bluuebunny/crossref_metadata_2025_split huggingface dataset:
1,4-Benzoquinone Reductase from
Phanerochaete chrysosporium
: cDNA Cloning and Regulation of Expression
Authors: Akileswaran Lakshmi , Brock Barry J. , Cereghino Joan Lin , Gold Michael H. 
Published: December 2019

ABSTRACT

A cDNA clone encoding a quinone reductase (QR) from the white rot basidiomycete
Phanerochaete chrysosporium
was isolated and sequenced. The cDNA consisted of 1,007 nucleotides and a poly(A) tail and encoded a deduced protein containing 271 amino acids. The experimentally determined eight-amino-acid N-terminal sequence of the purified QR protein from
P. chrysosporium
matched amino acids 72 to 79 of the predicted translation product of the cDNA. The
M
r
of the predicted translation product, beginning with Pro-72, was essentially identical to the experimentally determined
M
r
of one monomer of the QR dimer, and this finding suggested that QR is synthesized as a proenzyme. The results of in vitro transcription-translation experiments suggested that QR is synthesized as a proenzyme with a 71-amino-acid leader sequence. This leader sequence contains two potential KEX2 cleavage sites and numerous potential cleavage sites for dipeptidyl aminopeptidase. The QR activity in cultures of
P. chrysosporium
increased following the addition of 2-dimethoxybenzoquinone, vanillic acid, or several other aromatic compounds. An immunoblot analysis indicated that induction resulted in an increase in the amount of QR protein, and a Northern blot analysis indicated that this regulation occurs at the level of the
qr
mRNA.
Genotype Analysis of
Escherichia coli
Strains Isolated from Children and Chickens Living in Close Contact
Authors: Kariuki S. , Gilks C. , Kimari J. , Obanda A. , Muyodi J. , Waiyaki P. , Hart C. A. 
Published: December 2019

ABSTRACT

Escherichia coli
isolates from rectal swabs from 62 chickens and stools from 42 children living in close contact with chickens on the same farms in Kiambu district, Kenya, were compared for their genetic relatedness. Antibiotic susceptibility profiles broadly categorized isolates from the children and from the chickens into two separate clusters: the majority (144; 85.5%) of the
E. coli
isolates from children were multidrug resistant, while the majority (216; 87.1%) of the
E. coli
isolates from chickens were either fully susceptible or resistant only to tetracycline. Sixty- and 100- to 110-MDA plasmids were found to encode the transferable resistance to co-trimoxazole and tetracycline.
Hin
dIII restriction endonuclease digestion of the 60- and 100- to 110-MDA plasmids produced four distinct patterns for isolates from children and three distinct patterns for isolates from chickens.
Xba
I digestion of genomic DNA followed by pulsed-field gel electrophoresis (PFGE) analysis produced 14 distinct clusters. There were six distinct PFGE clusters among the isolates from children, while among the isolates from chickens there were seven distinct clusters. Only one PFGE cluster contained isolates from both children and chickens, with the isolates displaying an approximately 60% coefficient of similarity. This study showed that although several different genotypes of
E. coli
were isolated from children and chickens from the same farms, the
E. coli
strains from these two sources were distinct.
Ethanol Synthesis by Genetic Engineering in Cyanobacteria
Authors: Deng Ming-De , Coleman John R. 
Published: December 2019

ABSTRACT

Cyanobacteria are autotrophic prokaryotes which carry out oxygenic photosynthesis and accumulate glycogen as the major form of stored carbon. In this research, we introduced new genes into a cyanobacterium in order to create a novel pathway for fixed carbon utilization which results in the synthesis of ethanol. The coding sequences of pyruvate decarboxylase (
pdc
) and alcohol dehydrogenase II (
adh
) from the bacterium
Zymomonas mobilis
were cloned into the shuttle vector pCB4 and then used to transform the cyanobacterium
Synechococcus
sp. strain PCC 7942. Under control of the promoter from the
rbcLS
operon encoding the cyanobacterial ribulose-1,5-bisphosphate carboxylase/oxygenase, the
pdc
and
adh
genes were expressed at high levels, as demonstrated by Western blotting and enzyme activity analyses. The transformed cyanobacterium synthesized ethanol, which diffused from the cells into the culture medium. As cyanobacteria have simple growth requirements and use light, CO
2
, and inorganic elements efficiently, production of ethanol by cyanobacteria is a potential system for bioconversion of solar energy and CO
2
into a valuable resource.
Genetic Evidence That High Noninduced Maltase and Maltose Permease Activities, Governed by
MALx3
-Encoded Transcriptional Regulators, Determine Efficiency of Gas Production by Baker’s Yeast in Unsugared Dough
Authors: Higgins Vincent J. , Braidwood Mark , Bell Phil , Bissinger Peter , Dawes Ian W. , Attfield Paul V. 
Published: December 2019

