bluuebunny/crossref_metadata_2025 huggingface dataset:
Characterization of New Hybridoma Clones Producing Monoclonal Antibodies Reactive Against Both Live and Heat‐Killed Listeria monocytogenes
Authors: Heo Seok A. , Nannapaneni Ramakrishna , Story Robert P. , Johnson Michael G.
Published: December 2006

ABSTRACT:  The objective of this study is to develop high affinity monoclonal antibody (MAb) probes recognizing all major serotypes of Listeria monocytogenes cells. From 500 candidate hybridoma clones, 2 new monoclonal antibody‐producing hybridomas were selected and evaluated. MAbs 22D10 and 24F6 reacted strongly with live cells of most serotypes of L. monocytogenes except 4c and 4e and with some L. innocua strains; MAb 22D10 reacted strongly with both live and heat‐killed cells (100 °C for 20 min) of Listeria. Both MAbs 22D10 and 24F6 did not show any cross‐reactions with the other non‐Listeria G(+) bacteria tested in ELISA. The mixture of EM‐7G1 and 22D10 or 24F6 reacted with all 13 major serotypes of live L. monocytogenes except serotype 4c, while none of these 3 MAbs when tested alone did so. MAb 22D10 mixed with 7G1 reacted with all heat‐killed L. monocytogenes serotypes except 4c and 4e. In Western blots, MAbs 22D10 and 24F6 reacted with 1 major protein band of 66 kDa in extracts from L. monocytogenes, but with 2 major protein bands of 66 kDa and 76 kDa in extracts from L. innocua. These results suggest that MAbs 22D10 and 24F6 have high affinity for 11 of 13 serotypes of L. monocytogenes, both live and heat‐killed cells. MAbs 22D10 and 24F6—in combination with species‐specific MAb EM‐7G1—should be useful candidates for use in an ELISA sandwich assays for detecting L. monocytogenes in RTE meat and poultry products.
Relationship between Physical Properties and Sensory Attributes of Carbonated Beverages
Authors: Kappes S.M. , Schmidt S.J. , Lee S.‐Y.
Published: December 2006

ABSTRACT: Bulk sweeteners provide functional properties in beverages, including sweet taste, bulking, bitter masking, structure, and mouthfeel. Diet beverages come closer to the taste of regular beverages using a blend of high‐intensity sweeteners; however, some properties, including bulking, structure, and mouthfeel, remain significantly different. Relating physical properties to sensory characteristics is an important step in understanding why mouthfeel differences are apparent in beverages sweetened with alternative sweeteners compared to bulk sweeteners. The objectives of this research were to (1) measure sweetener profile, Brix, refractive index, viscosity,aw, carbonation, titratable acidity, and pH of commercial carbonated beverages; and (2) correlate the physical property measurements to descriptive analysis of the beverages. Correlation analysis, partial least squares, canonical correlation analysis, and cluster analysis were used to analyze the data. Brix, viscosity, and sweet taste were highly correlated among one another and were all negatively correlated toaw. Carbonated and decarbonated pH were highly correlated to each other and were both negatively correlated to mouthcoating. Numbing, burn, bite, and carbonation were highly correlated to total acidity, citric acid, and ascorbic acid and negatively correlated to phosphoric acid. The mouthfeel difference between diet and regular lemon/lime carbonated beverages is small and may be related to overall differences between flavor, acid, and sweetener types and usage levels. This research is significant because it demonstrates the use of both sensory attributes and physical properties to identify types of ingredients and levels that may decrease the mouthfeel perception differences between regular and diet carbonated beverages, which could consequently lead to higher acceptance of diet beverages by the consumers of regular.
Antioxidant Capacity of Extracts from Wild and Crop Plants of the Mediterranean Region
Authors: Mohamed Refaei , Pineda Manuel , Aguilar Miguel
Published: December 2006

ABSTRACT:  There is an increasing demand for natural antioxidants to replace synthetic additives in the food industry. The present work examines the potential of some wild and cultivated plants from the Mediterranean region as sources of natural antioxidants. Samples of different organs and tissues from each of these species were extracted with aqueous and organic solvents and analyzed for their total hydrosoluble and lipid‐soluble antioxidant capacity, measured by the phosphomolybdenum method, and for their content in tocopherols. Our results demonstrate that there is more than a 1000‐fold difference among total antioxidants in various plant sources. The highest level of water‐soluble antioxidant capacity was found in fruit peel and leaf samples, while seeds presented the highest levels of lipid‐soluble antioxidant capacity. Tocopherols were more abundant in leaf samples. Holm oak and olive tree leaves showed the highest yields of α‐tocopherol. These Mediterranean plant species could be used as a very good source of both water‐soluble and lipid‐soluble antioxidants, particularly α‐tocopherol.
Effect of Filtered Wood Smoke Treatment on Chemical and Microbial Changes in Mahi Mahi Fillets
Authors: Kristinsson Hordur G. , Danyali Nineveh , Ua‐Angkoon Siriporn
Published: December 2006

ABSTRACT:  A study was performed to investigate the effect of filtered wood smoke processing on quality and safety of mahi mahi compared to no treatment. Skinless mahi mahi fillet portions were either treated with filtered smoke (FS) or left untreated for 24 h, followed by either (a) aerobic storage at 4 °C for 8 d or (b) freezing for 30 d (–25 °C) followed by thawing and aerobic storage at 4 °C for 8 d. Results show that treating mahi mahi fillets with FS increased (P < 0.05) a* values (redness) of the muscle and stabilized it during frozen storage. The redness did, however, decay (P < 0.05) rapidly on cold storage for both defrosted and fresh filtered‐smoke‐treated products, and reached initial (presmoking) redness levels in 2 d. The FS process overall significantly (P < 0.05) improved microbial stability of the product. Stability toward lipid oxidation was also significantly (P < 0.05) increased for the FS products compared to untreated products, particularly after defrosting. Sensory studies supported the microbial and lipid oxidation findings, showing that products treated with FS were better accepted and had increased (P < 0.05) shelf life over the untreated products. The shelf life was, however, compromised when microbial levels increased; that is, the process did not mask microbial spoilage; the spoilage did become evident in the sensory trials.
Muscle Tuning During Running: Implications of an Un-tuned Landing
Authors: Boyer Katherine A. , Nigg Benno M.
Published: December 2006

