bluuebunny/crossref_metadata_2025 huggingface dataset:
DNA Probe-Mediated Detection of Resistant Bacteria from Soils Highly Polluted by Heavy Metals
Authors: Diels Ludo , Mergeay Max
Published: January 2020

Alcaligenes eutrophus
CH34 DNA fragments encoding resistance to Cd
2+
, Co
2+
, Zn
2+
(
czc
), or Hg
2+
(
merA
) were cloned and used as probes in colony hybridization procedures with bacteria isolated from polluted environments such as a zinc factory area (desertified because of the toxic effects of zinc contamination) and from sediments from factories of nonferrous metallurgy in Belgium and mine areas in Zaire. From the different soil samples, strains could be isolated and hybridized with the
czc
probe (resistance to Cd
2+
, Co
2+
, and Zn
2+
from plasmid pMOL30). Percentages of CFU isolated on nonselective plates which hybridized with the
czc
and the mercury resistance probes were, respectively, 25 and 0% for the zinc desert, 15 to 20 and 10 to 20% for the two Belgian factories, and 40 and 40% for the Likasi mine area. Most of these strains also carried two large plasmids of about the same size as those of
A. eutrophus
CH34 and shared many phenotypic traits with this strain. These findings indicated a certain correlation between the heavy-metal content in contaminated soils and the presence of heavy-metal-resistant megaplasmid-bearing
A. eutrophus
strains.
Simple, Sensitive Zymogram Technique for Detection of Xylanase Activity in Polyacrylamide Gels
Authors: Royer John C. , Nakas J. P.
Published: January 2020

A method capable of detecting as little as 0.11 U of xylanase activity in polyacrylamide gels was developed. The method entails incubation of protein gels in contact with substrate gels containing unmodified xylan, followed by immersion of substrate gels in 95% ethanol. Resulting zymograms contain transparent bands corresponding to enzymatic activity against an opaque background.
Fractionation of the β-Linked Glucans of
Bradyrhizobium japonicum
and Their Response to Osmotic Potential
Authors: Tully Raymond E. , Keister Donald L. , Gross Kenneth C.
Published: January 2020

Bradyrhizobium japonicum
USDA 110 synthesized both extracellular and periplasmic polysaccharides when grown on mannitol minimal medium. The extracellular polysaccharides were separated into a high-molecular-weight acidic capsular extracellular polysaccharide fraction (90% of total hexose) and three lower-molecular-weight glucan fractions by liquid chromatography. Periplasmic glucans, extracted from washed cells with 1% trichloroacetic acid, gave a similar pattern on liquid chromatography. Linkage analysis of the major periplasmic glucan fractions demonstrated mainly 6-linked glucose (63 to 68%), along with some 3,6- (8 to 18%), 3- (9 to 11%), and terminal (7 to 8%) linkages. The glucose residues were β-linked as shown by
1
H-nuclear magnetic resonance analysis. Glucan synthesis by
B. japonicum
cells grown on mannitol medium with 0 to 350 mM fructose as osmolyte was measured. Fructose at 150 mM or higher inhibited synthesis of periplasmic and extracellular 3- and 6-linked glucans but had no effect on the synthesis of capsular acidic extracellular polysaccharides.
Screening of aquatic samples for Vibrio cholerae serotype O1 by a dot-blot method and a latex agglutination test
Authors: Nishikawa Y , Hase A , Ishii E , Kishi T
Published: January 2020

A dot-blot, enzyme-linked immunosorbent method and a latex agglutination test were studied for their abilities to detect Vibrio cholerae serotype O1 in aquatic samples by testing artificially contaminated water as well as samples from natural potential sources. Water samples were preenriched with alkaline peptone and then enriched with Monsur peptone water. For the dot-blot test, enriched cultures of organisms in a small portion of the Monsur peptone water were transferred to a polyvinylidene difluoride membrane with a microfiltration apparatus. The enzyme-linked immunosorbent assay was performed by using biotin-labeled antibodies and avidin-biotin-peroxidase complex; brown dots developed in the wells that contained serotype O1 vibrios. Latex agglutination tests were performed by mixing 1 drop of the culture in Monsur with 1 drop of reagent coated with monoclonal antibody specific for antigen A. The sensitivities and specificities of the methods were compared with those of the colony-blot method, which identified individual colonies of V. cholerae O1 in mixed bacterial cultures on isolation media. Our results indicate that the dot-blot method is as sensitive as the colony-blot method and is useful for screening for V. cholerae serotype O1 even in specimens that are heavily contaminated with non-O1 vibrios.
Localization of pyruvate carboxylase in organic acid-producing Aspergillus strains
Authors: Bercovitz A , Peleg Y , Battat E , Rokem J S , Goldberg I
Published: January 2020

The localization of pyruvate carboxylase (cytosolic or mitochondrial) was studied in nine different Aspergillus species (14 strains). In some species (A. aculeatus, A. flavus, A. foetidus, A. nidulans, A. ochraceus, and A. sojae), the pyruvate carboxylase activity could be detected only in the cytosolic fraction of the cells. Pyruvate carboxylase has been found only in the mitochondrial fraction of two strains of Aspergillus wentii. In Aspergillus oryzae and in five strains of Aspergillus niger, pyruvate carboxylase activity was detected both in the mitochondrial fraction and in the cytosol. There was no quantitative or qualitative correlation between the activities of pyruvate carboxylase in the mitochondrial and cytosolic fractions of the cells and the ability of the various Aspergillus strains to accumulate different organic acids.
Rates of Microbial Metabolism in Deep Coastal Plain Aquifers
Authors: Chapelle Francis H. , Lovley Derek R.
Published: January 2020

