bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of Prechilling and Sequential Washing on Enumeration of Microorganisms from Refuse
Authors: Barlaz M. A. , Schaefer D. M. , Ham R. K.
Published: January 2020

Techniques were evaluated for formation of a liquid inoculum from shredded municipal refuse, including chilling the refuse at 4°C prior to blending and multiple washing and blending cycles. The average count of cellulolytic bacteria from six different detachment treatments was 5.1 × 10
4
cells per g (dry weight) of refuse with a range of 0.7 × 10
4
to 12.7 × 10
4
cells per g (dry weight). The liquid obtained from blending the refuse in phosphate buffer followed by hand squeezing was the selected detachment procedure. The inoculum formation procedure was validated by the addition of ruminal cellulolytic bacteria to refuse and recovery of the cellulolytic bacteria by most-probable-number enumerations. The ratio of measured to expected cell counts among tests in which different volumes of ruminal fluid were added to refuse ranged from 2.7 to 14.4. There was no evidence of anaerobic cellulolytic fungi in a refuse sample.
Field Study Comparing Growth and Viability of a Population of Phototrophic Bacteria
Authors: Folt Carol L. , Wevers Mary Jo , Yoder-Williams Michael P. , Howmiller Richard P.
Published: January 2020

The growth and viability of an anoxygenic, phototrophic bacterial community in the hypolimnion of Zaca Lake, Calif., were compared throughout the summer. The community is dominated by a single species, “
Thiopedia rosea
,” that inhabits the entire hypolimnion (6 to 8 m) for approximately 11 months. Suboptimal conditions in the hypolimnion (extremely low light intensity, high or low H
2
S levels) result in zero or extremely low growth rates (doubling times > 1 month) for most of the population, most of the time, yet cells remain viable and capable of high specific growth rates (doubling times of 1 to 10 days) when placed under favorable conditions (higher light intensities and temperatures). We first conclude that phototrophic bacterial populations in situ may frequently exist in a viable yet nongrowing state. Second, the viability of cells is likely to be reduced with depth owing to higher concentrations of potentially toxic chemicals and to changes in the physiological state associated with the prolonged periods of darkness commonly found at the bottom of bacterial plates.
Growth and Nodulation Responses of
Rhizobium meliloti
to Water Stress Induced by Permeating and Nonpermeating Solutes
Authors: Busse Matt D. , Bottomley Peter J.
Published: January 2020

Isolates of
Rhizobium meliloti
, representing antigenically distinct indigenous serogroups 31 and 17, were grown in yeast extract-mannitol broth (YEM) containing NaCl or polyethylene glycol (PEG) to provide external water potentials ranging from −0.15 to −1.5 MPa. Several differences were found between representatives of the two groups in their abilities to adapt to water stress induced by the nonpermeating solute PEG. At potentials below −0.5 MPa, strain 31 had a lower specific growth rate than strain 17 and an irregular cell morphology. In contrast, neither growth nor cell morphology of either strain was affected significantly over the same range of water potentials created by a permeating solute, NaCl. Despite the superior growth of strain 17 at the low water potentials imposed by PEG, upshock of water-stressed cells (−1.0 MPa; PEG) into normal YEM (−0.15 MPa) resulted in a faster recovery of growth by strain 31 than by strain 17. Different responses of the two strains to a water potential increase were also revealed in nodulation studies. Strain 31 required significantly fewer days to nodulate alfalfa than strain 17 did when the strains were transferred from YEM with PEG at −1.0 MPa onto the roots of alfalfa seedlings in plant growth medium (−0.1 MPa). The addition of supplemental calcium (0.1 mM) to growth medium with PEG (−1.0 MPa) reduced the differences between strains in their responses to water stress. The severe growth restriction and morphological abnormalities shown by strain 31 were corrected, and the prolonged recovery time shown by water-stressed cells (−1.0 MPa; PEG) of strain 17 upon transfer to normal YEM was shortened. The latter strain also nodulated earlier and more rapidly after growth in PEG medium at −1.0 MPa in the presence of supplemental calcium ions. These results indicate that the efficacy of osmoregulation can vary among strains of the same species and that the mechanism of osmoregulation may differ depending on the nature of the water stress.
Alternative Environmental Roles for Cellulose Produced by
Acetobacter xylinum
Authors: Williams W. Scott , Cannon Robert E.
Published: January 2020

The cellulose-producing bacterium
Acetobacter xylinum
has been considered a strict aerobe, and it has been suggested that the function of cellulose is to hold cells in an aerobic environment. In this study, we showed that
A. xylinum
is capable of growing microaerophilically. Cellulose pellicles provided significant protection to
A. xylinum
cells from the killing effects of UV light. In experiments measuring colonization by
A. xylinum
, molds, and other bacteria on pieces of apple, cellulose pellicles enhanced colonization of
A. xylinum
on the substrate and provided protection from competitors which use the same substrate as a source of nutrients. Cellulose pellicles produced by
A. xylinum
may have multiple functions in the growth and survival of the organism in nature.
Bradyrhizobium japonicum
Inoculant Mobility, Nodule Occupancy, and Acetylene Reduction in the Soybean Root System
Authors: McDermott Timothy R. , Graham Peter H.
Published: January 2020