Background: The impact force in heel-toe running is an input signal into the body that initiates vibrations of the soft tissue compartments of the leg. These vibrations are heavily damped and the paradigm of muscle tuning suggests the body adapts to different input signals to minimize these vibrations. The objectives of the present study were to investigate the implications of not tuning a muscle properly for a landing with a frequency close to the resonance frequency of a soft tissue compartment and to look at the effect of an unexpected surface change on the subsequent step of running. Method: Thirteen male runners were recruited and performed heel-toe running over two surface conditions. The peak accelerations and biodynamic responses of the soft tissue compartments of the leg along with the EMG activity of related muscles were determined for expected soft, unexpected hard and expected hard landings. Results and Conclusions: For the unexpected hard landing there was a change in the input frequency of the impact force, shifting it closer to the resonance frequency of the soft tissue compartments. For the unexpected landing there was no muscle adaptation, as subjects did not know the running surface was going to change. In support of the muscle-tuning concept an increase in the soft tissue acceleration did occur. This increase was greater when the proximity of the input signal frequency was closer to the resonance frequency of the soft tissue compartment. Following the unexpected change in the input signal a change in pre-contact muscle activity to minimize soft tissue compartment vibrations was not found. This suggests if muscle tuning does occur it is not a continuous feedback response that occurs with every small change in the landing surface properties. In previous studies with significant adaptation periods to new input signals significant correlations between the changes in the input signal frequency and the EMG intensity have been shown, however, changes in soft tissue accelerations have not been found. The results of the present study showed that changes in these soft tissue accelerations can occur in response to a resonance frequency input signal when a muscle reaction has not happened.
An EFE Model on Skin-Sleeve Interactions During Arm Rotation
Authors: Xing Malcolm M. Q. , Sun Zhiguo , Pan Ning , Zhong Wen , Maibach Howard I.
Published: December 2006

Skin and garment constitute a dynamic contact system for human body comfort and protection. Although dermatological injuries due to fabric actions during human body movement are common, there is still no general guidance or standard for measuring or evaluating skin/garment contact interactions, especially, during intense sports. A three-dimensional explicit finite element (EFE) model combined with Augmented Lagrange algorithm (ALA) is developed to simulate interactions between skin and fabric during rotation of the arm. Normalized effective shear stresses at the interface between skin and the sleeve during the arm rotation are provided to reflect the severity of the interactions. The effects due to changes in fabric properties, fabric-skin gap, and arm rotation rate are also illustrated. It has been demonstrated from our predictions that factors such as elastic modulus, friction coefficients, density of fabric, and the initial gap between skin and fabric influence significantly the shear stress and thus the discomfort and even injury potential to skin during intensive body movement such as sports and military. Thus this study for the first time confirms quantitatively that poorly chosen fabric with inappropriate garment design renders adverse actions on human skin.
Statistical Hemodynamics: A Tool for Evaluating the Effect of Fluid Dynamic Forces on Vascular Biology In Vivo
Authors: Friedman Morton H. , Himburg Heather A. , LaMack Jeffrey A.
Published: December 2006

Background. In vivo experimentation is the most realistic approach for exploring the vascular biological response to the hemodynamic stresses that are present in life. Post-mortem vascular casting has been used to define the in vivo geometry for hemodynamic simulation; however, this procedure damages or destroys the tissue and cells on which biological assays are to be performed. Method of Approach. Two statistical approaches, regional (RSH) and linear (LSH) statistical hemodynamics, are proposed and illustrated, in which flow simulations from one series of experiments are used to define a best estimate of the hemodynamic environment in a second series. As an illustration of the technique, RSH is used to compare the gene expression profiles of regions of the proximal external iliac arteries of swine exposed to different levels of time-average shear stress. Results. The results indicate that higher shears promote a more atheroprotective expression phenotype in porcine arterial endothelium. Conclusion. Statistical hemodynamics provides a realistic estimate of the hemodynamic stress on vascular tissue that can be correlated against biological response.
Effect of Viscosity on Instilled Perfluorocarbon Distribution in Rabbit Lungs
Authors: Tredici Stefano , Tredici Francesco , Brant David O. , Hirschl Ronald B. , Bull Joseph L.
Published: December 2006

The effect of viscosity on the distribution of perfluorocarbon instilled into the lungs for liquid ventilation was investigated. Perfluorocarbon (either perfluorodecalin or FC‐3283) was instilled into the trachea during ventilation at a constant infusion rate of 40ml∕min and radiographic images were obtained at 30frames∕s. Image analysis was performed and the homogeneity index of the distribution was computed for images at the end of inspiration of each breath to evaluate the evolution of perfluorocarbon distribution during filling. The higher viscosity perfluorocarbon (perfluorodecalin) resulted in a more homogeneous distribution. This was attributed to perfluorodecalin’s higher propensity to form liquid plugs in large airways and to those plugs leaving behind a thicker liquid layer as they propagated through the lungs.
Viscoelasticity of Esophageal Tissue and Application of a QLV Model
Authors: Yang W. , Fung T. C. , Chian K. S. , Chong C. K.
Published: December 2006