Rates of microbial metabolism in deep anaerobic aquifers of the Atlantic coastal plain of South Carolina were investigated by both microbiological and geochemical techniques. Rates of [2-
14
C]acetate and [U-
14
C]glucose oxidation as well as geochemical evidence indicated that metabolic rates were faster in the sandy sediments composing the aquifers than in the clayey sediments of the confining layers. In the sandy aquifer sediments, estimates of the rates of CO
2
production (millimoles of CO
2
per liter per year) based on the oxidation of [2-
14
C] acetate were 9.4 × 10
−3
to 2.4 × 10
−1
for the Black Creek aquifer, 1.1 × 10
−2
for the Middendorf aquifer, and <7 × 10
−5
for the Cape Fear aquifer. These estimates were at least 2 orders of magnitude lower than previously published estimates that were based on the accumulation of CO
2
in laboratory incubations of similar deep subsurface sediments. In contrast, geochemical modeling of groundwater chemistry changes along aquifer flowpaths gave rate estimates that ranged from 10
−4
to 10
−6
mmol of CO
2
per liter per year. The age of these sediments (ca. 80 million years) and their organic carbon content suggest that average rates of CO
2
production could have been no more than 10
−4
mmol per liter per year. Thus, laboratory incubations may greatly overestimate the in situ rates of microbial metabolism in deep subsurface environments. This has important implications for the use of laboratory incubations in attempts to estimate biorestoration capacities of deep aquifers. The rate estimates from geochemical modeling indicate that deep aquifers are among the most oligotrophic aquatic environments in which there is ongoing microbial metabolism.
Stable carbon isotope analysis of nucleic acids to trace sources of dissolved substrates used by estuarine bacteria
Authors: Coffin R B , Velinsky D J , Devereux R , Price W A , Cifuentes L A
Published: January 2020

The natural abundance of stable carbon isotopes measured in bacterial nucleic acids extracted from estuarine bacterial concentrates was used to trace sources of organic matter for bacteria in aquatic environments. The stable carbon isotope ratios of Pseudomonas aeruginosa and nucleic acids extracted from cultures resembled those of the carbon source on which bacteria were grown. The carbon isotope discrimination between the substrate and total cell carbon from bacterial cultures averaged 2.3% +/- 0.6% (n = 13). Furthermore, the isotope discrimination between the substrate and nucleic acids extracted from bacterial cultures was 2.4% +/- 0.4% (n = 10), not significantly different from the discrimination between bacteria and the substrate. Estuarine water samples were prefiltered through 1-micron-pore-size cartridge filters. Bacterium-sized particles in the filtrates were concentrated with tangential-flow filtration and centrifugation, and nucleic acids were then extracted from these concentrates. Hybridization with 16S rRNA probes showed that approximately 90% of the nucleic acids extracted on two sample dates were of eubacterial origin. Bacteria and nucleic acids from incubation experiments using estuarine water samples enriched with dissolved organic matter from Spartina alterniflora and Cyclotella caspia had stable carbon isotope values similar to those of the substrate sources. In a survey that compared diverse estuarine environments, stable carbon isotopes of bacteria grown in incubation experiments ranged from -31.9 to -20.5%. The range in isotope values of nucleic acids extracted from indigenous bacteria from the same waters was similar, -27.9 to -20.2%. Generally, the lack of isotope discrimination between bacteria and nucleic acids that was noted in the laboratory was observed in the field.(ABSTRACT TRUNCATED AT 250 WORDS)
Analysis of clinical and food-borne isolates of Listeria monocytogenes in the United States by multilocus enzyme electrophoresis and application of the method to epidemiologic investigations
Authors: Bibb W F , Gellin B G , Weaver R , Schwartz B , Plikaytis B D , Reeves M W , Pinner R W , Broome C V
Published: January 2020

To investigate the microbiology and epidemiology of the 1,700 sporadic cases of listeriosis that occur annually in the United States, we developed a multilocus enzyme electrophoresis (MEE) typing system for Listeria monocytogenes. We studied 390 isolates by MEE. Eighty-two electrophoretic types (ETs) were defined. Two distinct clusters of ETs, ET group A (ETGA) and ET group B (ETGB), separated at a genetic distance of 0.440, were identified. Strains of ETGB were associated with perinatal listeriosis (P = 0.03). All strains of H antigen type a were in ETGA, while all strains of H antigen type b were in ETGB. Among 328 clinical isolates from cases of literiosis, 55 ETs of L. monocytogenes were defined. Thirty-four ETs were identified among 62 isolates from food products. The mean number of strains per ET (5.2) was significantly higher among clinical isolates than among food-borne isolates. Examination of isolates from outbreaks further documented the link between cases and contaminated food products. In one investigation, we found 11 different ETs, ruling out a single common source as a cause of that outbreak. By examining a large number of isolates collected over a specified time in diverse geographic locations in the United States, we have begun to establish a baseline for the study of the epidemiology of listeriosis by MEE.
The Degree of Susceptibility of Nectary-Inoculated Cotton Flowers and Bolls to Subsequent Seed Infection by
Aspergillus flavus
Is Determined at or before Anthesis
Authors: Klich M. A.
Published: January 2020