In the American Midwest, superior inoculant rhizobia applied to soybeans usually occupy only 5 to 20% of nodules, and response to inoculation is the exception rather than the rule. Attempts to overcome this problem have met with limited success. We evaluated the ability of
Bradyrhizobium japonicum
, supplied as a seed coat inoculant, to stay abreast of the infectible region of the developing soybean root system. The rhizoplane population of the inoculant strain declined with distance from site of placement, the decrease being more pronounced on lateral than on taproots. This decline was paralleled by a decrease in inoculant-strain nodule occupancy. Inoculant bradyrhizobia contributed little to nodulation of lateral roots, which at pod-fill accounted for more than 50% of nodule number and mass, and were major contributors to acetylene reduction activity. From these data, it appears that inoculant bradyrhizobia are competitive with indigenous soil strains at the point of placement in the soil but have limited mobility and so are incapable of sustaining high populations throughout the developing root system. The result is low nodule occupancy by the inoculant strain in the tapand lateral roots. Future studies should address aspects of inoculant placement and establishment.
Evidence for a new pathway in the bacterial degradation of 4-fluorobenzoate
Authors: Oltmanns R H , Müller R , Otto M K , Lingens F
Published: January 2020

Six bacterial strains able to use 4-fluorobenzoic acid as their sole source of carbon and energy were isolated by selective enrichment from various water and soil samples from the Stuttgart area. According to their responses in biochemical and morphological tests, the organisms were assigned to the genera Alcaligenes, Pseudomonas, and Aureobacterium. To elucidate the degradation pathway of 4-fluorobenzoate, metabolic intermediates were identified. Five gram-negative isolates degraded this substrate via 4-fluorocatechol, as described in previous studies. In growth experiments, these strains excreted 50 to 90% of the fluoride from fluorobenzoate. Alcaligenes sp. strains RHO21 and RHO22 used all three isomers of monofluorobenzoate. Alcaligenes sp. strain RHO22 also grew on 4-chlorobenzoate. Aureobacterium sp. strain RHO25 transiently excreted 4-hydroxybenzoate into the culture medium during growth on 4-fluorobenzoate, and stoichiometric amounts of fluoride were released. In cell extracts from this strain, the enzymes for the conversion of 4-fluorobenzoate, 4-hydroxybenzoate, and 3,4-dihydroxybenzoate could be detected. All these enzymes were inducible by 4-fluorobenzoate. These data suggest a new pathway for the degradation of 4-fluorobenzoate by Aureobacterium sp. strain RHO25 via 4-hydroxybenzoate and 3,4-dihydroxybenzoate.
Alkaline Phosphatase and Other Hydrolyases Produced by
Cenococcum graniforme
, an Ectomycorrhizal Fungus
Authors: Bae Kwang-Sung , Barton Larry L.
Published: January 2020

Cell extracts of
Cenococcum graniforme
have been found to contain the following hydrolytic enzymes: protease, esterase, α-
d
-galactopyranosidase, β-
d
-galactopyranosidase, α-
d
-mannopyranosidase, β-
d
-xylopyranosidase, α-
d
-glucopyranosidase, β-
d
-glucopyranosidase, and alkaline phosphatase. Sulfatase, inorganic pyrophosphatase, and β-
d
-mannopyranosidase were not detected in the extracts. β-
d
-Xylopyranosidase and α-
d
-mannopyranosidase were most active in the neutral pH range, protease and phosphatase were most active in the alkaline pH range, and other enzymes were most active in the acidic pH range. These enzymes showed a high association with cell wall material, and the release of enzymes from the cells into the culture fluid appeared to occur only when the cells were undergoing autolysis. Alkaline phosphatase in
C. graniforme
is a constitutive enzyme, and examination of the alkaline phosphatase following a purification of 265-fold produced the following characteristics: pH optimum of 9.5,
M
r
of 60,000,
K

m

of 2.1 × 10
-4
M for
p
-nitrophenylphosphate, and activation energy for hydrolysis of the substrate at 9.9 kcal (1 cal = 4.184 J)/mol.
Host Plant Effects on Nodulation and Competitiveness of the
Bradyrhizobium japonicum
Serotype Strains Constituting Serocluster 123
Authors: Cregan P. B. , Keyser H. H. , Sadowsky M. J.
Published: January 2020

Strains in
Bradyrhizobium japonicum
serocluster 123 are the major indigenous competitors for nodulation in a large portion of the soybean production area of the United States. Serocluster 123 is defined by the serotype strains USDA 123, USDA 127, and USDA 129. The objective of the work reported here was to evaluate the ability of two soybean genotypes, PI 377578 and PI 417566, to restrict the nodulation and reduce the competitiveness of serotype strains USDA 123, USDA 127, and USDA 129 in favor of the highly effective strain CB1809 and to determine how these soybean genotypes alter the competitive relationships among the three serotype strains in the serocluster. The soybean genotypes PI 377578 and PI 417566 along with the commonly grown cultivar Williams were planted in soil essentially free of soybean rhizobia and inoculated with single-strain treatments of USDA 123, USDA 127, USDA 129, or CB1809 and six dual-strain competition treatments of USDA 123, USDA 127, or USDA 129 versus CB1809, USDA 123 versus USDA 127, USDA 123 versus USDA 129, and USDA 127 versus USDA 129. PI 377578 severely reduced the nodulation and competitiveness of USDA 123 and USDA 127, while PI 417566 similarly affected the nodulation and competitiveness of USDA 129. Thus, the two soybean genotypes can reduce the nodulation and competitiveness of each of the three serocluster 123 serotype strains. Our results indicate that host control of restricted nodulation and reduced competitiveness is quite specific and effectively discriminates between
B. japonicum
strains which are serologically related.
Microbial degradation of seven amides by suspended bacterial populations
Authors: Steen W C , Collette T W
Published: January 2020