The time-dependent mechanical properties of the porcine esophagus were investigated experimentally and theoretically. It was hypothesized that the viscoelasticity was quasilinear, i.e., the time and strain effects were independent. In order to verify the separability of time and strain effects, the stress-relaxation test was conducted at various strains and the data were fitted with the Fung’s quasilinear viscoelastic (QLV) model. By using the material parameters obtained from the stress relaxation test, the cyclic peak stress and hysteresis were predicted. Results showed that the stress relaxed by 20–30% of the peak stress within the first 10s and stabilized at ∼50% at the time of 300s. The relative stress relaxation R2 (i.e., the difference of stress at a particular time to the final equilibrium stress normalized by the total difference of the peak and final stress) was not different significantly for various strains. It was also found that, by using the stress-time data during both the ramp and relaxation phases, the correlation between parameters was substantially reduced. The model could also predict the cyclic peak stress and hysteresis except for the underestimate of valley stress. We conclude that the QLV model could be used as the material characterization of the esophageal tissue.
Protection of Mice from Lethal
Escherichia coli
Infection by ChimericHuman Bactericidal/Permeability-Increasing Protein and ImmunoglobulinG1 Fc Gene Delivery
Authors: Chen Jindong , Li Chengyao , Guan Yuanzhi , Kong Qingli , Li Chen , Guo Xianghua , Chen Qinghua , Jing Xuefang , Lv Zhe , An Yunqing
Published: December 2006

To
evaluate the potentiality of applying gene therapy to bacterial
infections, especially for preventing infection in high-risk patients,
we investigated protection of mice from challenge with lethal
Escherichia coli
infection by adeno-associated virus serotype
2 (AAV2)-mediated gene transfer of a chimeric
BPI
23
-Fcγ1 gene, which consisted of human
bactericidal/permeability-increasing protein (BPI) gene encoding the
functional N terminus (amino acid residues 1 to 199) of human BPI and
an Fcγ1 gene encoding the Fc segment of human immunoglobulin
G1. Here we show that the target protein that was expressed and
secreted into the serum of the gene-transferred mice demonstrated the
activity of a neutralizing endotoxin, killing
E. coli
and
mediating opsonization. After lethal
E. coli
infection, the
count of bacteria and the levels of endotoxin and proinflammatory
cytokines in the gene-transferred mice were decreased. The survival
rate of BPI
23
-Fcγ1 gene-transferred mice markedly
increased, especially in conjunction with antibiotics. Our data suggest
that AAV2-mediated chimeric BPI
23
-Fcγ1 gene delivery
could potentially be used clinically for the protection and treatment
of infection with gram-negative bacteria in high-risk
individuals.
Moxifloxacin, Ofloxacin, Sparfloxacin, and Ciprofloxacin against
Mycobacterium tuberculosis
: Evaluation of In Vitro and Pharmacodynamic Indices That Best Predict In Vivo Efficacy
Authors: Shandil Radha K. , Jayaram Ramesh , Kaur Parvinder , Gaonkar Sheshagiri , Suresh B. L. , Mahesh B. N. , Jayashree R. , Nandi Vrinda , Bharath Sowmya , Balasubramanian V.
Published: December 2006

Members of the fluoroquinolone class are being actively evaluated for inclusion in tuberculosis chemotherapy regimens, and we sought to determine the best in vitro and pharmacodynamic predictors of in vivo efficacy in mice. MICs for
Mycobacterium tuberculosis
H37Rv were 0.1 mg/liter (sparfloxacin [SPX]) and 0.5 mg/liter (moxifloxacin [MXF], ciprofloxacin [CIP], and ofloxacin [OFX]). The unbound fraction in the presence of murine serum was concentration dependent for MXF, OFX, SPX, and CIP. In vitro time-kill studies revealed a time-dependent effect, with the CFU reduction on day 7 similar for all four drugs. However, with a J774A.1 murine macrophage tuberculosis infection model, CIP was ineffective at up to 32× MIC. In addition, MXF, OFX, and SPX exhibited less activity than had been seen in the in vitro time-kill study. After demonstrating that the area under the concentration-time curve (AUC) and maximum concentration of drug in plasma were proportional to the dose in vivo, dose fractionation studies with total oral doses of 37.5 to 19,200 mg/kg of body weight (MXF), 225 to 115,200 mg/kg (OFX), 30 to 50,000 mg/kg (SPX), and 38 to 100,000 mg/kg (CIP) were performed with a murine aerosol infection model. MXF was the most efficacious agent (3.0 ± 0.2 log
10
CFU/lung reduction), followed by SPX (1.4 ± 0.1) and OFX (1.5 ± 0.1). CIP showed no effect. The ratio of the AUC to the MIC was the pharmacodynamic parameter that best described the in vivo efficacy. In summary, a lack of intracellular killing predicted the lack of in vivo activity of CIP. The in vivo rank order for maximal efficacy of the three active fluoroquinolones was not clearly predicted by the in vitro assays, however.
Length Effects in Antimicrobial Peptides of the (RW)nSeries
Authors: Liu Zhigang , Brady Anna , Young Anne , Rasimick Brian , Chen Kang , Zhou Chunhui , Kallenbach Neville R.
Published: December 2006