ABSTRACT

Strain selection and improvement in the baker’s yeast industry have aimed to increase the speed of maltose fermentation in order to increase the leavening activity of industrial baking yeast. We identified two groups of baker’s strains of
Saccharomyces cerevisiae
that can be distinguished by the mode of regulation of maltose utilization. One group (nonlagging strains), characterized by rapid maltose fermentation, had at least 12-fold more maltase and 130-fold-higher maltose permease activities than maltose-lagging strains in the absence of inducing sugar (maltose) and repressing sugar (glucose). Increasing the noninduced maltase activity of a lagging strain 13-fold led to an increase in CO
2
production in unsugared dough. This increase in CO
2
production also was seen when the maltose permease activity was increased 55-fold. Only when maltase and maltose permease activities were increased in concert was CO
2
production by a lagging strain similar to that of a nonlagging strain. The noninduced activities of maltase and maltose permease constitute the largest determinant of whether a strain displays a nonlagging or a lagging phenotype and are dependent upon the
MALx3
allele. Previous strategies for strain improvement have targeted glucose derepression of maltase and maltose permease expression. Our results suggest that increasing noninduced maltase and maltose permease levels is an important target for improved maltose metabolism in unsugared dough.
Diversity of Arbuscular Mycorrhizal Fungus Populations in Heavy-Metal-Contaminated Soils
Authors: Del Val C. , Barea J. M. , Azcón-Aguilar C. 
Published: December 2019

ABSTRACT
High concentrations of heavy metals have been shown to adversely affect the size, diversity, and activity of microbial populations in soil. The aim of this work was to determine how the diversity of arbuscular mycorrhizal (AM) fungi is affected by the addition of sewage-amended sludge containing heavy metals in a long-term experiment. Due to the reduced number of indigenous AM fungal (AMF) propagules in the experimental soils, several host plants with different life cycles were used to multiply indigenous fungi. Six AMF ecotypes were found in the experimental soils, showing consistent differences with regard to their tolerance to the presence of heavy metals. AMF ecotypes ranged from very sensitive to the presence of metals to relatively tolerant to high rates of heavy metals in soil. Total AMF spore numbers decreased with increasing amounts of heavy metals in the soil. However, species richness and diversity as measured by the Shannon-Wiener index increased in soils receiving intermediate rates of sludge contamination but decreased in soils receiving the highest rate of heavy-metal-contaminated sludge. Relative densities of most AMF species were also significantly influenced by soil treatments. Host plant species exerted a selective influence on AMF population size and diversity. We conclude based on the results of this study that size and diversity of AMF populations were modified in metal-polluted soils, even in those with metal concentrations that were below the upper limits accepted by the European Union for agricultural soils.
Enhancement of Solar Water Pasteurization with Reflectors
Authors: Safapour Negar , Metcalf Robert H. 
Published: December 2019

ABSTRACT
A simple and reliable method that could be used in developing countries to pasteurize milk and water with solar energy is described. A cardboard reflector directs sunshine onto a black jar, heating water to pasteurizing temperatures in several hours. A reusable water pasteurization indicator verifies that pasteurization temperatures have been reached.
Isolation of Fecal Coliform Bacteria from the Diamondback Terrapin (
Malaclemys terrapin centrata
)
Authors: Harwood Valerie J. , Butler Joseph , Parrish Danny , Wagner Victoria 
Published: December 2019

ABSTRACT

Total and fecal coliform bacteria were isolated from the cloaca and feces of the estuarine diamondback terrapin. The majority of samples contained fecal coliforms.
Escherichia coli
was the predominant fecal coliform species isolated, and members of the genus
Salmonella
were isolated from 2 of 39 terrapins. Fecal coliform numbers are used to regulate shellfish harvests, and diamondback terrapins inhabit the brackish-water habitats where oyster beds are found; therefore, these findings have implications for the efficacy of current regulatory parameters in shellfishing waters.
Degradation of Chloronitrobenzenes by a Coculture of
Pseudomonas putida
and a
Rhodococcus
sp
Authors: Park Hee-Sung , Lim Sung-Jin , Chang Young Keun , Livingston Andrew G. , Kim Hak-Sung 
Published: December 2019

