bluuebunny/crossref_metadata_2025 huggingface dataset:
Ultrahigh vacuum sample mount for x-ray photoelectron spectroscopy up to very high temperature (150–1400 K)
Authors: Williamsen Mark S. , Ray Shishir K. , Zou Ying , Dudek John A. , Sen Somaditya , Bissen Mark , Kretsch Laura , Palkar Vaijayanti R. , Onellion Marshall F. , Guptasarma Prasenjit
Published: March 2011

Spectroscopic studies are rarely performed at very high temperature, especially when combined with light from a synchrotron source. Demanding conditions of maintaining ultrahigh vacuum (UHV) during heating, together with the typically brief access to beam time at multiuser synchrotron end stations, may contribute to some of the reasons for the difficulty of such experiments. Consequently, a large number of materials with interesting properties and industrial applications at high temperature remain unexplored. The authors describe here a simple portable sample mount assembly that can be easily utilized at a beamline, with potential utility for a variety of spectroscopic measurements requiring elevated temperatures and an UHV environment. In the specific application described here, the authors use a resistive cartridge heater interfaced with a standard manipulator previously designed for cooling by liquid nitrogen with an UHV chamber and a cylindrical mirror analyzer for x-ray photoemission spectroscopy (XPS) [also known as electron spectroscopy for chemical analysis (ESCA)] at the Synchrotron Radiation Center in Stoughton, WI. The heater cartridge required only modest power to reach target temperatures using an open-loop temperature control. Finally, the authors describe the measurements of XPS (ESCA) and total-electron yield x-ray absorption spectroscopy on nanopowders and on single crystals grown by them. They emphasize the simplicity of the setup, which they believe would be of interest to groups performing measurements at large facilities, where access and time are both limited.
c-MYC Suppresses BIN1 to Release Poly(ADP-Ribose) Polymerase 1: A Mechanism by Which Cancer Cells Acquire Cisplatin Resistance
Authors: Pyndiah Slovénie , Tanida Satoshi , Ahmed Kazi M. , Cassimere Erica K. , Choe Chungyoul , Sakamuro Daitoku
Published: March 2011

c-MYC promotes cisplatin resistance by enabling the increased activity of a DNA repair enzyme.
Focus Issue: Rendering Resistance Futile
Authors: Adler Elizabeth M. , Gough Nancy R.
Published: March 2011

Understanding the pathways that mediate drug resistance is key to developing new cancer therapies.
Activated by Cleavage
Authors: Foley John F.
Published: March 2011

An avian Toll-like receptor is cleaved and activated by virulence-associated fungal and bacterial proteases.
Pancreatic Cancer Immunotherapy
Authors: Mueller Kristen L.
Published: March 2011

CD40 immunotherapy shows efficacy in treating pancreatic cancer in mice and humans by eliciting antitumor immunity.
From Fat Body to Glia to Neuroblasts
Authors: Adler Elizabeth M.
Published: March 2011

A fat-body–derived signal to glia leads to their release of insulin-like peptides to stimulate neuroblast reactivation.
FGF19 and Liver Metabolism
Authors: Ray L. Bryan
Published: March 2011

Fibroblast growth factor 19 regulates liver metabolism through a mechanism distinct from that of insulin.
Quantification of Methicillin-Resistant Staphylococcus aureus Strains in Marine and Freshwater Samples by the Most-Probable-Number Method
Authors: Levin-Edens Emily , Meschke John Scott , Roberts Marilyn C.
Published: March 2011

Recreational beach environments have been recently identified as a potential reservoir for methicillin-resistant
Staphylococcus aureus
(MRSA); however, accurate quantification methods are needed for the development of risk assessments. This novel most-probable-number approach for MRSA quantification offers improved sensitivity and specificity by combining broth enrichment with MRSA-specific chromogenic agar.
Validation of Alternative Methods for the Analysis of Drinking Water and Their Application to Escherichia coli
Authors: Boubetra Abdelkader , Le Nestour François , Allaert Corrie , Feinberg Max
Published: March 2011