A class of antimicrobial peptides involved in host defense consists of sequences rich in Arg and Trp-R and -W. Analysis of the pharmacophore in these peptides revealed that chains as short as trimers of sequences such as WRW and RWR have antimicrobial activity (M. B. Strom, B. E. Haug, M. L. Skar, W. Stensen, T. Stiberg, and J. S. Svendsen, J. Med. Chem. 46:1567-1570, 2003). To evaluate the effect of chain length on antimicrobial activity, we synthesized a series of peptides containing simple sequence repeats, (RW)n-NH2(wherenequals 1, 2, 3, 4, or 5), and determined their antimicrobial and hemolytic activity. The antimicrobial activity of the peptides increases with chain length, as does the hemolysis of red blood cells. Within the experimental error, longer peptides (nequals 3, 4, or 5) show similar values for the ratio of hemolytic activity to antibacterial activity, or the hemolytic index. The (RW)3represents the optimal chain length in terms of the efficacy of synthesis and selectivity as evaluated by the hemolytic index. Circular dichroism spectroscopy indicates that these short peptides appear to be unfolded in aqueous solution but acquire structure in the presence of phospholipids. Interaction of the peptides with model lipid vesicles was examined using tryptophan fluorescence. The (RW)npeptides preferentially interact with bilayers containing the negatively charged headgroup phosphatidylglycerol relative to those containing a zwitterionic headgroup, phosphatidylcholine.
Cathepsin A Is the Major Hydrolase Catalyzing the Intracellular Hydrolysis of the Antiretroviral Nucleotide Phosphonoamidate Prodrugs GS-7340 and GS-9131
Authors: Birkus Gabriel , Wang Ruth , Liu Xiaohong , Kutty Nilima , MacArthur Holly , Cihlar Tomas , Gibbs Craig , Swaminathan Swami , Lee William , McDermott Martin
Published: December 2006

GS-7340 and GS-9131 {9-[(R)-2-[[(S)-[[(S)-1-(isopropoxycarbonyl)ethyl]amino]phenoxyphosphinyl]methoxy]-propyl]adenine and 9-(R)-4′-(R)-[[[(S)-1-[(ethoxycarbonyl)ethyl]amino]phenoxyphosphinyl]methoxy]-2′-fluoro-1′-furanyladenine, respectively} are novel alkylalaninyl phenyl ester prodrugs of tenofovir {9-R-[(2-phosphonomethoxy)propyl]adenine} (TFV) and a cyclic nucleotide analog, GS-9148 (phosphonomethoxy-2′-fluoro-2′, 3′-dideoxydidehydroadenosine), respectively. Both prodrugs exhibit potent antiretroviral activity against both wild-type and drug-resistant human immunodeficiency virus type 1 strains and excellent in vivo pharmacokinetic properties. In this study, the main enzymatic activity responsible for the initial step in the intracellular activation of GS-7340 and GS-9131 was isolated from human peripheral blood mononuclear cells and identified as lysosomal carboxypeptidase A (cathepsin A [CatA]; EC 3.4.16.5). Biochemical properties of the purified hydrolase (native complex and catalytic subunit molecular masses of 100 and 29 kDa, respectively; isoelectric point [pI] of 5.5) matched those of CatA. Recombinant CatA and the isolated prodrug hydrolase displayed identical susceptibilities to inhibitors and identical substrate preferences towards a panel of tenofovir phosphonoamidate prodrugs. Incubation of both enzymes with14C-labeled GS-7340 or [3H]difluorophosphonate resulted in the covalent labeling of identical 29-kDa catalytic subunits. Finally, following a 4-h incubation with GS-7340 and GS-9131, the intracellular concentrations of prodrug metabolites detected in CatA-negative fibroblasts were approximately 7.5- and 3-fold lower, respectively, than those detected in normal control fibroblasts. Collectively, these data demonstrate the key role of CatA in the intracellular activation of nucleotide phosphonoamidate prodrugs and open new possibilities for further improvement of this important class of antiviral prodrugs.
Plasmid-Mediated KPC-2 in a
Klebsiella pneumoniae
Isolate from China
Authors: Wei Ze-Qing , Du Xiao-Xing , Yu Yun-Song , Shen Ping , Chen Ya-Gang , Li Lan-Juan
Published: December 2006

A carbapenem-resistant isolate of
Klebsiella pneumoniae
producing class A carbapenemase KPC-2 was identified in Zhejiang, China. The KPC-2 gene was located on an approximately 60-kb plasmid in a genetic environment partially different from that of
bla
KPC-2
in the isolates from the United States and Colombia.
Mechanism of Antimalarial Action of the Synthetic Trioxolane RBX11160 (OZ277)
Authors: Uhlemann Anne-Catrin , Wittlin Sergio , Matile Hugues , Bustamante Leyla Y. , Krishna Sanjeev
Published: December 2006

RBX11160 (OZ277) is a fully synthetic peroxidic antimalarial in clinical development. To study the possible mechanisms of action of RBX11160, we have examined its ability to inhibit PfATP6, a sarcoplasmic reticulum calcium ATPase and proposed target for semisynthetic peroxidic artemisinin derivatives. RBX11160 inhibits PfATP6 (apparent half-maximal inhibitory constant = 7,700 nM) less potently than artemisinin (79 nM). Inhibition of PfATP6 is abrogated by desferrioxamine, an iron-chelating agent. Consistent with this finding, the killing of
Plasmodium falciparum
organisms by RBX11160 in vitro is antagonized by desferrioxamine. Artesunate and RBX11160 also act antagonistically against
P. falciparum
in vitro. A fluorescent derivative of RBX11160 localizes to the parasite cytosol in some parasites and to the food vacuole in other parasites. These data demonstrate that there are both similarities and differences between the antimalarial properties of RBX11160 and those of semisynthetic antimalarials such as artesunate and artemisinin.
Aspirin Antagonism in Isoniazid Treatment of Tuberculosis in Mice
Authors: Byrne Sean T. , Denkin Steven M. , Zhang Ying
Published: December 2006