In greenhouse and field studies, cotton (
Gossypium hirsutum
L.) flowers were inoculated with
Aspergillus flavus
at the involucral nectaries. Bolls developing from early-season flowers had significantly higher percentages of
A. flavus
-infected seed than did bolls from flowers formed later in the season. Seeds from bolls inoculated 2 weeks after anthesis had the same infection levels as those from flowers inoculated at anthesis. These results indicate that early-season flowers are predisposed to
A. flavus
infection and that the degree of susceptibility at anthesis is retained through early boll development.
Common amino acid domain among endopolygalacturonases of ascomycete fungi
Authors: Keon J P , Waksman G
Published: January 2020

The endopolygalacturonase (EC 3.2.1.15) enzymes produced in vitro by three ascomycete fungi, Aspergillus niger, Sclerotinia sclerotiorum, and Colletotrichum lindemuthianum were studied by using thin-layer isoelectric focusing and activity stain overlay techniques. The polygalacturonases from A. niger and S. sclerotiorum consisted of numerous isoforms, whereas the endopolygalacturonase from C. lindemuthianum consisted of a single protein species. The most abundant endopolygalacturonase isoform produced by each of these organisms was purified and characterized. Biochemical parameters, including molecular weight, isoelectric point, kinetic parameters, temperature and pH optima, and thermal stability, were determined. Considerable differences in physical and chemical properties were demonstrated among these fungal polygalacturonases. Antibodies raised against individual proteins exhibited little cross-reaction, suggesting that these enzymes differ structurally as well as biochemically. In contrast, the analysis of the N-terminal amino acid sequences of the three proteins showed extensive homology, particularly in a region labeled domain 1 in which 84% of the amino acids were conserved.
Adhesion and motility of gliding bacteria on substrata with different surface free energies
Authors: Burchard R P , Rittschof D , Bonaventura J
Published: January 2020

The adhesion and motility of several aquatic and terrestrial gliding bacteria on slides differing in their critical surface energies have been examined. In general, adhesion was tenacious on low-critical surface energy (hydrophobic) surfaces and tenuous on hydrophilic surfaces. Gliding was inhibited on very hydrophobic substrata and skittish on very hydrophilic surfaces.
Coenzyme A-acylating aldehyde dehydrogenase from Clostridium beijerinckii NRRL B592
Authors: Yan R T , Chen J S
Published: January 2020

Acetaldehyde and butyraldehyde are substrates for alcohol dehydrogenase in the production of ethanol and 1-butanol by solvent-producing clostridia. A coenzyme A (CoA)-acylating aldehyde dehydrogenase (ALDH), which also converts acyl-CoA to aldehyde and CoA, has been purified under anaerobic conditions from Clostridium beijerinckii NRRL B592. The ALDH showed a native molecular weight (Mr) of 100,000 and a subunit Mr of 55,000, suggesting that ALDH is dimeric. Purified ALDH contained no alcohol dehydrogenase activity. Activities measured with acetaldehyde and butyraldehyde as alternative substrates were copurified, indicating that the same ALDH can catalyze the formation of both aldehydes for ethanol and butanol production. Based on the Km and Vmax values for acetyl-CoA and butyryl-CoA, ALDH was more effective for the production of butyraldehyde than for acetaldehyde. ALDH could use either NAD(H) or NADP(H) as the coenzyme, but the Km for NAD(H) was much lower than that for NADP(H). Kinetic data suggest a ping-pong mechanism for the reaction. ALDH was more stable in Tris buffer than in phosphate buffer. The apparent optimum pH was between 6.5 and 7 for the forward reaction (the physiological direction; aldehyde forming), and it was 9.5 or higher for the reverse reaction (acyl-CoA forming). The ratio of NAD(H)/NADP(H)-linked activities increased with decreasing pH. ALDH was O2 sensitive, but it could be protected against O2 inactivation by dithiothreitol. The O2-inactivated enzyme could be reactivated by incubating the enzyme with CoA in the presence or absence of dithiothreitol prior to assay.
Use of oligodeoxynucleotide signature probes for identification of physiological groups of methylotrophic bacteria
Authors: Tsien H C , Bratina B J , Tsuji K , Hanson R S
Published: January 2020

Oligodeoxynucleotide sequences that uniquely complemented 16S rRNAs of each group of methylotrophs were synthesized and used as hybridization probes for the identification of methylotrophic bacteria possessing the serine and ribulose monophosphate (RuMP) pathways for formaldehyde fixation. The specificity of the probes was determined by hybridizing radiolabeled probes with slot-blotted RNAs of methylotrophs and other eubacteria followed by autoradiography. The washing temperature was determined experimentally to be 50 and 52 degrees C for 9-alpha (serine pathway) and 10-gamma (RuMP pathway) probes, respectively. RNAs isolated from serine pathway methylotrophs bound to probe 9-alpha, and RNAs from RuMP pathway methylotrophs bound to probe 10-gamma. Nonmethylotrophic eubacterial RNAs did not bind to either probe. The probes were also labeled with fluorescent dyes. Cells fixed to microscope slides were hybridized with these probes, washed, and examined in a fluorescence microscope equipped with appropriate filter sets. Cells of methylotrophic bacteria possessing the serine or RuMP pathway specifically bind probes designed for each group. Samples with a mixture of cells of type I and II methanotrophs were detected and differentiated with single probes or mixed probes labeled with different fluorescent dyes, which enabled the detection of both types of cells in the same microscopic field.
Single-cell entrapment and microcolony development within uniform microspheres amenable to flow cytometry
Authors: Nir R , Lamed R , Gueta L , Sahar E
Published: January 2020

