clone 9 > 4 > 3 > 10 > 8 and was not restricted to strains of a single clone or serotype. Strain 8C from clone 8 was resistant to most test colicins. There is potential for using colicins in foods and agriculture to inhibit sensitive diarrheagenic E. coli strains, including serotype O157:H7.
Transformation Kinetics of Chlorinated Ethenes by Methylosinus trichosporium OB3b and Detection of Unstable Epoxides by On-Line Gas Chromatography
Authors: van Hylckama V J , de Koning W , Janssen D B
Published: January 2020

A rapid and accurate method for the determination of transformation kinetics of volatile organic substrates was developed. Concentrations were monitored by on-line gas chromatographic analysis of the headspace of well-mixed incubation mixtures. With this method, the kinetics of transformation of a number of C(inf1) and C(inf2) halogenated alkanes and alkenes by Methylosinus trichosporium OB3b expressing particulate methane monooxygenase or soluble methane monooxygenase (sMMO) were studied. Apparent specific first-order rate constants for cells expressing sMMO decreased in the order of dichloromethane, vinyl chloride, cis-1,2-dichloroethene, trans-1,2-dichloroethene, 1,1-dichloroethene, trichloroethene, chloroform, and 1,2-dichloroethane. During the degradation of trichloroethene, cis-1,2-dichloroethene, trans-1,2-dichloroethene, and vinyl chloride, the formation of the corresponding epoxides was observed. The epoxide of vinyl chloride and the epoxide of trichloroethene, which temporarily accumulated in the medium, were chemically degraded according to first-order kinetics, with half-lives of 78 and 21 s, respectively. Cells expressing sMMO actively degraded the epoxide of cis-1,2-dichloroethene but not the epoxide of trans-1,2-dichloroethene. Methane and acetylene inhibited degradation of the epoxide of cis-1,2-dichloroethene, indicating that sMMO was involved.
Evidence for two domains of growth temperature for the psychrotrophic bacterium Pseudomonas fluorescens MF0
Authors: Guillou C , Guespin-Michel J F
Published: January 2020

The variations in the maximal specific growth rate of the psychrotrophic bacterium Pseudomonas fluorescens MF0 with respect to temperature were studied between 0 and 30 degrees C (optimal for growth). The Arrhenius plot showed a drastic change in slope at the intermediate temperature of 17 degrees C. Over the cold domain from 0 to 17 degrees C, the temperature characteristic was twofold higher than over the suboptimal domain from 17 to 30 degrees C. The macromolecular composition of exponentially growing cells was invariant over the entire range from 0 to 30 degrees C. Variations of temperature and growth rate were independently investigated through chemostat experiments in order to characterize their respective effects on cell macromolecular composition and size. The effect of growth rate in this psychrotrophic strain is identical to that of all other bacteria assayed so far. In contrast, an original biphasic variation of total protein concentration was demonstrated in strain MF0 with respect to temperature, with a maximum at 17 to 20 degrees C. Indeed, increasing the temperature in the chemostat resulted in a biphasic decrease in the net protein production rate: a very slight decrease below 17 degrees C and a much larger decrease from 17 to 28 degrees C. These results could signify an increase in the cellular protein degradation rate with increasing temperature, especially above 17 degrees C.
Regio- and stereospecific oxidation of 9,10-dihydroanthracene and 9,10-dihydrophenanthrene by naphthalene dioxygenase: structure and absolute stereochemistry of metabolites
Authors: Resnick S M , Gibson D T
Published: January 2020

The oxidation of 9,10-dihydroanthracene and 9,10-dihydrophenanthrene was examined with mutant and recombinant strains expressing naphthalene dioxygenase from Pseudomonas putida (NCIB 9816.4. Salicylate-induced cells of P. putida strain 9816/11 and isopropylthiogalactopyranoside-induced cells of Escherichia coli JM109(DE3)(pDTG141) oxidized 9,10-dihydroanthracene to (+)-cis-1R,2S)-1,2-dihydroxy-1,2,9,10-tetrahydroanthracene (> 95% relative yield; > 95% enantiomeric excess) as the major product. 9-Hydroxy-9,10-dihydroanthracene (< 5% relative yield) was a minor product formed by both organisms. The same cells oxidized 9,10-dihydrophenanthrene to (+)-cis-(3S,4R)-3,4-dihydroxy-3,4,9,10-tetrahydrophenanthrene (70% relative yield; > 95% enantiomeric excess) and (+)-(S)-9-hydroxy-9,10-dihydrophenanthrene (30% relative yield). The major reaction catalyzed by naphthalene dioxygenase with 9,10-dihydroanthracene and 9,10-dihydrophenanthrene was stereospecific dihydroxylation in which both of the previously undescribed cis-diene diols were of R configuration at the benzylic center adjacent to the bridgehead carbon atom. The results suggest that for benzocylic substrates, the location of benzylic carbons influences the type of reaction(s) catalyzed by naphthalene dioxygenase.
Anaerobically controlled expression system derived from the arcDABC operon of Pseudomonas aeruginosa: application to lipase production
Authors: Winteler H V , Schneidinger B , Jaeger K E , Haas D
Published: January 2020