In Europe, the Drinking Water Directive of the European Commission indicates which methods (most of which are CEN/ISO-standardized methods) should be used for the analysis of microbiological parameters (European Commission, Environment, Council Directive 98/83/EC of 3 November 1998). According to the Directive, alternative methods “may be used, providing it can be demonstrated that the results obtained are at least as reliable as those produced by the methods specified.” The prerequisite for the routine use of any alternative method is to provide evidence that this method performs equivalently to the corresponding reference method. In this respect, the ISO 16140 standard (ISO,
ISO 16140. Microbiology of Food and Animal Feeding Stuffs—Protocol for the Validation of Alternative Methods
, 2003) represents a key issue in generating such a procedure based on an interlaboratory study. A new statistical tool, called the accuracy profile, has been developed to better interpret the data. The study presented here is based upon the enumeration of
Escherichia coli
bacteria in water. The reference method may require up to 72 h to provide a confirmed result. The aim of this publication is to present data for an alternative method by which results can be obtained in 18 h (Colilert-18/Quanti-Tray) based upon defined substrate technology (DST). The accuracy profile is a statistical and graphical decision-making tool and consists of simultaneously combining, in a single graphic, β expectation tolerance intervals (β-ETIs) and acceptability limits (λ). The study presents the validation criteria calculated at the three levels of contamination used in the trial for a β equal to 80% and a λ equal to ±0.3 and combines the accuracy profiles of
Escherichia coli
for a λ of ±0.3 log
10
unit/100 ml, a λ of ±0.4 log
10
unit/100 ml, and a β of 80% or 90%. Several interesting conclusions can be drawn from these data. The accuracy profile method has been applied to the validation of the Colilert-18/Quanti-Tray method against reference method ISO 9308-1 (ISO,
ISO 9308
-
1. Water Quality—Detection and Enumeration of Escherichia coli and Coliform Bacteria. Part 1. Membrane Filtration Method
, 2000), using a β of 80% and a λ of 0.4; the alternative method can be validated between 1.00 and 2.05 log
10
units/100 ml, equivalent to 10 to 112 CFU/100 ml.
Kinetics of Germination of Wet-Heat-Treated Individual Spores of Bacillus Species, Monitored by Raman Spectroscopy and Differential Interference Contrast Microscopy
Authors: Wang Guiwen , Zhang Pengfei , Setlow Peter , Li Yong-qing
Published: March 2011

Raman spectroscopy and differential interference contrast (DIC) microscopy were used to monitor the kinetics of nutrient and nonnutrient germination of multiple individual untreated and wet-heat-treated spores of
Bacillus cereus
and
Bacillus megaterium
, as well as of several isogenic
Bacillus subtilis
strains. Major conclusions from this work were as follows. (i) More than 90% of these spores were nonculturable but retained their 1:1 chelate of Ca
2+
and dipicolinic acid (CaDPA) when incubated in water at 80 to 95°C for 5 to 30 min. (ii) Wet-heat treatment significantly increased the time,
T
lag
, at which spores began release of the great majority of their CaDPA during the germination of
B. subtilis
spores with different nutrient germinants and also increased the variability of
T
lag
values. (iii) The time period, Δ
T
release
, between
T
lag
and the time,
T
release
, at which a spore germinating with nutrients completed the release of the great majority of its CaDPA, was also increased in wet-heat-treated spores. (iv) Wet-heat-treated spores germinating with nutrients had higher values of
I
release
, the intensity of a spore's DIC image at
T
release
, than did untreated spores and had much longer time periods, Δ
T
lys
, for the reduction in
I
release
intensities to the basal value due to hydrolysis of the spore's peptidoglycan cortex, probably due at least in part to damage to the cortex-lytic enzyme CwlJ. (v) Increases in
T
lag
and Δ
T
release
were also observed when wet-heat-treated
B. subtilis
spores were germinated with the nonnutrient dodecylamine, while the change in
I
release
was less significant. (vi) The effects of wet-heat treatment on nutrient germination of
B. cereus
and
B. megaterium
spores were generally similar to those on
B. subtilis
spores. These results indicate that (i) some proteins important in spore germination are damaged by wet-heat treatment, (ii) the cortex-lytic enzyme CwlJ is one germination protein damaged by wet heat, and (iii) the CaDPA release process itself seems likely to be the target of wet-heat damage which has the greatest effect on spore germination.
Inactivation of a Human Norovirus Surrogate, Human Norovirus Virus-Like Particles, and Vesicular Stomatitis Virus by Gamma Irradiation
Authors: Feng Kurtis , Divers Erin , Ma Yuanmei , Li Jianrong
Published: March 2011

Gamma irradiation is a nonthermal processing technology that has been used for the preservation of a variety of food products. This technology has been shown to effectively inactivate bacterial pathogens. Currently, the FDA has approved doses of up to 4.0 kGy to control food-borne pathogens in fresh iceberg lettuce and spinach. However, whether this dose range effectively inactivates food-borne viruses is less understood. We have performed a systematic study on the inactivation of a human norovirus surrogate (murine norovirus 1 [MNV-1]), human norovirus virus-like particles (VLPs), and vesicular stomatitis virus (VSV) by gamma irradiation. We demonstrated that MNV-1 and human norovirus VLPs were resistant to gamma irradiation. For MNV-1, only a 1.7- to 2.4-log virus reduction in fresh produce at the dose of 5.6 kGy was observed. However, VSV was more susceptible to gamma irradiation, and a 3.3-log virus reduction at a dose of 5.6 kGy in Dulbecco's modified Eagle medium (DMEM) was achieved. We further demonstrated that gamma irradiation disrupted virion structure and degraded viral proteins and genomic RNA, which resulted in virus inactivation. Using human norovirus VLPs as a model, we provide the first evidence that the capsid of human norovirus has stability similar to that of MNV-1 after exposure to gamma irradiation. Overall, our results suggest that viruses are much more resistant to irradiation than bacterial pathogens. Although gamma irradiation used to eliminate the virus contaminants in fresh produce by the FDA-approved irradiation dose limits seems impractical, this technology may be practical to inactivate viruses for other purposes, such as sterilization of medical equipment.
Production of
N
-Acetyl-
d
-Neuraminic Acid by Use of an Efficient Spore Surface Display System
Authors: Xu Xiaoman , Gao Chao , Zhang Xifeng , Che Bin , Ma Cuiqing , Qiu Jianhua , Tao Fei , Xu Ping
Published: March 2011