Salicylate has previously been shown to reduce the susceptibility of
Mycobacterium tuberculosis
to several drugs in vitro. In this study, aspirin, a salicylate anti-inflammatory, antagonized isoniazid treatment of murine pulmonary tuberculosis, whereas the nonsalicylate ibuprofen did not. These results may have implications on concurrent administration of anti-inflammatory and antituberculosis drugs.
Relationships between
Mycobacterium
Isolates from Patients with Pulmonary Mycobacterial Infection and Potting Soils
Authors: De Groote Mary Ann , Pace Norman R. , Fulton Kayte , Falkinham Joseph O.
Published: December 2006

High numbers of mycobacteria, including known pathogenic species such as
Mycobacterium avium
,
Mycobacterium intracellulare
, and
Mycobacterium chelonae
, were recovered from aerosols produced by pouring commercial potting soil products and potting soil samples provided by patients with pulmonary mycobacterial infections. The dominant mycobacteria in the soil samples corresponded to the dominant species implicated clinically. Profiles of large restriction fragments obtained by pulsed-field gel electrophoresis demonstrated a closely related pair of
M. avium
isolates recovered from a patient and from that patient's own potting soil. Thus, potting soils are potential sources of infection by environmental mycobacteria. Use of dust-excluding masks should be considered during potting or other activities that generate aerosol with soil.
Isolation and Molecular Characterization of Chitinase-Deficient
Bacillus licheniformis
Strains Capable of Deproteinization of Shrimp Shell Waste To Obtain Highly Viscous Chitin
Authors: Waldeck Jens , Daum Gabriele , Bisping Bernward , Meinhardt Friedhelm
Published: December 2006

Proteolytic but chitinase-deficient microbial cultures were isolated from shrimp shell waste and characterized. The most efficient isolate was found to be a mixed culture consisting of two
Bacillus licheniformis
strains, which were first determined microscopically and physiologically. Molecular characterization was carried out by sequencing the 16S rRNA gene of both strains. According to the residual protein and ash content, the chitin obtained by fermentation of such a mixed culture was found to be comparable to a commercially available, chemically processed product. However, the strikingly high viscosity (80 versus 10 mPa of the commercially available sample) indicates its superior quality. The two strains differed in colony morphology and in their secretion capabilities for degradative extracellular enzymes. Sequencing of the loci encoding amylase, cellulase, chitinases, and proteases, as well as the
degS/degU
operon, which is instrumental in the regulation of degradative enzymes, and the
pga
operon, which is responsible for polyglutamic acid production, revealed no differences. However, a frameshift mutation in
chiA
, encoding a chitinase, was validated for both strains, providing an explanation for the ascertained absence of chitinolytic activities and the concomitant possibility of producing highly viscous chitin in a fermentational deproteinization process.
A Naturally Occurring Novel Allele of
Escherichia coli
Outer Membrane Protein A Reduces Sensitivity to Bacteriophage
Authors: Power Michelle L. , Ferrari Belinda C. , Littlefield-Wyer Jane , Gordon David M. , Slade Martin B. , Veal Duncan A.
Published: December 2006

A novel
Escherichia coli
outer membrane protein A (OmpA) was discovered through a proteomic investigation of cell surface proteins. DNA polymorphisms were localized to regions encoding the protein's surface-exposed loops which are known phage receptor sites. Bacteriophage sensitivity testing indicated an association between bacteriophage resistance and isolates having the novel
ompA
allele.
Aerobic Growth of
Escherichia coli
with 2,4,6-Trinitrotoluene (TNT) as the Sole Nitrogen Source and Evidence of TNT Denitration by Whole Cells and Cell-Free Extracts
Authors: Stenuit Ben , Eyers Laurent , Rozenberg Raoul , Habib-Jiwan Jean-Louis , Agathos Spiros N.
Published: December 2006

Escherichia coli
grew aerobically with 2,4,6-trinitrotoluene (TNT) as sole nitrogen source and caused TNT's partial denitration. This reaction was enhanced in nongrowing cell suspensions with 0.516 mol nitrite released per mol TNT. Cell extracts denitrated TNT in the presence of NAD(P)H. Isomers of amino-dimethyl-tetranitrobiphenyl were detected and confirmed with U-
15
N-labeled TNT.
Resistance of
Listeria monocytogenes
Biofilms to Sanitizing Agents in a Simulated Food Processing Environment
Authors: Pan Y. , Breidt F. , Kathariou S.
Published: December 2006

The objective of this study was to evaluate the resistance of biofilms of
Listeria monocytogenes
to sanitizing agents under laboratory conditions simulating a food processing environment. Biofilms were initially formed on stainless steel and Teflon coupons using a five-strain mixture of
L. monocytogenes
. The coupons were then subjected to repeated 24-h daily cycles. Each cycle consisted of three sequential steps: (i) a brief (60 s) exposure of the coupons to a sanitizing agent (a mixture of peroxides) or saline as a control treatment, (ii) storage of the coupons in sterile plastic tubes without any nutrients or water for 15 h, (iii) and incubation of the coupons in diluted growth medium for 8 h. This regimen was repeated daily for up to 3 weeks and was designed to represent stresses encountered by bacteria in a food processing environment. The bacteria on the coupons were reduced in number during the first week of the simulated food processing (SFP) regimen, but then adapted to the stressful conditions and increased in number. Biofilms repeatedly exposed the peroxide sanitizer in the SFP regimen developed resistance to the peroxide sanitizer as well as other sanitizers (quaternary ammonium compounds and chlorine). Interestingly, cells that were removed from the biofilms on peroxide-treated and control coupons were not significantly different in their resistance to sanitizing agents. These data suggest that the resistance of the treated biofilms to sanitizing agents may be due to attributes of extracellular polymeric substances and is not an intrinsic attribute of the cells in the biofilm.
Induction of Heavy-Metal-Transporting CPX-Type ATPases during Acid Adaptation in
Lactobacillus bulgaricus
Authors: Penaud S. , Fernandez A. , Boudebbouze S. , Ehrlich S. D. , Maguin E. , van de Guchte M.
Published: December 2006