The anaerobically inducible arcDABC operon encodes the enzymes of the arginine deiminase pathway in Pseudomonas aeruginosa. Upon induction, the arcAB mRNAs and proteins reach high intracellular levels, because of a strong anaerobically controlled promoter and mRNA processing in arcD, leading to stable downstream transcripts. We explored the usefulness of this system for the construction of expression vectors. The lacZ gene of Escherichia coli was expressed to the highest levels when fused close to the arc promoter. Insertion of lacZ further downstream into arcA or arcB did not stabilize the intrinsically unstable lacZ mRNA. On the contrary, lacZ mRNA appeared to be a vulnerable endonuclease target destabilizing arcAB mRNAs in the 5'-to-3' direction in P. aeruginosa. The native arc promoter was modified for optional expression in the -10 sequence and in the -40 region, which is a binding site for the anaerobic regulator ANR. In P. aeruginosa grown either anaerobically or with oxygen limitation in unshaken cultures, this promoter was stronger than the induced tac promoter. The P. aeruginosa lipAH genes, which encode extracellular lipase and lipase foldase, respectively, were fused directly to the modified arc promoter in an IncQ vector plasmid. Semianaerobic static cultures of P. aeruginosa PAO1 carrying this recombinant plasmid overproduced extracellular lipase 30-fold during stationary phase compared with the production by strain PAO1 without the plasmid. Severe oxygen limitation, in contrast, resulted in poor lipase productivity despite effective induction of the ANR-dependent promoter, suggesting that secretion of active lipase is blocked by the absence of oxygen. In conclusion, the modified arc promoter is useful for driving the expression of cloned genes in P. aeruginosa during oxygen-limited growth and stationary phase.
Utility of microcosm studies for predicting phylloplane bacterium population sizes in the field
Authors: Kinkel L L , Wilson M , Lindow S E
Published: January 2020

Population sizes of two ice nucleation-active strains of Pseudomonas syringae were compared on leaves in controlled environments and in the field to determine the ability of microcosm studies to predict plant habitat preferences in the field. The P. syringae strains investigated were the parental strains of recombinant deletion mutant strains deficient in ice nucleation activity that had been field tested for their ability to control plant frost injury. The population size of the P. syringae strains was measured after inoculation at three field locations on up to 40 of the same plant species that were studied in the growth chamber. There was seldom a significant relationship between the mean population size of a given P. syringae strain incubated under either wet or dry conditions in microcosms and the mean population size which could be recovered from the same species when inoculated in the field. Specifically, on some plant species, the population size recovered from leaves in the field was substantially greater than from that species in a controlled environment, while for other plant species field populations were significantly smaller than those observed under controlled conditions. Population sizes of inoculated P. syringae strains, however, were frequently highly positively correlated with the indigenous bacterial population size on the same plant species in the field, suggesting that the ability of a particular plant species to support introduced bacterial strains is correlated with its ability to support large bacterial populations or that indigenous bacteria enhance the survival of introduced strains. Microcosm studies therefore seem most effective at assessing possible differences between parental and recombinant strains under a given environmental regime but are limited in their ability to predict the specific population sizes or plant habitat preferences of bacteria on leaves under field conditions.
Typing of bovine attaching and effacing Escherichia coli by multiplex in vitro amplification of virulence-associated genes
Authors: China B , Pirson V , Mainil J
Published: January 2020