Production of
N
-acetyl-
d
-neuraminic acid (Neu5Ac) via biocatalysis is traditionally conducted using isolated enzymes or whole cells. The use of isolated enzymes is restricted by the time-consuming purification process, whereas the application of whole cells is limited by the permeability barrier presented by the microbial cell membrane. In this study, a novel type of biocatalyst, Neu5Ac aldolase presented on the surface of
Bacillus subtilis
spores, was used for the production of Neu5Ac. Under optimal conditions, Neu5Ac at a high concentration (54.7 g liter
−1
) and a high yield (90.2%) was obtained under a 5-fold excess of pyruvate over
N
-acetyl-
d
-mannosamine. The novel biocatalyst system, which is able to express and immobilize the target enzyme simultaneously on the surface of
B. subtilis
spores, represents a suitable alternative for value-added chemical production.
comK
Prophage Junction Fragments as Markers for Listeria monocytogenes Genotypes Unique to Individual Meat and Poultry Processing Plants and a Model for Rapid Niche-Specific Adaptation, Biofilm Formation, and Persistence
Authors: Verghese Bindhu , Lok Mei , Wen Jia , Alessandria Valentina , Chen Yi , Kathariou Sophia , Knabel Stephen
Published: March 2011

Different strains of
Listeria monocytogenes
are well known to persist in individual food processing plants and to contaminate foods for many years; however, the specific genotypic and phenotypic mechanisms responsible for persistence of these unique strains remain largely unknown. Based on sequences in
comK
prophage junction fragments, different strains of epidemic clones (ECs), which included ECII, ECIII, and ECV, were identified and shown to be specific to individual meat and poultry processing plants. The
comK
prophage-containing strains showed significantly higher cell densities after incubation at 30°C for 48 h on meat and poultry food-conditioning films than did strains lacking the
comK
prophage (
P
< 0.05). Overall, the type of strain, the type of conditioning film, and the interaction between the two were all highly significant (
P
< 0.001). Recombination analysis indicated that the
comK
prophage junction fragments in these strains had evolved due to extensive recombination. Based on the results of the present study, we propose a novel model in which the concept of defective
comK
prophage was replaced with the rapid adaptation island (RAI). Genes within the RAI were recharacterized as “adaptons,” as these genes may allow
L. monocytogenes
to rapidly adapt to different food processing facilities and foods. If confirmed, the model presented would help explain
Listeria
's rapid niche adaptation, biofilm formation, persistence, and subsequent transmission to foods. Also,
comK
prophage junction fragment sequences may permit accurate tracking of persistent strains back to and within individual food processing operations and thus allow the design of more effective intervention strategies to reduce contamination and enhance food safety.
High-Yield Resveratrol Production in Engineered Escherichia coli
Authors: Lim Chin Giaw , Fowler Zachary L. , Hueller Thomas , Schaffer Steffen , Koffas Mattheos A. G.
Published: March 2011

Plant polyphenols have been the subject of several recent scientific investigations since many of the molecules in this class have been found to be highly active in the human body, with a plethora of health-promoting activities against a variety of diseases, including heart disease, diabetes, and cancer, and with even the potential to slow aging. Further development of these potent natural therapeutics hinges on the formation of robust industrial production platforms designed using specifically selected as well as engineered protein sources along with the construction of optimal expression platforms. In this work, we first report the investigation of various stilbene synthases from an array of plant species considering structure-activity relationships, their expression efficiency in microorganisms, and their ability to synthesize resveratrol. Second, we looked into the construct environment of recombinantly expressed stilbene synthases, including different promoters, construct designs, and host strains, to create an
Escherichia coli
strain capable of producing superior resveratrol titers sufficient for commercial usage. Further improvement of metabolic capabilities of the recombinant strain aimed at improving the intracellular malonyl-coenzyme A pool, a resveratrol precursor, resulted in a final improved titer of 2.3 g/liter resveratrol.
Sensitive Detection of Shiga Toxin 2 and Some of Its Variants in Environmental Samples by a Novel Immuno-PCR Assay
Authors: He Xiaohua , Qi Wenyuan , Quiñones Beatriz , McMahon Stephanie , Cooley Michael , Mandrell Robert E.
Published: March 2011