ABSTRACT

A single microorganism able to mineralize chloronitrobenzenes (CNBs) has not been reported, and degradation of CNBs by coculture of two microbial strains was attempted.
Pseudomonas putida
HS12 was first isolated by analogue enrichment culture using nitrobenzene (NB) as the substrate, and this strain was observed to possess a partial reductive pathway for the degradation of NB. From high-performance liquid chromatography-mass spectrometry and
1
H nuclear magnetic resonance analyses, NB-grown cells of
P. putida
HS12 were found to convert 3- and 4-CNBs to the corresponding 5- and 4-chloro-2-hydroxyacetanilides, respectively, by partial reduction and subsequent acetylation. For the degradation of CNBs,
Rhodococcus
sp. strain HS51, which degrades 4- and 5-chloro-2-hydroxyacetanilides, was isolated and combined with
P. putida
HS12 to give a coculture. This coculture was confirmed to mineralize 3- and 4-CNBs in the presence of an additional carbon source. A degradation pathway for 3- and 4-CNBs by the two isolated strains was also proposed.
Enhanced Bioaccumulation of Heavy Metal Ions by Bacterial Cells Due to Surface Display of Short Metal Binding Peptides
Authors: Kotrba Pavel , Dolečková Lucie , de Lorenzo Víctor , Ruml Tomas 
Published: December 2019

ABSTRACT

Metal binding peptides of sequences Gly-His-His-Pro-His-Gly (named HP) and Gly-Cys-Gly-Cys-Pro-Cys-Gly-Cys-Gly (named CP) were genetically engineered into LamB protein and expressed in
Escherichia coli
. The Cd
2+
-to-HP and Cd
2+
-to-CP stoichiometries of peptides were 1:1 and 3:1, respectively. Hybrid LamB proteins were found to be properly folded in the outer membrane of
E. coli
. Isolated cell envelopes of
E. coli
bearing newly added metal binding peptides showed an up to 1.8-fold increase in Cd
2+
binding capacity. The bioaccumulation of Cd
2+
, Cu
2+
, and Zn
2+
by
E. coli
was evaluated. Surface display of CP multiplied the ability of
E. coli
to bind Cd
2+
from growth medium fourfold. Display of HP peptide did not contribute to an increase in the accumulation of Cu
2+
and Zn
2+
. However, Cu
2+
ceased contribution of HP for Cd
2+
accumulation, probably due to the strong binding of Cu
2+
to HP. Thus, considering the cooperation of cell structures with inserted peptides, the relative affinities of metal binding peptide and, for example, the cell wall to metal ion should be taken into account in the rational design of peptide sequences possessing specificity for a particular metal.
Comparison of Flagellin Genes from Clinical and Environmental
Pseudomonas aeruginosa
Isolates
Authors: Morgan J. Alun W. , Bellingham Nessa F. , Winstanley Craig , Ousley Margaret A. , Hart C. Anthony , Saunders Jon R. 
Published: December 2019

ABSTRACT

Pseudomonas aeruginosa
, an important opportunistic pathogen, was isolated from environmental samples and compared to clinically derived strains. While
P. aeruginosa
was isolated readily from an experimental mushroom-growing unit, it was found only rarely in other environmental samples. A flagellin gene PCR-restriction fragment length polymorphism analysis of the isolates revealed that environmental and clinical
P. aeruginosa
strains are not readily distinguishable. The variation in the central regions of the flagellin genes of seven of the isolates was investigated further. The strains used included two strains with type a genes (998 bp), four strains with type b genes (1,258 bp), and one strain, K979, with a novel flagellin gene (2,199 bp). The route by which flagellin gene variation has occurred in
P. aeruginosa
is discussed.
Metabolic Engineering of a 1,2-Propanediol Pathway in
Escherichia coli
Authors: Altaras Nedim E. , Cameron Douglas C. 
Published: December 2019

ABSTRACT

1,2-Propanediol (1,2-PD) is a major commodity chemical that is currently derived from propylene, a nonrenewable resource. A goal of our research is to develop fermentation routes to 1,2-PD from renewable resources. Here we report the production of enantiomerically pure
R
-1,2-PD from glucose in
Escherichia coli
expressing NADH-linked glycerol dehydrogenase genes (
E. coli gldA
or
Klebsiella pneumoniae dhaD
). We also show that
E. coli
overexpressing the
E. coli
methylglyoxal synthase gene (
mgs
) produced 1,2-PD. The expression of either glycerol dehydrogenase or methylglyoxal synthase resulted in the anaerobic production of approximately 0.25 g of 1,2-PD per liter.
R
-1,2-PD production was further improved to 0.7 g of 1,2-PD per liter when methylglyoxal synthase and glycerol dehydrogenase (
gldA
) were coexpressed. In vitro studies indicated that the route to
R
-1,2-PD involved the reduction of methylglyoxal to
R
-lactaldehyde by the recombinant glycerol dehydrogenase and the reduction of
R
-lactaldehyde to
R
-1,2-PD by a native
E. coli
activity. We expect that
R
-1,2-PD production can be significantly improved through further metabolic and bioprocess engineering.
Organization of the Gene Cluster for Biosynthesis of Penicillin in
Penicillium nalgiovense
and Antibiotic Production in Cured Dry Sausages
Authors: Laich Federico , Fierro Francisco , Cardoza Rosa Elena , Martin Juan F. 
Published: December 2019