A method is presented for encapsulating single microbial cells in small spheres suitable for analysis and sorting by flow cytometry. The entrapped cells are able to multiply and form colonies contained within their respective microspheres. The system is based on ejecting the cells suspended in a gellable liquid through an orifice vibrating at ultrasonic frequencies, thus shearing the cell-containing jet into uniform droplets. When low-melting-temperature agarose was used, the droplets could be gelled into solid spheres during flight by appropriately directed colling air streams. This gelling was accompanied by significant dehydration, resulting in a twofold decrease in bead diameter and a corresponding increase in agarose concentration. Nevertheless, the microbeads obtained were highly uniform and had diameters which could be precisely controlled in the range of 10 to 40 microns. A variety of bacterial and yeast species were entrapped in agarose beads by using this system. In all cases the cells were able to develop into microcolonies containing as many as several hundred cells. This system enables one to apply the powerful method of flow cytometry to the analysis and sorting of whole microbial colonies. Potential applications of this technology in various areas of microbiology are considered.
Survival in seawater of Escherichia coli cells grown in marine sediments containing glycine betaine
Authors: Gauthier M J , Le Rudulier D
Published: January 2020

Considering both the protective effect of glycine betaine (GB) on enteric bacteria grown at high osmolarity and the possible presence of GB in marine sediments, we have analyzed the survival, in nutrient-free seawater, of Escherichia coli cells incubated in sediments supplemented with GB or not supplemented and measured the efficiency of GB uptake systems and the expression of proP and proU genes in both seawater and sediments. We did this by using strains harboring proP-lacZ and proU-lacZ operon or gene fusions. We found that the uptake of GB and the expression of both proP and proU were very weak in seawater. The survival ability of cells in seawater supplemented with GB was a linear function of GB concentration, although the overall protection by the osmolyte was low. In sediments, proP expression was weak and GB uptake and proU expression were variable, possibly depending on the availability of organic nutrients. In a sediment with a high total organic carbon content, GB uptake was very high and proU expression was enhanced; cells previously incubated in this sediment showed a higher resistance to decay in seawater. GB might therefore play a significant role in the long-term maintenance of enteric bacterial cells in some marine sediments.
Isolation and Structural Identification of a New Metabolite of
Fusarium equiseti
Authors: Xie Weiping , Mirocha Chester J. , Wen Yechun , Pawlosky Robert J.
Published: January 2020

A fluorescent compound was isolated and purified from rice cultures of
Fusarium equiseti
(Alaska 2-2). Mass spectrometry and nuclear magnetic resonance data indicated that its structure is 2,2-dimethyl-5-amino-6-(3′-hydroxyl-4′-methoxyl-butyryl)-4-chromone. It is an analog of the mycotoxin fusarochromanone, in which the amino group on C-3′ is replaced by a hydroxyl group and the hydroxyl group on C-4′ is replaced by a methoxyl group.
Resistance of Streptococcus bovis to acetic acid at low pH: relationship between intracellular pH and anion accumulation
Authors: Russell J B
Published: January 2020

Streptococcus bovis JB1, an acid-tolerant ruminal bacterium, was able to grow at pHs from 6.7 to 4.5, and 100 mM acetate had little effect on growth rate or proton motive force across the cell membrane. When S. bovis was grown in glucose-limited chemostats at pH 5.2, the addition of sodium acetate (as much as 100 mM) had little effect on the production of bacterial protein. At higher concentrations of sodium acetate (100 to 360 mM), production of bacterial protein declined, but this decrease could largely be explained by a shift in fermentation products (acetate, formate, and ethanol production to lactate production) and a decline in ATP production (3 ATP per glucose versus 2 ATP per glucose). YATP (grams of cells per mole of ATP) was not decreased significantly even by high concentrations of acetate. Cultures supplemented with 100 mM sodium acetate took up [14C]acetate and [14C]benzoate in accordance with the Henderson-Hasselbalch equation and gave similar estimates of intracellular pH. As the extracellular pH declined, S. bovis allowed its intracellular pH to decrease and maintained a relatively constant pH gradient across the cell membrane (0.9 unit). The decrease in intracellular pH prevented S. bovis from accumulating large amounts of acetate anion. On the basis of these results it did not appear that acetate was acting as an uncoupler. The sensitivity of other bacteria to volatile fatty acids at low pH is explained most easily by a high transmembrane pH gradient and anion accumulation.
Modification of the Bacillus sphaericus 51- and 42-kilodalton mosquitocidal proteins: effects of internal deletions, duplications, and formation of hybrid proteins
Authors: Clark M A , Baumann P
Published: January 2020

The 51- and 42-kDa proteins which constitute the binary mosquitocidal toxin of Bacillus sphaericus 2362 have a low overall sequence similarity but share several regions of near identity (L. Baumann, A. H. Broadwell, and P. Baumann, J. Bacteriol. 170:2045-2050, 1988). By using site-directed mutagenesis, deletions of 6 to 16 amino acids in three of these regions of the 51- and 42-kDa proteins were made, and the modified proteins were expressed in Bacillus subtilis. Deletions in both of these proteins resulted in a loss of toxicity for mosquito larvae. Hybrid proteins containing exchanged fragments of the 51- and 42-kDa proteins were inactive when tested in a variety of combinations, thereby indicating that potentially analogous fragments of these two proteins were not functionally equivalent. An internal duplication of 73 amino acids in the 51-kDa protein and 72 amino acids in the 42-kDa protein resulted in a major reduction in toxicity. These results indicate that the conserved regions of the 51- and 42-kDa proteins are necessary for toxicity to larvae and that the 51- and 42-kDa proteins, despite their sequence similarity, are unique, differing from each other by at least one essential attribute.
Cloning and DNA sequence analysis of an X-prolyl dipeptidyl aminopeptidase gene from Lactococcus lactis subsp. lactis NCDO 763
Authors: Nardi M , Chopin M C , Chopin A , Cals M M , Gripon J C
Published: January 2020