Shiga toxin-producing
Escherichia coli
(STEC) in the environment has been reported frequently. However, robust detection of STEC in environmental samples remains difficult because the numbers of bacteria in samples are often below the detection threshold of the method. We developed a novel and sensitive immuno-PCR (IPCR) assay for the detection of Shiga toxin 2 (Stx2) and Stx2 variants. The assay involves immunocapture of Stx2 at the B subunit and real-time PCR amplification of a DNA marker linked to a detection antibody recognizing the Stx2 A subunit. The qualitative detection limit of the assay is 0.1 pg/ml in phosphate-buffered saline (PBS), with a quantification range of 10 to 100,000 pg/ml. The IPCR method was 10,000-fold more sensitive than an analogue conventional enzyme-linked immunosorbent assay (ELISA) in PBS. Although the sensitivity of the IPCR for detection of Stx2 was affected by environmental sample matrices of feces, feral swine colons, soil, and water from watersheds, application of the IPCR assay to 23 enriched cultures of fecal, feral swine colon, soil, and watershed samples collected from the environment revealed that the IPCR detected Stx2 in all 15 samples that were shown to be STEC positive by real-time PCR and culture methods, demonstrating a 100% sensitivity and specificity. The modification of the sandwich IPCR we have described in this study will be a sensitive and specific screening method for evaluating the occurrence of STEC in the environment.
Influence of Antimicrobial Feed Additives on Broiler Commensal Posthatch Gut Microbiota Development and Performance
Authors: Torok Valeria A. , Allison Gwen E. , Percy Nigel J. , Ophel-Keller Kathy , Hughes Robert J.
Published: March 2011

The effects of avilamycin, zinc bacitracin, and flavophospholipol on broiler gut microbial community colonization and bird performance in the first 17 days posthatch were investigated. Significant differences in gut microbiota associated with gut section, dietary treatment, and age were identified by terminal restriction fragment length polymorphism (T-RFLP), although no performance-related differences between dietary treatments were detected. Similar age-related shifts in the gut microbiota were identified regardless of diet but varied between the ilea and ceca. Interbird variabilities in ileal bacterial communities were reduced (3 to 7 days posthatch) in chicks fed with feed containing antimicrobial agents. Avilamycin and flavophospholipol had the most consistent effect on gut microbial communities. Operational taxonomic units (OTU) linked to changes in gut microbiota in birds on antimicrobial-supplemented diets were characterized and identified. Some OTUs could be identified to the species level; however, the majority could be only tentatively classified to the genus, family, order, or domain level. OTUs 140 to 146 (
Lachnospiraceae
), OTU 186/188 (
Lactobacillus johnsonii
), OTU 220 (
Lachnospiraceae
), OTUs 284 to 288 (unclassified bacterial spp. or
Ruminococcaceae
), OTU 296/298 (unclassified bacterium or
Clostridiales
), and OTU 480/482 (
Oxalobacteraceae
) were less prevalent in the guts of chicks fed antimicrobial-supplemented diets. OTU 178/180 (
Lactobacillus crispatus
), OTU 152 (
Lactobacillus reuteri
or unclassified
Clostridiales
), OTU 198/200 (
Subdoligranulum
spp.), and OTU 490/492 (unclassified bacterium or
Enterobacteriaceae
) were less prevalent in the gut of chicks raised on the antimicrobial-free diet. The identification of key bacterial species influenced by antimicrobial-supplemented feed immediately posthatch may assist in the formulation of diets that facilitate beneficial gut microbial colonization and, hence, the development of alternatives to current antimicrobial agents in feed for sustainable poultry production.
Increased In Vitro Adherence and On-Farm Persistence of Predominant and Persistent Listeria monocytogenes Strains in the Milking System
Authors: Latorre Alejandra A. , Van Kessel Jo Ann S. , Karns Jeffrey S. , Zurakowski Michael J. , Pradhan Abani K. , Boor Kathryn J. , Adolph Evin , Sukhnanand Sharinne , Schukken Ynte H.
Published: March 2011

Dairy farms are a reservoir for
Listeria monocytogenes
, and the reduction of this pathogen at the farm level is important for reducing human exposure. The objectives of this research were to study the diversity of
L. monocytogenes
strains on a single dairy farm, assess strain dynamics within the farm, identify potential sources of
L. monocytogenes
in bulk tank milk and milk filters, and assess the adherence abilities of representative strains. A total of 248
L. monocytogenes
isolates were analyzed by pulsed-field gel electrophoresis (PFGE). Combined AscI and ApaI restriction analysis yielded 40 PFGE types (strains). The most predominant strains were T (28.6%), D (22.6%), and F (14.9%). A high level of heterogeneity of strains among isolates from fecal (Simpson's index of diversity [SID] = 0.96) and environmental (SID = 0.96) samples was observed. A higher homogeneity of strains was observed among isolates from milk filters (SID = 0.71) and bulk tank milk (SID = 0.65). Six of 17
L. monocytogenes
isolates (35.3%) were classified in an
in vitro
assay as having a “low adherence ability,” 9 (52.9%) were classified as having a “medium adherence ability,” and 2 (11.8%) were classified as having a “high adherence ability.” The
L. monocytogenes
strains that were predominant and persistent showed significantly better adherence than did strains that were only sporadic, predominant, or persistent (
P
= 0.0006). Our results suggest that the milking system was exposed to several
L. monocytogenes
strains from different sources. Only 3 strains, however, were successful in persisting within the milking system, suggesting that some strains are more suitable to that particular ecological environment than others.
Novel Bacillus thuringiensis δ-Endotoxin Active against Locusta migratoria manilensis
Authors: Wu Yan , Lei Cheng-Feng , Yi Dan , Liu Peng-Ming , Gao Mei-Ying
Published: March 2011