ABSTRACT

Several fungal isolates obtained from two cured meat products from Spain were identified as
Penicillium nalgiovense
by their morphological features and by DNA fingerprinting. All
P. nalgiovense
isolates showed antibiotic activity in agar diffusion assays, and their penicillin production in liquid complex medium ranged from 6 to 38 μg · ml
−1
. We constructed a restriction map of the penicillin gene cluster of
P. nalgiovense
and found that the organization of the penicillin biosynthetic genes (
pcbAB
,
pcbC
, and
penDE
) is the same as in
Penicillium chrysogenum
and
Aspergillus nidulans
. The
pcbAB
gene is located in an orientation opposite that of the
pcbC
and
penDE
genes in all three species. Significant amounts of penicillin were found in situ in the casing and the outer layer of salami meat during early stages of the curing process, coinciding with fungal colonization, but no penicillin was detected in the cured salami. The antibiotic produced in situ was sensitive to penicillinase.
Identification of Epichloë Endophytes In Planta by a Microsatellite-Based PCR Fingerprinting Assay with Automated Analysis
Authors: Moon Christina D. , Tapper Brian A. , Scott Barry 
Published: December 2019

ABSTRACTEpichloë endophytes are a group of filamentous fungi that include both sexual (Epichloë) and asexual (Neotyphodium) species. As a group they are genetically diverse and form both antagonistic and mutualistic associations with temperate grasses. We report here on the development of a microsatellite-based PCR system for fingerprinting this group of fungi with template isolated from either culture or infected plant material. M13mp19 partial genomic libraries were constructed for size-fractionated genomic DNA from two endophyte strains. These libraries were screened with a mixture of DIG-labeled dinucleotide and trinucleotide repeat probes. Positive clones were sequenced, and nine unique microsatellite loci were identified. An additional microsatellite was serendipitously identified in the 3′ untranscribed region of the 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase gene fromN. loliiLp19. Primers were designed for each locus and a panel of endophytes, from different taxonomic groupings, was screened to determine the degree of polymorphism. On the basis of these results a multiplex assay was developed for strain identification with fluorescently labeled primers for five of these loci. Using this system the size of the products amplified can be precisely determined by automated analysis, and an allele profile for each strain can be readily generated. The assay was shown to resolve endophyte groupings to the level of known isozyme phenotype groupings. In a blind test the assay was used successfully to identify a set of endophytes in planta. A reference database of allele sizes has been established for the panel of endophytes examined, and this will be expanded as new strains are analyzed.
A Randomly Amplified Polymorphic DNA Marker Specific for the
Bacillus cereus
Group Is Diagnostic for
Bacillus anthracis
Authors: Daffonchio Daniele , Borin Sara , Frova Giuseppe , Gallo Romina , Mori Elena , Fani Renato , Sorlini Claudia 
Published: December 2019

ABSTRACT

Aiming to develop a DNA marker specific for
Bacillus anthracis
and able to discriminate this species from
Bacillus cereus
,
Bacillus thuringiensis
, and
Bacillus mycoides
, we applied the randomly amplified polymorphic DNA (RAPD) fingerprinting technique to a collection of 101 strains of the genus
Bacillus
, including 61 strains of the
B. cereus
group. An 838-bp RAPD marker (SG-850) specific for
B. cereus
,
B. thuringiensis
,
B. anthracis
, and
B. mycoides
was identified. This fragment included a putative (366-nucleotide) open reading frame highly homologous to the
ypuA
gene of
Bacillus subtilis
. The restriction analysis of the SG-850 fragment with
Alu
I distinguished
B. anthracis
from the other species of the
B. cereus
group.
Thermus aquaticus
ATCC 33923 Amylomaltase Gene Cloning and Expression and Enzyme Characterization: Production of Cycloamylose
Authors: Terada Yoshinobu , Fujii Kazutoshi , Takaha Takeshi , Okada Shigetaka 
Published: December 2019