Microbial transformation rate constants were determined for seven amides in natural pond water. A second-order mathematical rate expression served as the model for describing the microbial transformation. Also investigated was the relationship between the infrared spectra and the second-order rate constants for these amides. Second-order rate constants (k2) ranged from a low of 2.0 X 10(-14) to a high of 1.1 X 10(-9) liters organism-1 h-1 for niclosamide (2',5-dichloro-4'-nitrosalicylanilide) and propachlor (2-chloro-N-isopropylacetanilide), respectively. The mechanism of degradation (i.e., microbially mediated hydrolysis) of the amides was consistent with that of other organic chemicals previously studied in a variety of natural waters. Preliminary investigations indicate that temporal variations in measured second-order rate constants are small. A simple linear regression of the infrared carbonyl-stretching frequency with log K2 gave a correlation coefficient (r2) of 0.962.
Alteration of the catalytic efficiency of penicillin amidase from Escherichia coli
Authors: Forney L J , Wong D C
Published: January 2020

Ampicillin and cephalexin are beta-lactam antibiotics that are synthesized by the condensation of D-(-)-alpha-aminophenylacetic acid with 6-aminopenicillanic acid or 7-aminodeacetoxycephalosporanic acid, respectively. The rates at which the penicillin amidase of Escherichia coli catalyzes these reactions are too low to be of practical use. The objective of this study was to determine whether it is possible to alter the substrate specificity of penicillin amidase and select enzymes that efficiently hydrolyze substrates with alpha-aminophenylacetyl moieties at low pH, at which the alpha-amino group is nearly completely protonated. In this study, D-(-)-alpha-aminophenylacetyl-(L)-leucine (APAL) was used as a substrate analog of ampicillin and cephalexin. The gene for the penicillin amidase of E. coli ATCC 11105 was cloned and transferred to a leucine auxotroph of E. coli; numerous amidase mutants were selected by their ability to cleave APAL and provide leucine for growth in low-pH medium. The plasmid encoding one of the mutant amidases (pA135) was used to transform naive cells, and transformants that expressed the mutant amidase were shown to grow more rapidly in medium at pH 6.5 containing 0.1 mM APAL as the sole leucine source than did cells with the wild-type amidase. The mutant amidase was purified, and the second-order rate constant (kcat/Km) for APAL hydrolysis at pH 6.5 was found to be 10-fold greater than the rate observed with the wild-type enzyme. The difference between the rates of APAL hydrolysis by the mutant and wild-type amidases increased as the pH of the reactions decreased.(ABSTRACT TRUNCATED AT 250 WORDS)
Oxygen consumption by ruminal microorganisms: protozoal and bacterial contributions
Authors: Ellis J E , Williams A G , Lloyd D
Published: January 2020

The relative contributions to O2 consumption made by the protozoal and bacterial populations present within the rumen were determined by using an open-type oxygen electrode system. Measurements indicated that two separate microbial populations contributed approximately equally to ruminal O2 consumption over the O2 concentration range experienced in situ (0.25 to 1.0 microM). The populations were observed to consume O2 under liquid-phase O2 concentrations of up to 7 microM, above which point rapid inactivation of O2 utilization was observed. Km values for the mixed population of bacteria and protozoa were 0.36 +/- 0.17 and 3.2 +/- 0.4 microM at concentrations of less than 1.6 and greater than 1.6 microM, respectively. O2 affinity values obtained for both the protozoal and bacterial populations were similar. O2 affinities of the isolated entodiniomorphid ciliates Polyplastron multivesiculatum and Eudiplodinium maggii showed O2 inhibition thresholds of 10 and 5, respectively, and apparent half-saturation constants (Km values) of 1.7 and 5.2 microM O2, respectively. Corresponding Vmax values were 7.8 microM O2 per min per 10(5) organisms for P. multivesiculatum and 3.6 microM O2 per min per 10(5) organisms for E. maggii. Mass spectroscopic analysis detected average rates of H2 production of 12.0 and 3.7 microM H2 per min per 10(5) organisms for P. multivesiculatum and E. maggii, respectively. Trace levels of dissolved O2 (less than 0.25 microM) stimulated the H2 production rate of E. maggii eightfold but inhibited that of P. multivesiculatum by 18%.
Inactivation of gerbil-cultured Giardia lamblia cysts by free chlorine
Authors: Rubin A J , Evers D P , Eyman C M , Jarroll E L
Published: January 2020

Giardia lamblia cysts were harvested from Mongolian gerbils and exposed to free chlorine in buffered water at pH 5, 7, and 9 at 15 degrees C. The contact times required to obtain a 2-log reduction in cyst survival (i.e., a 99% kill) were interpolated from survival curves generated at fixed concentrations of chlorine in the range of 0.25 to about 16 mg/liter. Concentration-time (C.t') products for 99% inactivation ranged from about 120 to nearly 1,500 mg.min/liter. These values are higher than those reported previously for free chlorine using G. lamblia cysts from infected humans. The cysts isolated from gerbils, as with other Giardia cysts, were unusually sensitive to chlorine in alkaline solutions.
Growth and luminescence of the bacterium Xenorhabdus luminescens from a human wound
Authors: Colepicolo P , Cho K W , Poinar G O , Hastings J W
Published: January 2020

Xenorhabdus luminescens, a newly isolated luminous bacterium collected from a human wound, was characterized. The effects of ionic strength, temperature, oxygen, and iron on growth and development of the bioluminescent system were studied. The bacteria grew and emitted light best at 33 degrees C in a medium with low salt, and the medium after growth of cells to a high density was found to have antibiotic activity. The emission spectrum peaked at 482 nm in vivo and at 490 nm in vitro. Both growth and the development of luminescence in X. luminescens required oxygen and iron. The isolated luciferase itself exhibited a temperature optimum at about 40 degrees C; after purification by affinity chromatography, it showed two bands (52 and 41 kilodaltons) on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, indicative of an alpha and beta subunit structure. Reduced flavin mononucleotide (Km of 1.4 microM) and tetradecanal (Km of 2.1 microM) were the best substrates for the luciferase, and the first-order decay constant under these conditions at 37 degrees C was 0.79 s-1.
Characterization of toxigenic vibrios isolated from the freshwater environment of Hiroshima, Japan
Authors: Venkateswaran K , Kiiyukia C , Takaki M , Nakano H , Matsuda H , Kawakami H , Hashimoto H
Published: January 2020