A novel δ-endotoxin gene was cloned from a
Bacillus thuringiensis
strain with activity against
Locusta migratoria manilensis
by PCR-based genome walking. The sequence of the
cry
gene was 3,432 bp long, and it encoded a Cry protein of 1,144 amino acid residues with a molecular mass of 129,196.5 kDa, which exhibited 62% homology with Cry7Ba1 in the amino acid sequence. The δ-endotoxin with five conserved sequence blocks in the amino-terminal region was designated Cry7Ca1 (GenBank accession no.
EF486523
). Protein structure analysis suggested that the activated toxin of Cry7Ca1 has three domains: 227 residues forming 7 α-helices (domain I); 213 residues forming three antiparallel β-sheets (domain II); and 134 residues forming a β-sandwich (domain III). The three domains, respectively, exhibited 47, 44, and 34% sequence identity with corresponding domains of known Cry toxins. SDS-PAGE and Western blot analysis showed that Cry7Ca1, encoded by the full-length open reading frame of the
cry
gene, the activated toxin 1, which included three domains but without the N-terminal 54 amino acid residues and the C terminus, and the activated toxin 2, which included three domains and N-terminal 54 amino acid residues but without the C terminus, could be expressed in
Escherichia coli
. Bioassay results indicated that the expressed proteins of Cry7Ca1 and the activated toxins (toxins 1 and 2) showed significant activity against 2nd instar locusts, and after 7 days of infection, the estimated 50% lethal concentrations (LC
50
s) were 8.98 μg/ml for the expressed Cry7Ca1, 0.87 μg/ml for the activated toxin 1, and 4.43 μg/ml for the activated toxin 2. The δ-endotoxin also induced histopathological changes in midgut epithelial cells of adult
L. migratoria manilensis
.
Transcriptomic Responses of the Softwood-Degrading White-Rot Fungus Phanerochaete carnosa during Growth on Coniferous and Deciduous Wood
Authors: MacDonald Jacqueline , Doering Matt , Canam Thomas , Gong Yunchen , Guttman David S. , Campbell Malcolm M. , Master Emma R.
Published: March 2011

To identify enzymes that could be developed to reduce the recalcitrance of softwood resources, the transcriptomes of the softwood-degrading white-rot fungus
Phanerochaete carnosa
were evaluated after growth on lodgepole pine, white spruce, balsam fir, and sugar maple and compared to the transcriptome of
P. carnosa
after growth on liquid nutrient medium. One hundred fifty-two million paired-end reads were obtained, and 63% of these reads were mapped to 10,257 gene models from
P. carnosa.
Five-hundred thirty-three of these genes had transcripts that were at least four times more abundant during growth on at least one wood medium than on nutrient medium. The 30 transcripts that were on average over 100 times more abundant during growth on wood than on nutrient medium included 6 manganese peroxidases, 5 cellulases, 2 hemicellulases, a lignin peroxidase, glyoxal oxidase, and a P450 monooxygenase. Notably, among the genes encoding putative cellulases, one encoding a glycosyl hydrolase family 61 protein had the highest relative transcript abundance during growth on wood. Overall, transcripts predicted to encode lignin-degrading activities were more abundant than those predicted to encode carbohydrate-active enzymes. Transcripts predicted to encode three MnPs represented the most highly abundant transcripts in wood-grown cultivations compared to nutrient medium cultivations. Gene set enrichment analyses did not distinguish transcriptomes resulting from softwood and hardwood cultivations, suggesting that similar sets of enzyme activities are elicited by
P. carnosa
grown on different wood substrates, albeit to different expression levels.
Root-Associated Ectomycorrhizal Fungi Shared by Various Boreal Forest Seedlings Naturally Regenerating after a Fire in Interior Alaska and Correlation of Different Fungi with Host Growth Responses
Authors: Bent Elizabeth , Kiekel Preston , Brenton Rebecca , Taylor D. Lee
Published: March 2011