Lactococcus lactis subsp. lactis NCDO 763 (also designated ML3) possesses an X-prolyl dipeptidyl aminopeptidase (X-PDAP; EC 3.4.14.5). X-PDAP mutants were selected by an enzymatic plate assay on the basis of their inability to hydrolyze an L-phenylalanyl-L-proline-beta-naphthylamide substrate. A DNA bank from L. lactis subsp. lactis NCDO 763 was constructed in one of these X-PDAP mutants, and one clone in which the original X-PDAP phenotype was restored was detected by the enzymatic plate assay. The X-PDAP gene, designated pepXP, was further subcloned and sequenced. It codes for a protein containing 763 residues. Comparison of the amino-terminal sequence of the X-PDAP enzyme with the amino acid sequence deduced from the pepXP gene indicated that the enzyme is not subjected to posttranslational modification or exported via processing of a signal peptide. The pepXP gene from L. lactis subsp. lactis NCDO 763 in more than 99% homologous to the pepXP gene from L. lactis subsp. cremoris P8-2-47 described elsewhere (B. Mayo, J. Kok, K. Venema, W. Bockelmann, M. Teuber, H. Reinke, and G. Venema, Appl. Environ. Microbiol. 57:38-44, 1991) and is also conserved in other lactococcal strains.
Degradation of organochlorine compounds in spent sulfite bleach plant effluents by actinomycetes
Authors: Winter B , Fiechter A , Zimmermann W
Published: January 2020

Actinomycetes isolated from different soil samples were tested for their abilities to utilize spent sulfite bleach effluents from a paper mill. Degradation and dechlorination of the chlorinated compounds in the effluents of the first two bleaching stages, i.e., chlorination stage [(C + D)red.] and alkaline extraction stage (E1O), were monitored by determining total organic carbon (TOC) and activated-carbon-adsorbable organic-bound halogen (AOX). The isolates showed increased degradation rates after repeated incubations in the effluent-containing medium. Separation of the culture supernatants by ultrafiltration into three fractions of different molecular weights revealed substantial AOX and TOC reductions in the low-molecular-weight fraction. The AOX values of the higher-molecular-weight fractions were also reduced. Extracellular peroxidase and cell wall-bound catalase activities were produced during growth of the microorganisms on bleach effluents.
Subcloning of bph genes from Pseudomonas testosteroni B-356 in Pseudomonas putida and Escherichia coli: evidence for dehalogenation during initial attack on chlorobiphenyls
Authors: Ahmad D , Sylvestre M , Sondossi M
Published: January 2020

The bphA, -B, -C, and -D genes from Pseudomonas testosteroni B-356 were mapped to a 5.5-kb DNA fragment of cloned plasmids pDA1 and pDA2 by use of deletion and insertion mutants of these plasmids. The expression of each of these genes was evaluated in Escherichia coli and in Pseudomonas putida, and it was found that the bphC and bphD genes are well expressed in both E. coli and P. putida cells while the bphA and bphB genes are very poorly expressed in E. coli, even when placed downstream of a tac promotor. P. putida clones carrying the bphA gene were used to study the metabolites produced from 4,4'-dichlorobiphenyl, 2,2'-dichlorobiphenyl, and 2,4'-dichlorobiphenyl. It was shown that dehalogenation of 4-Cl and 2-Cl occurs in the course of the initial oxygenase attack on these molecules, which always occurs on carbons 2 and 3, independently of the positions of the chlorine atoms. Our data also suggest that in the case of polychlorobiphenyl congeners carrying chlorine atoms on both rings, it appears that, depending on the chlorine positions, dioxygenation will occur predominantly on one ring over the other. However, attack of the more resistant ring is not excluded, resulting in multiple conversion pathways.
Distribution of Legionella spp. in hydrothermal areas in continental Portugal and the island of São Miguel, Azores
Authors: Veríssimo A , Marrão G , da Silva F G , da Costa M S
Published: January 2020

Nineteen aquatic environment sites from three hydrothermal areas on continental Portugal and one area on the island of São Miguel, Azores, were examined for the recovery of Legionella spp. Physicochemical and bacteriological parameters were also determined for each site. Water temperatures varied between 22 and 67.5 degrees C, although the majority had temperatures above 40 degrees C; the pH varied between 5.5 and 9.2. The number of Legionella spp. recovered varied between 5.0 x 10(2) and 2.3 x 10(6) CFU/liter. A total of 288 isolates from 14 sites were identified by indirect immunofluorescence assay. The majority of the isolates belonged to Legionella pneumophila (74.3%), of which most belong to serogroup 1, but the relative proportion of L. pneumophila serogroups varied considerably. L. pneumophila serogroup 1 constituted 96.2% of the isolates in area 2 from central Portugal, but no isolates of this serogroup were recovered from São Miguel, where serogroup 6 strains were the predominant isolates. Ninety-six percent of the L. pneumophila serogroup 1 isolates belonged to monoclonal antibody subgroups OLDA and Bellingham. Other species identified were L. bozemanii serogroup 2, L. dumoffii, L. micdadei, L. moravica, L. oakridgensis, L. sainticrucis, and L. sainthelensi. Two undescribed species, which react by indirect immunofluorescence assay to antisera to "L. londoniensis" and "L. nautarum" and a group of isolates with strong cross-reaction to L. cincinnatiensis/L. sainticrucis/L. longbeachae by indirect immunofluorescence assay were also recovered. The latter were the only isolates recovered from area 3, in east central Portugal, over a period of 1 year.
Reverse Sample Genome Probing, a New Technique for Identification of Bacteria in Environmental Samples by DNA Hybridization, and Its Application to the Identification of Sulfate-Reducing Bacteria in Oil Field Samples
Authors: Voordouw Gerrit , Voordouw Johanna K. , Karkhoff-Schweizer Roxann R. , Fedorak Phillip M. , Westlake Donald W. S.
Published: January 2020