ABSTRACT

The amylomaltase gene of the thermophilic bacterium
Thermus aquaticus
ATCC 33923 was cloned and sequenced. The open reading frame of this gene consisted of 1,503 nucleotides and encoded a polypeptide that was 500 amino acids long and had a calculated molecular mass of 57,221 Da. The deduced amino acid sequence of the amylomaltase exhibited a high level of homology with the amino acid sequence of potato disproportionating enzyme (D-enzyme) (41%) but a low level of homology with the amino acid sequence of the
Escherichia coli
amylomaltase (19%). The amylomaltase gene was overexpressed in
E. coli
, and the enzyme was purified. This enzyme exhibited maximum activity at 75°C in a 10-min reaction with maltotriose and was stable at temperatures up to 85°C. When the enzyme acted on amylose, it catalyzed an intramolecular transglycosylation (cyclization) reaction which produced cyclic α-1,4-glucan (cycloamylose), like potato D-enzyme. The yield of cycloamylose produced from synthetic amylose with an average molecular mass of 110 kDa was 84%. However, the minimum degree of polymerization (DP) of the cycloamylose produced by
T. aquaticus
enzyme was 22, whereas the minimum DP of the cycloamylose produced by potato D-enzyme was 17. The
T. aquaticus
enzyme also catalyzed intermolecular transglycosylation of maltooligosaccharides. A detailed analysis of the activity of
T. aquaticus
ATCC 33923 amylomaltase with maltooligosaccharides indicated that the catalytic properties of this enzyme differ from those of
E. coli
amylomaltase and the plant D-enzyme.
Mechanism of Peroxidase Inactivation in Liquid Cultures of the Ligninolytic Fungus
Pleurotus pulmonarius
Authors: Böckle Brigitte , Martínez María Jesús , Guillén Francisco , Martínez Ángel T. 
Published: December 2019

ABSTRACT

It has recently been reported that
Pleurotus pulmonarius
secretes a versatile peroxidase that oxidizes Mn
2+
, as well as different phenolic and nonphenolic aromatic compounds; this enzyme has also been detected in other
Pleurotus
species and in
Bjerkandera
species. During culture production of the enzyme, the activity of the main peak was as high as 1,000 U/liter (measured on the basis of the Mn
3+
-tartrate formation) but this peak was very ephemeral due to enzyme instability (up to 80% of the activity was lost within 15 h). In culture filtrates inactivation was even faster; all peroxidase activity was lost within a few hours. Using different inhibitor compounds, we found that proteases were not responsible for the decrease in peroxidase activity. Peroxidase instability coincided with an increase in the H
2
O
2
concentration, which reached 200 μM when filtrates were incubated for several hours. It also coincided with the onset of biosynthesis of anisylic compounds and a decrease in the pH of the culture. Anisyl alcohol is the natural substrate of the enzyme aryl-alcohol oxidase, the main source of extracellular H
2
O
2
in
Pleurotus
cultures, and addition of anisyl alcohol to filtrates containing stable peroxidase activity resulted in rapid inactivation. A decrease in the culture pH could also dramatically affect the stability of the
P. pulmonarius
peroxidase, as shown by using pH values ranging from 6 to 3.25, which resulted in an increase in the level of inactivation by 10 μM H
2
O
2
from 5 to 80% after 1 h. Moreover, stabilization of the enzyme was observed after addition of catalase, Mn
2+
, or some phenols or after dialysis of the culture filtrate. We concluded that extracellular H
2
O
2
produced by the fungus during oxidation of aromatic metabolites is responsible for inactivation of the peroxidase and that the enzyme can protect itself in the presence of different reducing substrates.
Identification of Aerobically and Anaerobically Induced Genes in
Enterococcus faecalis
by Random Arbitrarily Primed PCR
Authors: Shepard Brett D. , Gilmore Michael S. 
Published: December 2019

ABSTRACT

Enterococci have emerged among the leading causes of nosocomial infection. With the goal of analyzing enterococcal genes differentially expressed in environments related to commensal or environmental colonization and infection sites, we adapted and optimized a method more commonly used in the study of eukaryotic gene expression, random arbitrarily primed PCR (RAP-PCR). The RAP-PCR method was systematically optimized, allowing the technique to be used in a highly reproducible manner with gram-positive bacterial RNA. In the present study, aerobiosis was chosen as a variable for the induction of changes in gene expression by
Enterococcus faecalis
. Aerobically and anaerobically induced genes were detected and identified to the sequence level, and differential gene expression was confirmed by quantitative, specifically primed RT-PCR. Differentially expressed genes included several sharing identity with those of other organisms related to oxygen metabolism, as well as hypothetical genes lacking identity to known genes.