The occurrence and characterization of toxigenic vibrios in surface water and sediment samples of the fresh water environment of the Ohta River were studied. The membrane filter, pad preenrichment technique, followed by the placement of membranes onto thiosulfate citrate-bile salt-sucrose agar, was used for the enumeration of total vibrios. Qualitative examination of pathogenic vibrios was also attempted. In addition, a survey was conducted to determine the incidence of Clostridium botulinum in sediment samples of the Ohta River and the Hiroshima coast. In the identification of 361 strains, 12 species of Vibrio and two species of Listonella were observed. Non-01 Vibrio cholerae was prevalent among the members of the genus Vibrio. Vibrio parahaemolyticus (serotype 04:K34), isolated in fresh water, is significant and suggests that some still unknown conditions promote the survival of these organisms in fresh water. An estimated 132 strains were hemolytic by a simple agar method, and further characterization revealed that 82% of the hemolytic vibrios (107 strains) produced various toxins. About 71% (93 strains) elaborated cytotoxin, 55% (72 strains) produced hemolysin, and 44% (58 strains) responded for both cytotoxin and hemolysin in the crude toxin extracts. All the non-01 V. cholerae showed cytotoxic activity, and the virulent strains of Vibrio fluvialis and Vibrio spp. showed cytotonic responses in RK-13 cells. Of 36 sediment samples tested, 10 harbored C. botulinum spores (28%) and were isolated invariably in all the regions of the Hiroshima coast and in the Ohta River, except the upper region of the Ohta River.
Improved medium for recovery and enumeration of the farmer's lung organism, Saccharomonospora viridis
Authors: Amner W , Edwards C , McCarthy A J
Published: January 2020

A new medium, which we propose to call R8, was developed for the isolation and enumeration of the thermophilic actinomycete, Saccharomonospora viridis. This organism has been implicated in a range of hypersensitivity pneumonitides, including farmer's lung, but is generally isolated in small numbers from contaminated environments. Recovery of S. viridis from moldy hay and mushroom compost on R8 medium was compared with recovery on conventional media. S. viridis was isolated from both substrates but in highest numbers and most consistently on the R8 medium. The selectivity of this medium was best observed when the sedimentation chamber method was used for hay samples. Here S. viridis accounted for up to 80% of the total number of actinomycetes recovered on R8 and could not be recovered on rifampin selective medium under the same conditions. R8 was also found to be an efficient recovery medium for a range of thermophilic actinomycetes from mushroom compost and for another allergenic species, Faenia rectivirgula, from moldy hay. Contamination of isolation plates by thermophilic bacilli was reduced on R8 compared with the activity on half-strength tryptone soy agar, supplemented with 0.2% casein hydrolysate, and this, together with specific improvements in S. viridis growth, accounts for the selective effect. It is possible that the occurrence of S. viridis and its role as a causative agent of hypersensitivity pnuemonitis have been underestimated by the use of suboptimal recovery protocols. It is hoped that use of R8 in conjunction with dilution plate techniques will generate information on the ecology of S. viridis and contribute to health risk assessment studies.
Simple Method for Determination of Biodegradable Dissolved Organic Carbon in Water
Authors: Servais Pierre , Anzil Adriana , Ventresque Claire
Published: January 2020

Determination of biodegradable dissolved organic carbon in waters is of particular importance for the water treatment industry. A simple method for determining biodegradable dissolved organic carbon which is applicable to surface and drinking water is proposed. It consists of sterilizing the water sample, inoculating it with autochthonous bacteria, and measuring the decrease in dissolved organic carbon concentration due to the carbon oxidization by bacteria. The detailed experimental procedure is discussed, and validation of the method is presented. The method has been used for studying river waters and for drinking water treatment plant design.
Production of fusarin C on cereal and soybean by Fusarium moniliforme
Authors: Bacon C W , Marijanovic D R , Norred W P , Hinton D M
Published: January 2020

Two isolates of Fusarium moniliforme were compared with respect to production of a mutagenic compound, fusarin C, on seven corn varieties as well as on soybean, wheat, rye, barley, and a liquid culture medium. The isolates were originally obtained from corn and barley. Both isolates produced fusarin C on seed of all five crops within a 21-day period, and one isolate produced the largest amount on oats. Soybean was the poorest substrate for both isolates. Although the quantity of fusarin C produced on grain was isolate dependent, specific substrate requirements for each strain were suggested. The isolates differed in their ability to grow and produce fusarin C on corn with different moisture contents (16, 20, 24, and 28%). One isolate was more xerotolerant and grew at 16% moisture but did not produce the mutagen.
DNA hybridization assay for detection of gypsy moth nuclear polyhedrosis virus in infected gypsy moth (Lymantria dispar L.) larvae
Authors: Keating S T , Burand J P , Elkinton J S
Published: January 2020