A novel method for the identification of bacteria in environmental samples by DNA hybridization is presented. It is based on the fact that, even within a genus, the genomes of different bacteria may have little overall sequence homology. This allows the use of the labeled genomic DNA of a given bacterium (referred to as a “standard”) to probe for its presence and that of bacteria with highly homologous genomes in total DNA obtained from an environmental sample. Alternatively, total DNA extracted from the sample can be labeled and used to probe filters on which denatured chromosomal DNA from relevant bacterial standards has been spotted. The latter technique is referred to as reverse sample genome probing, since it is the reverse of the usual practice of deriving probes from reference bacteria for analyzing a DNA sample. Reverse sample genome probing allows identification of bacteria in a sample in a single step once a master filter with suitable standards has been developed. Application of reverse sample genome probing to the identification of sulfate-reducing bacteria in 31 samples obtained primarily from oil fields in the province of Alberta has indicated that there are at least 20 genotypically different sulfate-reducing bacteria in these samples.
Increased Bacterial Uptake of Macromolecular Substrates with Fluid Shear
Authors: Confer David R. , Logan Bruce E.
Published: January 2020

To investigate the effect of fluid shear on uptake rates of low-diffusivity macromolecular substrates by suspended cultures, we measured the uptake of two compounds as models of macromolecules, a protein (bovine serum albumin [BSA]) and a polysaccharide (dextran), using pure cultures of
Zoogloea ramigera
and
Escherichia coli
, respectively. Oxygen utilization rates of stirred samples grown on BSA and dextran were 2.3 and 2.9 times higher, respectively, than those of undisturbed (still) samples. Uptake rates of
3
H-BSA and [
3
H]dextran by stirred samples were 12.6 and 6.2 times higher, respectively, than those by still samples. These experimentally obtained increases are larger than those predicted with a mass transfer model. Model results indicated that stirring would increase uptake by factors of 1.6 and 1.8 for BSA and dextran. As predicted by the model, we also found that uptake rates of low-molecular-weight substrates with high diffusivities, such as leucine and glucose, were only slightly affected by fluid shear. Since macromolecules can make up a major portion of bacterial substrate in natural, laboratory, and engineered systems, the demonstrated effect of fluid shear has wide implications for kinetic studies performed in basic metabolic research as well as in the evaluation of engineered bioreactors used for wastewater treatment.
Effects of Genetically Engineered Microorganisms on Nitrogen Transformations and Nitrogen-Transforming Microbial Populations in Soil
Authors: Jones R. A. , Broder M. W. , Stotzky G.
Published: January 2020

The principal concern about releasing genetically engineered microorganisms (GEMs) into the environment is their potential adverse effects on the environment, whether caused directly or indirectly by the GEMs. The effects of five GEMs on ammonification, nitrification, and denitrification in soil were studied. With the possible exception of a strain of
Enterobacter cloacae
carrying a plasmid, no consistent statistically or ecologically significant differences in effects on these processes or on the population dynamics of the microorganisms responsible for the processes were observed between soils inoculated with the GEMs or their homologous plasmidless hosts and those that were not inoculated. Increasing the concentration of montmorillonite in the soil enhanced the rate of nitrification, regardless of the inoculum, indicating that the perfusion technique used was sensitive enough to detect changes in nitrification rates when they occurred.
Identification of mesophilic lactic acid bacteria by using polymerase chain reaction-amplified variable regions of 16S rRNA and specific DNA probes
Authors: Klijn N , Weerkamp A H , de Vos W M
Published: January 2020

Specific DNA probes based on variable regions V1 and V3 of 16S rRNA of lactic acid bacteria were designed. These probes were used in hybridization experiments with variable regions amplified by using the polymerase chain reaction. In this way, a rapid and sensitive method was developed for the identification and classification of Lactococcus and Leuconostoc species.
Cellular fatty acid analysis as a potential tool for predicting mosquitocidal activity of Bacillus sphaericus strains
Authors: Frachon E , Hamon S , Nicolas L , de Barjac H
Published: January 2020

Gas-liquid chromatography of fatty acid methyl esters and numerical analysis were carried out with 114 Bacillus sphaericus strains. Since only two clusters harbored mosquitocidal strains, this technique could be developed in screening programs to limit bioassays on mosquito larvae. It also allows differentiation of highly homologous strains.
Complete degradation of polychlorinated hydrocarbons by a two-stage biofilm reactor
Authors: Fathepure B Z , Vogel T M
Published: January 2020