Lactobacillus bulgaricus
is a lactic acid bacteria (LAB) that, through the production of lactic acid, gradually acidifies its environment during growth. In the course of this process,
L. bulgaricus
acquires an improved tolerance to acidity. A survey of the recently established genome sequence shows that this bacterium possesses few of the pH control functions that have been described in other LAB and raises the question of what other mechanisms could be involved in its adaptation to the decreasing environmental pH. In some bacteria other than LAB, ion transport systems have been implicated in acid adaptation. We therefore studied the expression of this type of transport system during acid adaptation in
L. bulgaricus
by reverse transcription and real-time quantitative PCR and mapped transcription start sites. Intriguingly, the most significantly induced were three ATPases carrying the CPX signature of heavy-metal transporters. Protein homology and the presence of a conserved sequence motif in the promoter regions of the genes encoding these proteins strongly suggest that they are involved in copper homeostasis. Induction of this system is thought to assist in avoiding indirect damage that could result from medium acidification.
Different Levels of Transcriptional Regulation Due to Trophic Constraints in the Reduced Genome of
Buchnera aphidicola
APS
Authors: Reymond Nancie , Calevro Federica , Viñuelas José , Morin Nicolas , Rahbé Yvan , Febvay Gérard , Laugier Christian , Douglas Angela , Fayard Jean-Michel , Charles Hubert
Published: December 2006

Symbiotic associations involving intracellular microorganisms and animals are widespread, especially for species feeding on poor or unbalanced diets.
Buchnera aphidicola
, the obligate intracellular bacterium associated with most aphid species, provides its hosts with essential amino acids (EAAs), nutrients in short supply in the plant phloem sap. The
Buchnera
genome has undergone severe reductions during intracellular evolution. Genes for EAA biosynthesis are conserved, but most of the transcriptional regulatory elements are lost. This work addresses two main questions: is transcription in
Buchnera
(i) regulated and (ii) scaled to aphid EAA demand? Two microarray experiments were designed for profiling the gene expression in
Buchnera
. The first one was characterized by a specific depletion of tyrosine and phenylalanine in the aphid diet, and the second experiment combined a global diminution of EAAs in the aphid diet with a sucrose concentration increase to manipulate the aphid growth rate. Aphid biological performance and budget analysis (the balance between EAAs provided by the diet and those synthesized by
Buchnera
) were performed to quantify the nutritional demand from the aphids toward their symbiotic bacteria. Despite the absence of known regulatory elements, a significant transcriptional regulation was observed at different levels of organization in the
Buchnera
genome: between genes, within putative transcription units, and within specific metabolic pathways. However, unambiguous evidence for transcriptional changes underpinning the scaling of EAA biosynthesis to aphid demand was not obtained. The phenotypic relevance of the transcriptional response from the reduced genome of
Buchnera
is addressed.
Molecular Detection of Epiphytic
Acaryochloris
spp. on Marine Macroalgae
Authors: Ohkubo Satoshi , Miyashita Hideaki , Murakami Akio , Takeyama Haruko , Tsuchiya Tohru , Mimuro Mamoru
Published: December 2006

A molecular method for detecting the epiphyte community on marine macroalgae was developed by using PCR-denaturing gradient gel electrophoresis. Selective amplification of 16S rRNA gene fragments from either cyanobacteria or algal plastids improved the detection of minor epiphytes. Two phylotypes of
Acaryochloris
, a chlorophyll
d
-containing cyanobacterium, were found not only on red macroalgae but also on green and brown macroalgae.
Virus-Specific Responses of
Heterosigma akashiwo
to Infection
Authors: Lawrence Janice E. , Brussaard Corina P. D. , Suttle Curtis A.
Published: December 2006

We used flow cytometry to examine the process of cell death in the bloom-forming alga
Heterosigma akashiwo
during infection by a double-stranded DNA virus (OIs1) and a single-stranded RNA virus (
H. akashiwo
RNA virus [HaRNAV]). These viruses were isolated from the same geographic area and infect the same strain of
H. akashiwo
. By use of the live/dead stains fluorescein diacetate and SYTOX green as indicators of cellular physiology, cells infected with OIs1 showed signs of infection earlier than HaRNAV-infected cultures (6 to 17 h versus 23 to 29 h). Intracellular esterase activity was lost prior to increased membrane permeability during infection with OIs1, while the opposite was seen with HaRNAV-infected cultures. In addition, OIs1-infected cells accumulated in the cultures while HaRNAV-infected cells rapidly disintegrated. Progeny OIs1 viruses consisted of large and small morphotypes with estimated latent periods of 11 and 17 h, respectively, and about 1,100 and 16,000 viruses produced per cell, respectively. In contrast, HaRNAV produced about 21,000 viruses per cell and had a latent period of 29 h. This study reveals that the characteristics of viral infection in algae are virus dependent and therefore are variable among viruses infecting the same species. This is an important consideration for ecosystem modeling exercises; calculations based on in situ measurements of algal physiology must be sensitive to the diverse responses of algae to viral infection.
Evidence for Horizontal Gene Transfer as Origin of Putrescine Production in
Oenococcus oeni
RM83
Authors: Marcobal Ángela , de las Rivas Blanca , Moreno-Arribas M. Victoria , Muñoz Rosario
Published: December 2006