Radiolabeled Lymantria dispar nuclear polyhedrosis virus DNA probes were used in a DNA hybridization assay to detect the presence of viral DNA in extracts from infected larvae. Total DNA was extracted from larvae, bound to nitrocellulose filters, and assayed for the presence of viral DNA by two methods: slot-blot vacuum filtration and whole-larval squashes. To test the assays, neonate larvae were fed droplets containing a known concentration of L. dispar nuclear polyhedrosis virus and observed for up to 10 days to determine the percentage of infected larvae. The average percent mortalities were 88.0, 60.7, 26.0, and 5.3% for larvae fed droplets containing 4.0 x 10(4), 1.0 x 10(4), 2.5 x 10(3), and 6.25 x 10(2) polyhedral inclusion bodies (PIBs) per ml, respectively. Other larvae treated with the same virus concentrations were frozen at 2, 4, and 6 days postinoculation and examined by the hybridization techniques. The average percentage of slot blots containing viral DNA equaled 81.0, 58.0, 18.0, and 6.0% for larvae blotted 4 days after treatment with 4.0 x 10(4), 1.0 x 10(4), 2.5 x 10(3), and 6.25 x 10(2) PIBs per ml, respectively, and 89.9, 52.1, 26.6, and 6.0%, respectively at 6 days postinoculation. Thus, the hybridization results were closely correlated with mortality observed in reared larvae. Hybridization of squashes of larvae frozen 4 days after receiving the above virus treatments also produced accurate measures of the incidence of virus infection.
Quantitative comparison of the laboratory and field competitiveness of Rhizobium leguminosarum biovar phaseoli
Authors: Beattie G A , Clayton M K , Handelsman J
Published: January 2020

Rhizobium leguminosarum bv. phaseoli KIM5s outcompeted strain CE3 in bean (Phaseolus vulgaris L.) root nodulation when plants were grown at any of three field sites, each with a different soil type and indigenous population, or in the laboratory in a sterilized sand, a sterilized peat-vermiculite mixture, or a nonsterile field soil. A mathematical model describing nodulation competitiveness was empirically derived to evaluate the relative competitiveness of the two strains under these conditions. This model relates the proportional representation of the two strains in the inoculum to the proportional representation of nodules occupied by each strain or both strains and provides a measure of competitiveness, which is referred to as the competitiveness index. Statistical comparisons of competitiveness indices showed that the relative competitiveness of KIM5s and CE3 remained constant when the two strains were applied in a constant ratio over a range of inoculum concentrations, from 10(3) to 10(7) cells per seed, and when they were applied in various ratios to six P. vulgaris cultivars. Furthermore, the relative competitiveness of KIM5s and CE3 in the laboratory did not differ significantly from their relative competitiveness at the three field sites studied. Thus, a study of the basis for nodulation competitiveness of KIM5s and CE3 in the laboratory has the potential to provide an understanding of competitiveness both in the laboratory and in the field.
Evaluation of automated threshold selection methods for accurately sizing microscopic fluorescent cells by image analysis
Authors: Sieracki M E , Reichenbach S E , Webb K L
Published: January 2020

The accurate measurement of bacterial and protistan cell biomass is necessary for understanding their population and trophic dynamics in nature. Direct measurement of fluorescently stained cells is often the method of choice. The tedium of making such measurements visually on the large numbers of cells required has prompted the use of automatic image analysis for this purpose. Accurate measurements by image analysis require an accurate, reliable method of segmenting the image, that is, distinguishing the brightly fluorescing cells from a dark background. This is commonly done by visually choosing a threshold intensity value which most closely coincides with the outline of the cells as perceived by the operator. Ideally, an automated method based on the cell image characteristics should be used. Since the optical nature of edges in images of light-emitting, microscopic fluorescent objects is different from that of images generated by transmitted or reflected light, it seemed that automatic segmentation of such images may require special considerations. We tested nine automated threshold selection methods using standard fluorescent microspheres ranging in size and fluorescence intensity and fluorochrome-stained samples of cells from cultures of cyanobacteria, flagellates, and ciliates. The methods included several variations based on the maximum intensity gradient of the sphere profile (first derivative), the minimum in the second derivative of the sphere profile, the minimum of the image histogram, and the midpoint intensity. Our results indicated that thresholds determined visually and by first-derivative methods tended to overestimate the threshold, causing an underestimation of microsphere size. The method based on the minimum of the second derivative of the profile yielded the most accurate area estimates for spheres of different sizes and brightnesses and for four of the five cell types tested. A simple model of the optical properties of fluorescing objects and the video acquisition system is described which explains how the second derivative best approximates the position of the edge.
Rhizobacteria of Cotton and Their Repression of Seedling Disease Pathogens
Authors: Hagedorn C. , Gould W. D. , Bardinelli T. R.
Published: January 2020

During the 1983 field season, the rhizobacteria (including organisms from rhizosphere soil and the root rhizoplane) of cotton plants at one location in Mississippi were inventoried at different plant growth stages. Isolates (1,000) were identified to the genus level and characterized for repression of
Pythium ultimum
and
Rhizoctonia solani.
Cotton seedlings were initially colonized by bacteria of many different genera, and populations quickly reached 10
8
CFU/g of root tissue. As the season progressed, the bacterial populations declined as root mass increased and the roots became more woodlike in consistency. Fluorescent pseudomonads were the most numerous gram-negative rhizobacterial isolates of those that were randomly collected and identified, and they provided the largest number of isolates with fungal repressive activity. Several other gram-negative bacterial genera were recovered throughout the growing season, and some gram-positive bacteria were also isolated routinely, but at lower numbers. There was no correlation between the proportion of rhizobacterial isolates that possessed fungal repressive activity and the plant growth stage from which the isolates were obtained. Approximately twice as many bacterial isolates demonstrated fungal repression in the agar assay compared with the inplanta assay, and isolates were found more frequently with fungal repressive activity against
P. ultimum
than against
R. solai.
Response of microbial adhesives and biofilm matrix polymers to chemical treatments as determined by interference reflection microscopy and light section microscopy
Authors: Marshall P A , Loeb G I , Cowan M M , Fletcher M
Published: January 2020