The role of common mycorrhizal networks (CMNs) in postfire boreal forest successional trajectories is unknown. We investigated this issue by sampling a 50-m by 40-m area of naturally regenerating black spruce (
Picea mariana
), trembling aspen (
Populus tremuloides
), and paper birch (
Betula papyrifera
) seedlings at various distances from alder (
Alnus viridis
subsp.
crispa
), a nitrogen-fixing shrub, 5 years after wildfire in an Alaskan interior boreal forest. Shoot biomasses and stem diameters of 4-year-old seedlings were recorded, and the fungal community associated with ectomycorrhizal (ECM) root tips from each seedling was profiled using molecular techniques. We found distinct assemblages of fungi associated with alder compared with those associated with the other tree species, making the formation of CMNs between them unlikely. However, among the spruce, aspen, and birch seedlings, there were many shared fungi (including members of the
Pezoloma ericae
[
Hymenoscyphus ericae
] species aggregate,
Thelephora terrestris
, and
Russula
spp.), raising the possibility that these regenerating seedlings may form interspecies CMNs. Distance between samples did not influence how similar ECM root tip-associated fungal communities were, and of the fungal groups identified, only one of them was more likely to be shared between seedlings that were closer together, suggesting that the majority of fungi surveyed did not have a clumped distribution across the small scale of this study. The presence of some fungal ribotypes was associated with larger or smaller seedlings, suggesting that these fungi may play a role in the promotion or inhibition of seedling growth. The fungal ribotypes associated with larger seedlings were different between spruce, aspen, and birch, suggesting differential impacts of some host-fungus combinations. One may speculate that wildfire-induced shifts in a given soil fungal community could result in variation in the growth response of different plant species after fire and a shift in regenerating vegetation.
Formyltetrahydrofolate Synthetase Gene Diversity in the Guts of Higher Termites with Different Diets and Lifestyles
Authors: Ottesen Elizabeth A. , Leadbetter Jared R.
Published: March 2011

In this study, we examine gene diversity for formyl-tetrahydrofolate synthetase (FTHFS), a key enzyme in homoacetogenesis, recovered from the gut microbiota of six species of higher termites. The “higher” termites (family
Termitidae
), which represent the majority of extant termite species and genera, engage in a broader diversity of feeding and nesting styles than the “lower” termites. Previous studies of termite gut homoacetogenesis have focused on wood-feeding lower termites, from which the preponderance of FTHFS sequences recovered were related to those from acetogenic treponemes. While sequences belonging to this group were present in the guts of all six higher termites examined, treponeme-like FTHFS sequences represented the majority of recovered sequences in only two species (a wood-feeding
Nasutitermes
sp. and a palm-feeding
Microcerotermes
sp.). The remaining four termite species analyzed (a
Gnathamitermes
sp. and two
Amitermes
spp. that were recovered from subterranean nests with indeterminate feeding strategies and a litter-feeding
Rhynchotermes
sp.) yielded novel FTHFS clades not observed in lower termites. These termites yielded two distinct clusters of probable purinolytic
Firmicutes
and a large group of potential homoacetogens related to sequences previously recovered from the guts of omnivorous cockroaches. These findings suggest that the gut environments of different higher termite species may select for different groups of homoacetogens, with some species hosting treponeme-dominated homoacetogen populations similar to those of wood-feeding, lower termites while others host
Firmicutes
-dominated communities more similar to those of omnivorous cockroaches.
Antifungal Activity of Wickerhamomyces anomalus and Lactobacillus plantarum during Sourdough Fermentation: Identification of Novel Compounds and Long-Term Effect during Storage of Wheat Bread
Authors: Coda Rossana , Cassone Angela , Rizzello Carlo G. , Nionelli Luana , Cardinali Gianluigi , Gobbetti Marco
Published: March 2011

This study aimed at investigating the antifungal activity of
Wickerhamomyces anomalus
and sourdough lactic acid bacteria to extend the shelf life of wheat flour bread. The antifungal activity was assayed by agar diffusion, growth rate inhibition, and conidial germination assays, using
Penicillium roqueforti
DPPMAF1 as the indicator fungus. Sourdough fermented by
Lactobacillus plantarum
1A7 (S1A7) and dough fermented by
W. anomalus
LCF1695 (D1695) were selected and characterized. The water/salt-soluble extract of S1A7 was partially purified, and several novel antifungal peptides, encrypted into sequences of
Oryza sativa
proteins, were identified. The water/salt-soluble extract of D1695 contained ethanol and, especially, ethyl acetate as inhibitory compounds. As shown by growth inhibition assays, both water/salt-soluble extracts had a large inhibitory spectrum, with some differences, toward the most common fungi isolated from bakeries. Bread making at a pilot plant was carried out with S1A7, D1695, or a sourdough started with a combination of both strains (S1A7-1695). Slices of the bread manufactured with S1A7-1695 did not show contamination by fungi until 28 days of storage in polyethylene bags at room temperature, a level of protection comparable to that afforded by 0.3% (wt/wt) calcium propionate. The effect of sourdough fermentation with
W. anomalus
LCF1695 was also assessed based on rheology and sensory properties.
Quantification and Evaluation of Infectivity of Shiga Toxin-Encoding Bacteriophages in Beef and Salad
Authors: Imamovic Lejla , Muniesa Maite
Published: March 2011