Attaching and effacing Escherichia coli is a new causal agent of diarrhea in calves. Its major virulence factors are the intimin protein, encoded by the eaeA gene, and the Shiga-like toxins, encoded by slt genes. Because the sequences of these genes are available, we selected specific primers to amplify each virulence gene so as to develop a new identification test based on multiplex amplification of virulence-associated genes. Of 30 tested strains, 14 were eaeA+, 15 were eaeA+ slt-I+, 1 was eaeA+ slt-I+ slt-II+, and 1 was eaeA+ slt-II+. The method proved in our hands to be fast and specific and in perfect correlation with the hybridization method.
Overproduction of recombinant ribulose 1,5-bisphosphate carboxylase/oxygenase from Synechococcus sp. strain PCC6301 in glucose-controlled high-cell-density fermentations by Escherichia coli K-12
Authors: Kleman G L , Horken K M , Tabita F R , Strohl W R
Published: January 2020

A predictive and feedback glucose feed controller, previously developed for nutrient-sufficient growth of Escherichia coli to high cell densities, was used to produce large quantities of a heterologous, cyanobacterial recombinant hexadecameric (L8S8) protein, ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) in E. coli. Culture and plasmid stability conditions were optimized to yield the production of approximately 1 g of soluble, active recombinant RubisCO per liter. Recombinant RubisCO also was produced in lactose-induced high-cell-density fermentation of E. coli K-12.
A simple procedure for isolation of cloning vectors and endogenous plasmids from viridans group streptococci and Staphylococcus aureus
Authors: Vriesema A J , Zaat S A , Dankert J
Published: January 2020

Isolation of plasmid DNA from viridans group streptococci is difficult, and preparations are often heavily contaminated with chromosomal DNA. We developed a simple protocol to isolate pure plasmid DNA for use in different molecular techniques, including automated sequencing. The protocol is also applicable for plasmid isolation from Staphylococcus aureus. In addition, the protocol allows isolation of pure endogenous plasmids from streptococci and S. aureus.
DNA polymorphism in morels: complete sequences of the internal transcribed spacer of genes coding for rRNA in Morchella esculenta (yellow morel) and Morchella conica (black morel)
Authors: Wipf D , Munch J C , Botton B , Buscot F
Published: January 2020

The internal transcribed spacer (ITS) of the gene coding for rRNA was sequenced in both directions with the gene walking technique in a black morel (Morchella conica) and a yellow morel (M. esculenta) to elucidate the ITS length discrepancy between the two species groups (750-bp ITS in black morels and 1,150-bp ITS in yellow morels.
Use of sulfite and hydrogen peroxide to control bacterial contamination in ethanol fermentation
Authors: Chang I S , Kim B H , Shin P K
Published: January 2020

Lactic acid bacteria isolated from an industrial-scale ethanol fermentation process were used to evaluate sulfite as a bacterial-contamination control agent in a cell-recycled continuous ethanol fermentation process. The viabilities of bacteria were decreased by sulfite at concentrations of 100 to 400 mg liter-1, while sulfite at the same concentrations did not change the viability of the Saccharomyces cerevisiae strain used in this process. Sulfite was effective only in the presence of oxygen. Bacteria showed differences in their susceptibilities to sulfite. Facultatively heterofermentative Lactobacillus casei 4-3 was more susceptible than was obligatory heterofermentative Lactobacillus fermentum 7-1. The former showed higher enzyme activities involved in the production and consumption of hydrogen peroxide than did the latter. The viability of L. fermentum 7-1 could be selectively controlled by hydrogen peroxide at concentrations of 1 to 10 mM. Based on these findings, it is hypothesized that the sulfur trioxide radical anions formed by peroxidase in the presence of hydrogen peroxide are responsible for the control of contaminating bacteria. Sulfite did not kill the yeast strain, which has catalase to degrade hydrogen peroxide. A cell-recycled continuous ethanol fermentation process was run successfully with sulfite treatments.
Coliphage and indigenous phage in Mamala Bay, Oahu, Hawaii
Authors: Paul J H , Rose J B , Jiang S C , London P , Xhou X , Kellogg C
Published: January 2020