A two-stage anaerobic-aerobic biofilm reactor successfully degraded a mixture of chlorinated organic compounds to water-soluble metabolic intermediates and carbon dioxide. Reductive dechlorination of hexachlorobenzene (HCB), tetrachloroethylene (PCE), and chloroform (CF) occurred on all tested primary carbon sources such as glucose, methanol, and acetate. However, the extent of dechlorination was maximum when the anaerobic biofilm column was fed acetate as a primary carbon source. HCB, PCE, and CF were dechlorinated to the levels of tri- and dichlorinated products (99, 80, and 32%, respectively) with acetate in the feed. This is important, since these less-chlorinated compounds can be metabolized by the aerobic biofilm. The effluent from the anaerobic biofilm column was fed directly into the aerobic column. After both columns, the total amount transformed into nonvolatile intermediates and carbon dioxide was 94, 96, and 83% for [14C]HCB, [14C]trichloroethylene, and [14C]CF, respectively. This research shows the potential application of this novel two-stage bioreactor system for treating groundwaters and industrial effluents composed of highly chlorinated aliphatic and aromatic hydrocarbons.
Citrate Fermentation by
Lactococcus
and
Leuconostoc
spp
Authors: Starrenburg Marjo J. C. , Hugenholtz Jeroen
Published: January 2020

Citrate and lactose fermentation are subject to the same metabolic regulation. In both processes, pyruvate is the key intermediate.
Lactococcus lactis
subsp.
lactis
biovar diacetylactis homofermentatively converted pyruvate to lactate at high dilution (growth) rates, low pH, and high lactose concentrations. Mixed-acid fermentation with formate, ethanol, and acetate as products was observed under conditions of lactose limitation in continuous culture at pH values above 6.0. An acetoin/butanediol fermentation with α-acetolactate as an intermediate was found upon mild aeration in continuous culture and under conditions of excess pyruvate production from citrate.
Leuconostoc
spp. showed a limited metabolic flexibility. A typical heterofermentative conversion of lactose was observed under all conditions in both continuous and batch cultures. The pyruvate produced from either lactose or citrate was converted to
d
-lactate. Citrate utilization was pH dependent in both
L. lactis
and
Leuconostoc
spp., with maximum rates observed between pH 5.5 and 6.0. The maximum specific growth rate was slightly stimulated by citrate, in
L. lactis
and greatly stimulated by citrate in
Leuconostoc
spp., and the conversion of citrate resulted in increased growth yields on lactose for both
L. lactis
and
Leuconostoc
spp. This indicates that energy is conserved during the metabolism of citrate.
Improved vector for promoter screening in lactococci
Authors: Bojovic B , Djordjevic G , Topisirovic L
Published: January 2020

Fragments of Lactococcus lactis subsp. lactis NP45 chromosomal DNA provided promoter activity in Escherichia coli when cloned into the promoter probe vector pGKV210. Only 13% of these recombinant plasmids promoted detectable cat-86 activity when transferred to L. lactis, i.e., expressed chloramphenicol resistance. In these promoter-containing versions of pGKV210, the cat-86 gene specifies chloramphenicol-inducible chloramphenicol acetyltransferase expression. This could be a limiting factor for cloning of promoters with lower activity in L. lactis. Therefore, we have constructed a new promoter probe vector, pBV5030, with the mutated version of the cat-86 gene, which is constitutively expressed when transcriptionally activated by the insertion of a promoter. We found that in L. lactis IL1403 the constitutively expressed cat-86 gene (on a pBV5030 derivative) has four times higher activity than the inducible version of the same gene (on a pGKV210 derivative) when both have the same promoter inserted upstream of the cat-86 gene. These results suggest that plasmid pBV5030 could be a more efficient vector for the cloning of promoters from lactococci.
Isolation and Identification of Vesicular-Arbuscular Mycorrhiza-Stimulatory Compounds from Clover (
Trifolium repens
) Roots
Authors: Nair Muraleedharan G. , Safir Gene R. , Siqueira Jose O.
Published: January 2020

Two isoflavonoids isolated from clover roots grown under phosphate stress were characterized as formononetin (7-hydroxy,4′-methoxy isoflavone) and biochanin A (5,7-dihydroxy,4′-methoxy isoflavone). At 5 ppm, these compounds stimulated hyphal growth in vitro and root colonization of an undescribed vesicular-arbuscular mycorrhiza, a
Glomus
sp. (INVAM-112). The permethylated products of the two compounds were inactive. These findings suggest that the isoflavonoids studied may act as signal molecules in vesicular-arbuscular mycorrhiza symbiosis.
Ecophysiological and Trophic Implications of Light-Stimulated Amino Acid Utilization in Marine Picoplankton
Authors: Paerl Hans W.
Published: January 2020

By using microautoradiography, light-stimulated utilization of dissolved amino acids for natural marine phytoplankton assemblages was demonstrated. The <2-μm-size (diameter) picoplankton, known to be a dominant fraction of marine primary production, revealed a widespread capability for this process. Autofluorescent (chlorophyll
a
-containing) picoplankton and some larger phytoplankton from diverse oceanic locations, as well as isolates of the representative cyanobacterial picoplankton
Synechococcus
spp. (WH7803, WH8101), showed light-stimulated incorporation of amino acids at trace concentrations. Dark-mediated amino acid utilization was dominated by nonfluorescent bacterial populations. Among autofluorescent picoplankton, light-stimulated exceeded dark-mediated amino acid incorporation by 5 to 75%; light-stimulated amino acid incorporation was only partially blocked by the photosystem II inhibitor 3(3,4-dichloro-phenyl)-1,1-dimethy-lurea (2 × 10
-5
M), suggesting a photoheterotrophic incorporation mechanism. Parallel light versus dark incubations with glucose and mannitol indicated a lack of light-stimulated utilization of these nonnitrogenous compounds. Since marine primary production is frequently nitrogen limited, light-mediated auxotrophic utilization of amino acids and possibly other dissolved organic nitrogen (DON) constituents may represent exploitation of the relatively large DON pool in the face of dissolved inorganic nitrogen depletion. This process (i) increases the efficiency of DON retention at the base of oceanic food webs and (ii) may in part be responsible for relatively high rates of picoplankton production under conditions of chronic dissolved inorganic nitrogen limitation. Picoplanktonic recycling of organic matter via this process has important ramifications with respect to trophic transfer via the “microbial loop.”
Production and localization of beta-fructosidase in asynchronous and synchronous chemostat cultures of yeasts
Authors: Rouwenhorst R J , van der Baan A A , Scheffers W A , Van Dijken J P
Published: January 2020