The polymers involved in the adhesion of Pseudomonas fluorescens H2S to solid surfaces were investigated to determine whether differences between cell surface adhesives and biofilm matrix polymers could be detected. Two optical techniques, i.e., interference reflection microscopy (IRM) and light section microscopy (LSM), were used to compare the responses of the two types of polymer to treatment with electrolytes, dimethyl sulfoxide (DMSO), and Tween 20. To evaluate initial adhesive polymers, P. fluorescens H2S cells were allowed to attach to glass cover slip surfaces and were immediately examined with IRM, and their response to chemical solutions was tested. With IRM, changes in cell-substratum separation distance between 0 and ca. 100 nm are detectable as changes in relative light intensity of the image; a contraction of the polymer would be detected as a darkening of the image, whereas expansion would appear as image brightening. To evaluate the intercellular polymer matrix in biofilms, 3-day-old biofilms were exposed to similar solutions, and the resultant change in biofilm thickness was measured with LSM, which measures film thicknesses between 10 and 1,000 microns. The initial adhesive and biofilm polymers were similar in that both appeared to contract when treated with electrolytes and to expand when treated with Tween 20. However, with DMSO treatment, the initial adhesive polymer appeared to contract, whereas there was no change in thickness of the biofilm polymer. These results indicate that both polymers bear acidic groups and thus act electrostatically with cations and are able to enter into hydrophobic interactions.(ABSTRACT TRUNCATED AT 250 WORDS)
Metabolism of Styrene Oxide and 2-Phenylethanol in the Styrene-Degrading
Xanthobacter
Strain 124X
Authors: Hartmans S. , Smits J. P. , van der Werf M. J. , Volkering F. , de Bont J. A. M.
Published: January 2020

Styrene oxide and 2-phenylethanol metabolism in the styrene-degrading
Xanthobacter
sp. strain 124X was shown to proceed via phenylacetaldehyde and phenylacetic acid. In cell extracts 2-phenylethanol was oxidized by a phenazine methosulfate-dependent enzyme, probably a pyrroloquinoline quinone enzyme.
Xanthobacter
sp. strain 124X also contains a novel enzymatic activity designated as styrene oxide isomerase. Styrene oxide isomerase catalyzes the isomerization of styrene oxide to phenylacetaldehyde. The enzyme was partially purified and shown to have a very high substrate specificity. Of the epoxides tested, styrene oxide was the only substrate transformed. The initial step in styrene metabolism in
Xanthobacter
sp. strain 124X is oxygen dependent and probably involves oxidation of the aromatic nucleus.
Nitrate Reduction in a Groundwater Microcosm Determined by
15
N Gas Chromatography-Mass Spectrometry
Authors: Bengtsson Göran , Annadotter Heléne
Published: January 2020

Aerobic and anaerobic groundwater continuous-flow microcosms were designed to study nitrate reduction by the indigenous bacteria in intact saturated soil cores from a sandy aquifer with a concentration of 3.8 mg of NO
3
−
-N liter
−1
. Traces of
15
NO
3
−
were added to filter-sterilized groundwater by using a Darcy flux of 4 cm day
−1
. Both assimilatory and dissimilatory reduction rates were estimated from analyses of
15
N
2
,
15
N
2
O,
15
NH
4
+
, and
15
N-labeled protein amino acids by capillary gas chromatography-mass spectrometry. N
2
and N
2
O were separated on a megabore fused-silica column and quantified by electron impact-selected ion monitoring. NO
3
−
and NH
4
+
were analyzed as pentafluorobenzoyl amides by multiple-ion monitoring and protein amino acids as their
N
-heptafluorobutyryl isobutyl ester derivatives by negative ion-chemical ionization. The numbers of bacteria and their [
methyl
-
3
H]thymidine incorporation rates were simultaneously measured. Nitrate was completely reduced in the microcosms at a rate of about 250 ng g
−1
day
−1
. Of this nitrate, 80 to 90% was converted by aerobic denitrification to N
2
, whereas only 35% was denitrified in the anaerobic microcosm, where more than 50% of NO
3
−
was reduced to NH
4
+
. Assimilatory reduction was recorded only in the aerobic microcosm, where N appeared in alanine in the cells. The nitrate reduction rates estimated for the aquifer material were low in comparison with rates in eutrophic lakes and coastal sediments but sufficiently high to remove nitrate from an uncontaminated aquifer of the kind examined in less than 1 month.
Influence of pH, nutrient availability, and growth rate on amine production by Bacteroides fragilis and Clostridium perfringens
Authors: Allison C , Macfarlane G T
Published: January 2020

Dimethylamine, methylamine, propylamine, and pyrrolidine were the major amines formed by Bacteroides fragilis NCDO 2217 during the active phase of growth in batch culture. Production of these metabolites was strongly pH dependent and was optimal under acidic conditions (pH 6.0). Low pH also favored the formation of pyrrolidine, cadaverine, and dimethylamine by Clostridium perfringens C523, but the reverse was the case with putrescine, butylamine, and propylamine, where production was maximal at neutral pH. B. fragilis was grown in continuous culture under either starch or casein limitation. Amine formation was influenced by carbohydrate availability and was greatest when the bacteria were grown at high growth rates (dilution rate, 0.20/h) under starch limitation, where they constituted about 18% of the total fermentation products measured. Amine production was optimal and increased concomitantly with growth rate when C. perfringens was grown in glucose-limited continuous culture. Under conditions of high growth rate and glucose limitation, amines accounted for approximately 27% of the fermentation products measured. When glucose in the feed medium was increased from 5 to 15 g/liter, amine production was repressed, and under these nutritional conditions the growth rate had little effect on the process.
Bacterial Metabolism of 2,6-Xylenol
Authors: Ewers Jens , Rubio Miguel Angel , Knackmuss Hans-Joachim , Freier-Schröder Doris
Published: January 2020