Public concern over the discharge of primarily treated sewage by two offshore outfalls in Mamala Bay, Oahu, prompted a multidisciplinary study to determine the impact of such activities on the water quality in the bay and at adjacent recreational beaches. As part of this study, we determined the abundance of coliphage as an indicator of fecal pollution along with total viral direct counts and phages infective for Vibrio parahaemoltyicus 16 at stations in Mamala Bay in four quarterly samplings over 13 months. Coliphage (< 1 to 1.2 x 10(3)/liter) were found during each quarterly sampling along an offshore transect to the Sand Island waste treatment facility outfall. The nonpoint coastal stations (Pearl Harbor, Ala Wai Canal, and Ke'ehi Lagoon) had high levels of coliphage during the storm event sampling in February 1994 but much lower levels or none when sampled during dry weather. Coliphage were absent at all samplings at Waikiki Beach and at the control station off Diamond Head. Viral direct counts in eutrophic coastal stations (Pearl Harbor, Ke'ehi Lagoon, Ala Moana Beach, and Ala Wai canal) averaged 10(9)/liter, while counts at offshore stations ranged from 9 x 10(7) to 1 x 10(9) viruses/liter, values similar to those for other marine environments. Vibriophage were found mainly in eutrophic coastal environments (Ala Wai Canal, Pearl Harbor, and Ke'ehi Lagoon) and at the Sand Island Transect stations D1 and D2. The greatest abundance was found during the storm event (February 1994) sampling. These results suggest that the Sand Island outfall influenced the water quality of the immediate surrounding waters but had little effect on the quality of the recreational beaches. Nonpoint discharge sources appeared to be more important in the distribution of fecal indicators in the coastal zone.
Location and Survival of Mycorrhiza Helper Pseudomonas fluorescens during Establishment of Ectomycorrhizal Symbiosis between Laccaria bicolor and Douglas Fir
Authors: Frey-Klett P , Pierrat J C , Garbaye J
Published: January 2020

The mycorrhiza helper bacterium Pseudomonas fluorescens BBc6, isolated from a Laccaria bicolor sporocarp, consistently promotes L. bicolor-Douglas fir (Pseudotsuga menziesii) ectomycorrhizal formation, even with low doses of bacterial inoculum. In order to describe this phenomenon more accurately, we have looked at the location and survival of the introduced bacterial strain in the soil and in the rhizosphere during the establishment of mycorrhizal symbiosis in glasshouse and nursery experiments. Bacterial populations were quantified with a spontaneous, stable, rifampin-resistant mutant, BBc6R8, which phenotypically conformed to the parental strain. BBc6R8 populations declined rapidly, reaching the detection limit after 19 weeks, and did not increase either when L. bicolor sporocarps were forming in autumn or when Douglas fir roots resumed growing in spring. BBc6R8 was neither an endophyte nor a rhizobacterium. Furthermore, it was not particularly associated with either mycorrhizas of Douglas fir-L. bicolor or L. bicolor sporocarps. Surprisingly, a significant mycorrhiza helper effect was observed when the inoculated BBc6R8 population had dropped as low as 30 CFU g of dry matter(sup-1) in the soil. This study raises questions concerning the bacterial concentration in the soil which is effective for promotion of mycorrhizal establishment and the timing of the bacterial effect. It allows us to develop working hypotheses, which can be tested experimentally, to identify the mechanisms of the mycorrhiza helper effect.
Metabolism of Naphthalene, 1-Naphthol, Indene, and Indole by Rhodococcus sp. Strain NCIMB 12038
Authors: Boyd C , Larkin M J , Reid K A , Sharma N D , Wilson K
Published: January 2020

The regulation of naphthalene and 1-naphthol metabolism in a Rhodococcus sp. (NCIMB 12038) has been investigated. The microorganism utilizes separate pathways for the degradation of these compounds, and they are regulated independently. Naphthalene metabolism was inducible, but not by salicylate, and 1-naphthol metabolism, although constitutive, was also repressed during growth on salicylate. The biochemistry of naphthalene degradation in this strain was otherwise identical to that found in Pseudomonas putida, with salicylate as a central metabolite and naphthalene initially being oxidized via a naphthalene dioxygenase enzyme to cis-(1R,2S)-1,2-dihydroxy-1,2-dihydronaphthalene (naphthalene cis-diol). A dioxygenase enzyme was not expressed under growth conditions which facilitate 1-naphthol degradation. However, biotransformations with indene as a substrate suggested that a monooxygenase enzyme may be involved in the degradation of this compound. Indole was transformed to indigo by both naphthalene-grown NCIMB 12038 and by cells grown in the absence of an inducer. Therefore, the presence of a naphthalene dioxygenase enzyme activity was not necessary for this reaction. Thus, the biotransformation of indole to indigo may be facilitated by another type of enzyme (possibly a monooxygenase) in this organism.