In synchronized continuous cultures of Saccharomyces cerevisiae CBS 8066, the production of the extracellular invertase (EC 3.2.1.26) showed a cyclic behavior that coincided with the budding cycle. The invertase activity increased during bud development and ceased at bud maturation and cell scission. The cyclic changes in invertase production resulted in cyclic changes in amounts of invertase localized in the cell wall. However, the amount of enzyme invertase present in the culture liquid remained constant throughout the budding cycle. Also, in asynchronous continuous cultures of S. cerevisiae, the production and localization of invertase showed significant fluctuation. The overall invertase production in an asynchronous culture was two to three times higher than in synchronous cultures. This could be due to more-severe invertase-repressive conditions in a synchronous chemostat culture. Both the intracellular glucose-6-phosphate concentration and residual glucose concentration were significantly higher in synchronous chemostat cultures than in asynchronous chemostat cultures. In the asynchronous and synchronous continuous cultures of S. cerevisiae, about 40% of the invertase was released into the culture liquid; it has generally been believed that S. cerevisiae releases only about 5% of its invertase. In contrast to invertase production and localization in the chemostat cultures of S. cerevisiae, no significant changes in inulinase (EC 3.2.1.7) production and localization were observed in chemostat cultures of Kluyveromyces maxianus CBS 6556. In cultures of K. marxianus about 50% of the inulinase was present in the culture liquid.
In situ bacterial selenate reduction in the agricultural drainage systems of western Nevada
Authors: Oremland R S , Steinberg N A , Presser T S , Miller L G
Published: January 2020

Dissimilatory in situ selenate reduction to elemental selenium in sediments from irrigated agricultural drainage regions of western Nevada was measured at ambient Se oxyanion concentrations. Selenate reduction was rapid, with turnover rate constants ranging from 0.04 to 1.8 h-1 at total Se concentrations in pore water of 13 to 455 nM. Estimates of removal rates of selenium oxyanions were 14.38, and 155 mumol m-2 day-1 for South Lead Lake, Massie Slough, and Hunter Drain, respectively.
Chemotactic Behavior of
Azotobacter vinelandii
Authors: Haneline Stephen , Connelly Carla J. , Melton Thoyd
Published: January 2020

Chemotaxis was exhibited by
Azotobacter vinelandii
motile cells. Exposure of cells to sudden increases in attractant concentration suppressed the frequency of tumbling and resulted in smooth swimming. Cells responded chemotactically to a chemical gradient produced during metabolism. Motility occurred over a temperature range of 25 to 37°C with an optimum pH range of between pH 7.0 and 8.0. The average speed of motile cells was determined to be 74 μm/s or 37 body lengths per s. The speed of cells appeared to increase as a function of attractant concentration. Chemotactic systems for fructose, glucose, xylitol, and mannitol were inducible.
A. vinelandii
exhibited chemotaxis for a number of compounds, including hexoses, hexitols, pentitols, pentoses, disaccharides, and amino sugars. We conclude from these studies that
A. vinelandii
exhibits a temporal chemotactic sensing system.
Portable Device for Preparation and Delivery of Gas Mixtures
Authors: Misaghi I. J. , Stowell L. J.
Published: January 2020

A simple, portable device for the preparation and delivery of gas mixtures has been designed and constructed. The basic feature of the device is the use of gas flow controllers to maintain stable flow rates over a wide range of downstream pressures, instead of the capillary tubes and water-filled barostats commonly used in gas-mixing devices. Elimination of the barostat avoids problems such as water leakage, the loss of gases through the barostat, and changes in gas pressure due to evaporative loss of water from the barostat. The absence of a barostat also provides a closed system, allowing the use of the device for mixing and delivering of toxic gases. The prototype of the device has been used to prepare mixtures of different gases for more than 1 year and has been found to operate consistently and reproducibly. The actual concentrations of O
2
, CO
2
, and N
2
in gas mixtures (determined by gas chromatography) immediately after mixing were between 2.2 and 6.6% of the desired values in four performance tests. Fluctuations in concentration of gases in mixtures after 9 days of continuous gas delivery was less than 2% in four performance tests.
Bacterial nutrients in drinking water
Authors: LeChevallier M W , Schulz W , Lee R G
Published: January 2020

Regrowth of coliform bacteria in distribution systems has been a problem for a number of water utilities. Efforts to solve the regrowth problem have not been totally successful. The current project, which was conducted at the New Jersey American Water Co.-Swimming River Treatment Plant, showed that the occurrence of coliform bacteria in the distribution system could be associated with rainfall, water temperatures greater than 15 degrees C, total organic carbon levels greater than 2.4 mg/liter, and assimilable organic carbon levels greater than 50 micrograms of acetate carbon equivalents per liter. A multiple linear regression model based on free chlorine residuals present in dead-end sections of the distribution system and temperature predicted 83.8% of the heterotrophic plate count bacterial variation. To limit the growth of coliform bacteria in drinking water, the study concludes that assimilable organic carbon levels should be reduced to less than 50 micrograms/liter.