Stx bacteriophages in 68 samples of beef and salad were quantified by real-time quantitative PCR (qPCR). Stx phages from the samples were propagated in
Escherichia coli
C600,
E. coli
O157:H7, and
Shigella
strains and further quantified. Fifty percent of the samples carried infectious Stx phages that were isolated from plaques generated by lysis.
Bacteriophage-Mediated Dispersal of Campylobacter jejuni Biofilms
Authors: Siringan Patcharin , Connerton Phillippa L. , Payne Robert J. H. , Connerton Ian F.
Published: March 2011

Bacteria in their natural environments frequently exist as mixed surface-associated communities, protected by extracellular material, termed biofilms. Biofilms formed by the human pathogenCampylobacter jejunimay arise in the gastrointestinal tract of animals but also in water pipes and other industrial situations, leading to their possible transmission into the human food chain either directly or via farm animals. Bacteriophages are natural predators of bacteria that usually kill their prey by cell lysis and have potential application for the biocontrol and dispersal of target bacteria in biofilms. The effects of virulentCampylobacterspecific-bacteriophages CP8 and CP30 onC. jejunibiofilms formed on glass by strains NCTC 11168 and PT14 at 37°C under microaerobic conditions were investigated. Independent bacteriophage treatments (n≥ 3) led to 1 to 3 log10CFU/cm2reductions in the viable count 24 h postinfection compared with control levels. In contrast, bacteriophages applied under these conditions effected a reduction of less than 1 log10CFU/ml in planktonic cells. Resistance to bacteriophage in bacteria surviving bacteriophage treatment ofC. jejuniNCTC 11168 biofilms was 84% and 90% for CP8 and CP30, respectively, whereas bacteriophage resistance was not found in similarly recoveredC. jejuniPT14 cells. Dispersal of the biofilm matrix by bacteriophage was demonstrated by crystal violet staining and transmission electron microscopy. Bacteriophage may play an important role in the control of attachment and biofilm formation byCampylobacterin situations where biofilms occur in nature, and they have the potential for application in industrial situations leading to improvements in food safety.
New Method Using a Positively Charged Microporous Filter and Ultrafiltration for Concentration of Viruses from Tap Water
Authors: Ikner Luisa A. , Soto-Beltran Marcela , Bright Kelly R.
Published: March 2011

The methods used to concentrate enteric viruses from water have remained largely unchanged for nearly 30 years, with the most common technique being the use of 1MDS Virozorb filters followed by organic flocculation for secondary concentration. Recently, a few studies have investigated alternatives; however, many of these methods are impractical for use in the field or share some of the limitations of this traditional method. In the present study, the NanoCeram virus sampler, an electropositive pleated microporous filter composed of microglass filaments coated with nanoalumina fibers, was evaluated. Test viruses were first concentrated by passage of 20 liters of seeded water through the filter (average filter retention efficiency was ≥99.8%), and then the viruses were recovered using various salt-based or proteinaceous eluting solutions. A 1.0% sodium polyphosphate solution with 0.05 M glycine was determined to be the most effective. The recovered viruses were then further concentrated using Centricon Plus-70 centrifugal ultrafilters to a final volume of 3.3 (±0.3 [standard deviation]) ml; this volume compares quite favorably to that of previously described methods, such as organic flocculation (∼15 to 40 ml). The overall virus recovery efficiencies were 66% for poliovirus 1, 83% for echovirus 1, 77% for coxsackievirus B5, 14% for adenovirus 2, and 56% for MS2 coliphage. In addition, this method appears to be compatible with both cell culture and PCR assays. This new approach for the recovery of viruses from water is therefore a viable alternative to currently used methods when small volumes of final concentrate are an advantage.
Transient Fecal Shedding and Limited Animal-to-Animal Transmission of Clostridium difficile by Naturally Infected Finishing Feedlot Cattle
Authors: Rodriguez-Palacios Alexander , Pickworth Carrie , Loerch Steve , LeJeune Jeffrey T.
Published: March 2011