Strain DM1, a
Mycobacterium
sp. that utilizes 2,6-xylenol, 2,3,6-trimethylphenol, and
o
-cresol as sources of carbon and energy, was isolated. Intact cells of
Mycobacterium
strain DM1 grown with 2,6-xylenol cooxidized 2,4,6-trimethylphenol to 2,4,6-trimethylresorcinol. 4-Chloro-3,5-dimethylphenol prevents 2,6-xylenol from being totally degraded; it was quantitatively converted to 2,6-dimethylhydroquinone by resting cells. 2,6-Dimethylhydroquinone, citraconate, and an unidentified metabolite were detected as products of 2,6-xylenol oxidation in cells that were partially inactivated by EDTA. Under oxygen limitation, 2,6-dimethylhy-droquinone, citraconate, and an unidentified metabolite were released during 2,6-xylenol turnover by resting cells. Cell extracts of 2,6-xylenol-grown cells contained a 2,6-dimethylhydroquinone-converting enzyme. When supplemented with NADH, cell extracts catalyzed the reduction of 2,6-dimethyl-3-hydroxyquinone to 2,6-dimethyl-3-hydroxyhydroquinone. Since a citraconase was also demonstrated in cell extracts, a new metabolic pathway with 2,6-dimethyl-3-hydroxyhydroquinone as the ring fission substrate is proposed.
Standardized method for evaluation of hand disinfection by surgical scrub formulations
Authors: Crémieux A , Reverdy M E , Pons J L , Savage C , Chevalier J , Fleurette J , Mossé M
Published: January 2020

A standardized protocol for the evaluation of hand disinfection by surgical scrub formulations was applied to volunteers in a multicenter trial. Povidone iodine (PVI), chlorhexidine (CHX), and a nonmedicated soap (NMS) were tested. The scrubbing procedure involved three daily hand washings for five consecutive days; surviving bacteria were counted daily after being collected in a suitable neutralizing solution. Immediate efficacy (IE), cumulative efficacy (CE), and remanent effect (RE) were calculated by reference to the control hand. Statistical analyses of IE, CE, and RE showed significant differences among the three scrub formulations. IEs of PVI and CHX were equivalent and different from IE of NMS; CE and RE of CHX were higher than those of PVI and NMS. On the basis of the statistical analysis, the population size required for further studies aimed at detecting significant differences between surgical scrub formulations could be estimated.
Survey of microbial oxygenases: trichloroethylene degradation by propane-oxidizing bacteria
Authors: Wackett L P , Brusseau G A , Householder S R , Hanson R S
Published: January 2020

Microorganisms that biosynthesize broad-specificity oxygenases to initiate metabolism of linear and branched-chain alkanes, nitroalkanes, cyclic ketones, alkenoic acids, and chromenes were surveyed for the ability to biodegrade trichloroethylene (TCE). The results indicated that TCE oxidation is not a common property of broad-specificity microbial oxygenases. Bacteria that contained nitropropane dioxygenase, cyclohexanone monooxygenase, cytochrome P-450 monooxygenases, 4-methoxybenzoate monooxygenase, and hexane monooxygenase did not degrade TCE. However, one new unique class of microorganisms removed TCE from incubation mixtures. Five Mycobacterium strains that were grown on propane as the sole source of carbon and energy degraded TCE. Mycobacterium vaccae JOB5 degraded TCE more rapidly and to a greater extent than the four other propane-oxidizing bacteria. At a starting concentration of 20 microM, it removed up to 99% of the TCE in 24 h. M. vaccae JOB5 also biodegraded 1,1-dichloroethylene, trans-1,2-dichloroethylene, cis-1,2-dichloroethylene, and vinyl chloride.
Relationship between the Unicellular Red Alga
Porphyridium
sp. and Its Predator, the Dinoflagellate
Gymnodinium
sp
Authors: Ucko Michal , Cohen Ephraim , Gordin Hillel , Arad Shoshana (Malis)
Published: January 2020

Contamination of algae cultivated outdoors by various microorganisms, such as bacteria, fungi, algae, and protozoa, can affect growth and product quality, sometimes causing fast collapse of the cultures. The main contaminant of
Porphyridium
cultures grown outdoors in Israel is a
Gymnodinium
sp., a dinoflagellate that feeds on the alga. Comparison of the effects of various environmental conditions, i.e., pH, salinity, and temperature, on
Gymnodinium
and
Porphyridium
species revealed that the
Gymnodinium
sp. has sharp optimum curves, whereas the
Porphyridium
sp. has a wider range of optimum conditions and is also more resistant to extreme environmental variables. The mode of preying on the alga was observed, and the specificity of the
Gymnodinium
sp. for the
Porphyridium
sp. was shown. In addition,
Gymnodinium
extract was shown to contain enzymatic degrading activity specific to the
Porphyridium
sp. cell wall polysaccharide.
Degradation of barley straw, ryegrass, and alfalfa cell walls by Clostridium longisporum and Ruminococcus albus
Authors: Varel V H , Richardson A J , Stewart C S
Published: January 2020