The nucleotide sequence of a 17.2-kb chromosomal DNA fragment containing the
odc
gene encoding ornithine decarboxylase has been determined in the putrescine producer
Oenococcus oeni
RM83. This DNA fragment contains 13 open reading frames, including genes coding for five transposases and two phage proteins. This description might represent the first evidence of a horizontal gene transfer event as the origin of a biogenic amine biosynthetic locus.
Intracellular Accumulation of Trehalose Protects
Lactococcus lactis
from Freeze-Drying Damage and Bile Toxicity and Increases Gastric Acid Resistance
Authors: Termont Sofie , Vandenbroucke Klaas , Iserentant Dirk , Neirynck Sabine , Steidler Lothar , Remaut Erik , Rottiers Pieter
Published: December 2006

Interleukin-10 (IL-10) is a promising candidate for the treatment of inflammatory bowel disease. Intragastric administration of
Lactococcus lactis
genetically modified to secrete IL-10 in situ in the intestine was shown to be effective in healing and preventing chronic colitis in mice. However, its use in humans is hindered by the sensitivity of
L. lactis
to freeze-drying and its poor survival in the gastrointestinal tract. We expressed the trehalose synthesizing genes from
Escherichia coli
under control of the nisin-inducible promoter in
L. lactis
. Induced cells accumulated intracellular trehalose and retained nearly 100% viability after freeze-drying, together with a markedly prolonged shelf life. Remarkably, cells producing trehalose were resistant to bile, and their viability in human gastric juice was enhanced. None of these effects were seen with exogenously added trehalose. Trehalose accumulation did not interfere with IL-10 secretion or with therapeutic efficacy in murine colitis. The newly acquired properties should enable a larger proportion of the administered bacteria to reach the gastrointestinal tract in a bioactive form, providing a means for more effective mucosal delivery of therapeutics.
Metabolism of Thioamides byRalstonia pickettiiTA
Authors: Dodge Anthony G. , Richman Jack E. , Johnson Gilbert , Wackett Lawrence P.
Published: December 2006

Information on bacterial thioamide metabolism has focused on transformation of the antituberculosis drug ethionamide and related compounds byMycobacterium tuberculosis. To study this metabolism more generally, a bacterium that grew using thioacetamide as the sole nitrogen source was isolated via enrichment culture. The bacterium was identified asRalstonia pickettiiand designated strain TA. Cells grown on thioacetamide also transformed other thioamide compounds. Transformation of the thioamides tested was dependent on oxygen. During thioamide degradation, sulfur was detected in the medium at the oxidation level of sulfite, further suggesting an oxygenase mechanism.R. pickettiiTA did not grow on thiobenzamide as a nitrogen source, but resting cells converted thiobenzamide to benzamide, with thiobenzamide S-oxide and benzonitrile detected as intermediates. Thioacetamide S-oxide was detected as an intermediate during thioacetamide degradation, but the only accumulating metabolite of thioacetamide was identified as 3,5-dimethyl-1,2,4-thiadiazole, a compound shown to derive from spontaneous reaction of thioacetamide and oxygenated thioacetamide species. This dead-end metabolite accounted for only ca. 12% of the metabolized thioacetamide. Neither acetonitrile nor acetamide was detected during thioacetamide degradation, butR. pickettiigrew on both compounds as nitrogen and carbon sources. It is proposed thatR. pickettiiTA degrades thioamides via a mechanism involving consecutive oxygenations of the thioamide sulfur atom.
Identification of Novel Benzoylformate Decarboxylases by Growth Selection
Authors: Henning Helge , Leggewie Christian , Pohl Martina , Müller Michael , Eggert Thorsten , Jaeger Karl-Erich
Published: December 2006

A growth selection system was established using
Pseudomonas putida
, which can grow on benzaldehyde as the sole carbon source. These bacteria presumably metabolize benzaldehyde via the β-ketoadipate pathway and were unable to grow in benzoylformate-containing selective medium, but the growth deficiency could be restored by expression in
trans
of genes encoding benzoylformate decarboxylases. The selection system was used to identify three novel benzoylformate decarboxylases, two of them originating from a chromosomal library of
P. putida
ATCC 12633 and the third from an environmental-DNA library. The novel
P. putida
enzymes BfdB and BfdC exhibited 83% homology to the benzoylformate decarboxylase from
P. aeruginosa
and 63% to the enzyme MdlC from
P. putida
ATCC 12633, whereas the metagenomic BfdM exhibited 72% homology to a putative benzoylformate decarboxylase from
Polaromonas naphthalenivorans
. BfdC was overexpressed in
Escherichia coli
, and the enzymatic activity was determined to be 22 U/ml using benzoylformate as the substrate. Our results clearly demonstrate that
P. putida
KT2440 is an appropriate selection host strain suitable to identify novel benzoylformate decarboxylase-encoding genes. In principle, this system is also applicable to identify a broad range of different industrially important enzymes, such as benzaldehyde lyases, benzoylformate decarboxylases, and hydroxynitrile lyases, which all catalyze the formation of benzaldehyde.
Characterization of a
Vibrio alginolyticus
Strain, Isolated from Alaskan Oysters, Carrying a Hemolysin Gene Similar to the Thermostable Direct Hemolysin-Related Hemolysin Gene (
trh
) of
Vibrio parahaemolyticus
Authors: González-Escalona Narjol , Blackstone George M. , DePaola Angelo
Published: December 2006