The recently isolated ruminal sporeforming cellulolytic anaerobe Clostridium longisporum B6405 was examined for its ability to degrade barley straw, nonlignified cell walls (mesophyll and epidermis) and lignified cell walls (fiber) of ryegrass, and alfalfa cell walls in comparison with strains of Ruminococcus albus. R. albus strains degraded 20 to 28% of the dry matter in barley straw in 10 days, while the clostridium degraded less than 2%. A combined inoculum of R. albus SY3 and strain B6405 was no more efficient than SY3 alone, and the presence of Methanobacterium smithii PS did not increase the amount of dry matter degraded. In contrast, with alfalfa cell walls as the substrate, the clostridium was twice as active (28% weight loss) as R. albus SY3 (15%). The percentages of dry matter degraded from ryegrass cell walls of mesophyll, epidermis, and fiber for the clostridium were 50, 47, and 32%, respectively, and for R. albus SY3 they were 77, 73, and 63%, respectively. Analyses of the predominant neutral sugars (arabinose, xylose, and glucose) in the plant residues after bacterial attack were consistent with the values for dry matter weight loss. Measurements of the amount of carbon appearing in the fermentation products indicated that R. albus SY3 degraded ryegrass mesophyll cell walls most rapidly, with epidermis and fiber cell walls being degraded at similar rates. Strain B6405 attacked alfalfa cell walls at a rate greater than that of any of the ryegrass substrates. These results indicate an unexpected degree of substrate specificity in the ability of C. longisporum to degrade plant cell wall material.
Fermentation of Cellulosic Substrates in Batch and Continuous Culture by
Clostridium thermocellum
Authors: Lynd Lee Rybeck , Grethlein Hans E. , Wolkin Richard H.
Published: January 2020

Fermentation of dilute-acid-pretreated mixed hardwood and Avicel by
Clostridium thermocellum
was compared in batch and continuous cultures. Maximum specific growth rates per hour obtained on cellulosic substrates were 0.1 in batch culture and >0.13 in continuous culture. Cell yields (grams of cells per gram of substrate) in batch culture were 0.17 for pretreated wood and 0.15 for Avicel. Ethanol and acetate were the main products observed under all conditions. Ethanol:acetate ratios (in grams) were approximately 1.8:1 in batch culture and generally slightly less than 1:1 in continuous culture. Utilization of cellulosic substrates was essentially complete in batch culture. A prolonged lag phase was initially observed in batch culture on pretreated wood; the length of the lag phase could be shortened by addition of cell-free spent medium. In continuous culture with ∼5 g of glucose equivalent per liter in the feed, substrate conversion relative to theoretical ranged from 0.86 at a dilution rate (
D
) of 0.05/h to 0.48 at a
D
of 0.167/h for Avicel and from 0.75 at a
D
of 0.05/h to 0.43 at a
D
of 0.11/h for pretreated wood. At feed concentrations of <4.5 g of glucose equivalent per liter, conversion of pretreated wood was 80 to 90% at
D
= 0.083/h. Lower conversion was obtained at higher feed substrate concentrations, consistent with a limiting factor other than cellulose. Free Avicelase activities of 12 to 84 mU/ml were observed, with activity increasing in this order: batch cellobiose, batch pretreated wood < batch Avicel, continuous pretreated wood < continuous Avicel. Free cellulase activity was higher at increasing extents of substrate utilization for both pretreated wood and Avicel under all conditions tested. The results indicate that fermentation parameters, with the exception of free cellulase activity, are essentially the same for pretreated mixed hardwood and Avicel under a variety of conditions. Hydrolysis yields obtained with
C. thermocellum
cellulase acting either in vitro or in vivo were comparable to those previously reported for
Trichoderma reesei
on the same substrates.
Adsorption-Desorption Characteristics of Polychlorinated Biphenyls on Various Polymers Commonly Found in Laboratories
Authors: Cseh Tibor , Sanschagrin Sylvie , Hawari Jalal , Samson Réjean
Published: January 2020

Adsorption and desorption of Aroclor 1254 (a mixture of polychlorinated biphenyls [PCBs]) from 0.25% (wt/vol) aqueous solutions of Triton X-100 on polymeric materials common in laboratories (red vacuum rubber, latex, Tygon, Norprene, manosil, polyethylene, polypropylene, phenoxy resin, nylon, and Teflon) is described. Teflon, nylon, and phenoxy resins showed the lowest adsorptions, with efficiencies of 3.4, 22.9, and 33.0%, respectively. The other polymers adsorbed more than 90% of Aroclor 1254 under similar conditions. Adsorption of PCBs was found to increase with the lipophilicity of the polymer and to be irreversible on soft polymers. The variation in the adsorption efficiencies of these polymers toward PCBs is apparently related to variations in the chemical composition, electronic properties, and degree of softness of these polymers. The present study showed that Teflon, with less than 4% adsorption and more than 99% desorption of Aroclor 1254, is the best candidate for use in biological laboratory work.
Natural occurrence of the mycotoxin fusarochromanone, a metabolite of Fusarium equiseti, in cereal feed associated with tibial dyschondroplasia
Authors: Krogh P , Christensen D H , Hald B , Harlou B , Larsen C , Pedersen E J , Thrane U
Published: January 2020

The mycotoxin fusarochromanone, a metabolite of Fusarium fungi, is able to induce tibial dyschondroplasia (TD) in chickens under experimental conditions. On the basis of health surveillance data on TD, two broiler farms with TD prevalence rates of up to 56% were identified. In the corresponding pelleted feed samples, fusarochromanone was detected in all 12 samples analyzed by column purification and TLC, with concentrations 4 to 59 micrograms/kg. No Fusarium fungi were available from the feed because of the pelleting process, but seven Fusarium equiseti strains previously isolated from Danish cereals were checked for fusarochromanone production, and all produced fusarochromanone at 57 to 1,435 mg/kg. Thus, the potential for fusarochromanone production by F. equiseti is considerable. The identification of fusarochromanone from feed and F. equiseti was confirmed by mass, infrared, and nuclear magnetic resonance spectral analyses. This is the first report of fusarochromanone as a naturally occurring contaminant.