A
Vibrio
strain isolated from Alaskan oysters and classified by its biochemical characteristics as
Vibrio alginolyticus
possessed a thermostable direct hemolysin-related hemolysin (
trh
) gene previously reported only in
Vibrio parahaemolyticus
. This
trh
-like gene was cloned and sequenced and was 98% identical to the
trh2
gene of
V. parahaemolyticus
. This gene seems to be functional since it was transcriptionally active in early-stationary-phase growing cells. To our knowledge, this is the first report of
V. alginolyticus
possessing a
trh
gene.
Tvbgn3, a β-1,6-Glucanase from the Biocontrol Fungus
Trichoderma virens
, Is Involved in Mycoparasitism and Control of
Pythium ultimum
Authors: Djonović Slavica , Pozo Maria J. , Kenerley Charles M.
Published: December 2006

Even though β-1,6-glucanases have been purified from several filamentous fungi, the physiological function has not been conclusively established for any species. In the present study, the role of Tvbgn3, a β-1,6-glucanase from
Trichoderma virens
, was examined by comparison of wild-type (WT) and transformant strains in which Tv
bgn3
was disrupted (GKO) or constitutively overexpressed (GOE). Gene expression analysis revealed induction of Tv
bgn3
in the presence of host fungal cell walls, indicating regulation during mycoparasitism. Indeed, while deletion or overexpression of Tv
bgn3
had no evident effect on growth and development, GOE and GKO strains showed an enhanced or reduced ability, respectively, to inhibit the growth of the plant pathogen
Pythium ultimum
compared to results with the WT. The relevance of this activity in the biocontrol ability of
T. virens
was confirmed in plant bioassays. Deletion of the gene resulted in levels of disease protection that were significantly reduced from WT levels, while GOE strains showed a significantly increased biocontrol capability. These results demonstrate the involvement of β-1,6-glucanase in mycoparasitism and its relevance in the biocontrol activity of
T. virens
, opening a new avenue for biotechnological applications.
Diversity and Significance of Mold Species in Norwegian Drinking Water
Authors: Hageskal Gunhild , Knutsen Ann Kristin , Gaustad Peter , de Hoog G. Sybren , Skaar Ida
Published: December 2006

In order to determine the occurrence, distribution, and significance of mold species in groundwater- and surface water-derived drinking water in Norway, molds isolated from 273 water samples were identified. Samples of raw water, treated water, and water from private homes and hospital installations were analyzed by incubation of 100-ml membrane-filtered samples on dichloran-18% glycerol agar. The total count (number of CFU per 100 ml) of fungal species and the species diversity within each sample were determined. The identification of mold species was based on morphological and molecular methods. In total, 94 mold species belonging to 30 genera were identified. The mycobiota was dominated by species of
Penicillium
,
Trichoderma
, and
Aspergillus
, with some of them occurring throughout the drinking water system. Several of the same species as isolated from water may have the potential to cause allergic reactions or disease in humans. Other species are common contaminants of food and beverages, and some may cause unwanted changes in the taste or smell of water. The present results indicate that the mycobiota of water should be considered when the microbiological safety and quality of drinking water are assessed. In fact, molds in drinking water should possibly be included in the Norwegian water supply and drinking water regulations.
Roles for Cell Wall Glycopeptidolipid in Surface Adherence and Planktonic Dispersal ofMycobacterium avium
Authors: Freeman Robert , Geier Henriette , Weigel Kris M. , Do Julie , Ford Timothy E. , Cangelosi Gerard A.
Published: December 2006

The opportunistic pathogenMycobacterium aviumis a significant inhabitant of biofilms in drinking water distribution systems.M. aviumexpresses on its cell surface serovar-specific glycopeptidolipids (ssGPLs). Studies have implicated the core GPL in biofilm formation byM. aviumand by otherMycobacteriumspecies. In order to test this hypothesis in a directed fashion, three model systems were used to examine biofilm formation by mutants ofM. aviumwith transposon insertions intopstAB(also known asnrpandmps).pstABencodes the nonribosomal peptide synthetase that catalyzes the synthesis of the core GPL. The mutants did not adhere to polyvinyl chloride plates; however, they adhered well to plastic and glass chamber slide surfaces, albeit with different morphologies from the parent strain. In a model that quantified surface adherence under recirculating water, wild-type andpstABmutant cells accumulated on stainless steel surfaces in equal numbers. Unexpectedly,pstABmutant cells were >10-fold less abundant in the recirculating-water phase than parent strain cells. These observations show that GPLs are directly or indirectly required for colonization of some, but by no means all, surfaces. Under some conditions, GPLs may play an entirely different role by facilitating the survival or dispersal of nonadherentM. aviumcells in circulating water. Such a function could contribute to waterborneM. aviuminfection.
In Vitro Deoxynucleotidylation of the Terminal Protein of
Streptomyces
Linear Chromosomes
Authors: Yang Chien-Chin , Chen Yi-Hong , Tsai Hsiu-Hui , Huang Chih-Hung , Huang Tzu-Wen , Chen Carton W.
Published: December 2006

Single-stranded gaps at the 3′ ends of
Streptomyces
linear replicons are patched by DNA synthesis primed by terminal proteins (TP) during replication. We devised an in vitro system that specifically incorporated dCMP, the first nucleotide at the 5′ ends, onto a threonine residue of the TP of
Streptomyces coelicolor
.
Giardia
Cysts and
Cryptosporidium
Oocysts in Membrane-Filtered Municipal Wastewater Used for Irrigation
Authors: Lonigro A. , Pollice A. , Spinelli R. , Berrilli F. , Di Cave D. , D'Orazi C. , Cavallo P. , Brandonisio O.
Published: December 2006

A wastewater tertiary treatment system based on membrane ultrafiltration and fed with secondary-treated municipal wastewater was evaluated for its
Giardia
cyst and
Cryptosporidium
oocyst removal efficiency.
Giardia duodenalis
(assemblages A and B) and
Cryptosporidium parvum
were identified in feed water but were found in filtered water only during occasional failure of the filtration system.