To longitudinally assess fecal shedding and animal-to-animal transmission of
Clostridium difficile
among finishing feedlot cattle as a risk for beef carcass contamination, we tested 186 ± 12 steers (mean ± standard deviation; 1,369 samples) in an experimental feedlot facility during the finishing period and at harvest.
Clostridium difficile
was isolated from 12.9% of steers on arrival (24/186; 0 to 33% among five suppliers). Shedding decreased to undetectable levels a week later (0%;
P
< 0.001), and remained low (<3.6%) until immediately prior to shipment for harvest (1.2%). Antimicrobial use did not increase fecal shedding, despite treatment of 53% of animals for signs of respiratory disease. Animals shedding
C. difficile
on arrival, however, had 4.6 times higher odds of receiving antimicrobials for respiratory signs than nonshedders (95% confidence interval for the odds ratio, 1.4 to 14.8;
P
= 0.01). Neither the toxin genes nor toxin A or B was detected in most (39/42) isolates based on two complementary multiplex PCRs and enzyme-linked immunosorbent assay testing, respectively. Two linezolid- and clindamycin-resistant PCR ribotype 078 (
tcdA
+
/
tcdB
+
/
cdtB
+
/39-bp-type deletion in
tcdC
) isolates were identified from two steers (at arrival and week 20), but these ribotypes did not become endemic. The other toxigenic isolate (
tcdA
+
/
tcdB
+
/
cdtB
+
/classic
tcdC
; PCR ribotype 078-like) was identified in the cecum of one steer at harvest. Spatio-temporal analysis indicated transient shedding with no evidence of animal-to-animal transmission. The association between
C. difficile
shedding upon arrival and the subsequent need for antimicrobials for respiratory disease might indicate common predisposing factors. The isolation of toxigenic
C. difficile
from bovine intestines at harvest highlights the potential for food contamination in meat processing plants.
Core and Intact Polar Glycerol Dibiphytanyl Glycerol Tetraether Lipids of Ammonia-Oxidizing Archaea Enriched from Marine and Estuarine Sediments
Authors: Pitcher Angela , Hopmans Ellen C. , Mosier Annika C. , Park Soo-Je , Rhee Sung-Keun , Francis Christopher A. , Schouten Stefan , Sinninghe Damsté Jaap S.
Published: March 2011

Glycerol dibiphytanyl glycerol tetraether (GDGT)-based intact membrane lipids are increasingly being used as complements to conventional molecular methods in ecological studies of ammonia-oxidizing archaea (AOA) in the marine environment. However, the few studies that have been done on the detailed lipid structures synthesized by AOA in (enrichment) culture are based on species enriched from nonmarine environments, i.e., a hot spring, an aquarium filter, and a sponge. Here we have analyzed core and intact polar lipid (IPL)-GDGTs synthesized by three newly available AOA enriched directly from marine sediments taken from the San Francisco Bay estuary (“
Candidatus
Nitrosoarchaeum limnia”), and coastal marine sediments from Svalbard, Norway, and South Korea. Like previously screened AOA, the sedimentary AOA all synthesize crenarchaeol (a GDGT containing a cyclohexane moiety and four cyclopentane moieties) as a major core GDGT, thereby supporting the hypothesis that crenarchaeol is a biomarker lipid for AOA. The IPL headgroups synthesized by sedimentary AOA comprised mainly monohexose, dihexose, phosphohexose, and hexose-phosphohexose moieties. The hexose-phosphohexose headgroup bound to crenarchaeol was common to all enrichments and, in fact, the only IPL common to every AOA enrichment analyzed to date. This apparent specificity, in combination with its inferred lability, suggests that it may be the most suitable biomarker lipid to trace living AOA. GDGTs bound to headgroups with a mass of 180 Da of unknown structure appear to be specific to the marine group I.1a AOA: they were synthesized by all three sedimentary AOA and “
Candidatus
Nitrosopumilus maritimus”; however, they were absent in the group I.1b AOA “
Candidatus
Nitrososphaera gargensis.”
Improvement of Fitness and Efficacy of a Fire Blight Biocontrol Agent via Nutritional Enhancement Combined with Osmoadaptation
Authors: Cabrefiga J. , Francés J. , Montesinos E. , Bonaterra A.
Published: March 2011

The efficacy of
Pseudomonas fluorescens
EPS62e in the biocontrol of
Erwinia amylovora
was improved by a procedure of physiological adaptation to increase colonization and survival in the phytosphere of rosaceous plants. The procedure consisted of osmoadaptation (OA) and nutritional enhancement (NE). OA was based on saline stress and osmolyte amendment of the growth medium during inoculum preparation. NE consisted of addition of glycine and Tween 80 to the formulation. NE and OA increased the growth rate and carrying capacity of EPS62e under high-relative-humidity (RH) conditions and improved survival at low RH on flowers under controlled environmental conditions. NE did not promote growth or affect infection capacity of
E. amylovora
. The effect of both methods was tested in the field by following the population of EPS62e using quantitative PCR (Q-PCR) (total population) and CFU counting (culturable population) methods. Following field application, EPS62e colonized blossoms, but it was stressed, as indicated by a sharp decrease in culturable compared to total population levels. However, once established in flowers and at the end of bloom, almost all the total population was culturable. The physiological adaptation treatments increased population levels of EPS62e over those of nonadapted cells during the late stage of the flowering period. Control of fire blight infections in flowers and immature fruits was tested by field application of EPS62e and subsequent inoculation with
E. amylovora
under controlled-environment conditions. The efficacy of fire blight control increased significantly with the combination of nutritional enhancement and osmoadaptation, in comparison with the absence of physiological adaptation.
Integrated Regulation of Acetoin Fermentation by Quorum Sensing and pH in Serratia plymuthica RVH1
Authors: Moons Pieter , Van Houdt Rob , Vivijs Bram , Michiels Chris M. , Aertsen Abram
Published: March 2011

During fermentation of sugars, a number of bacterial species are able to switch from mixed acid production to acetoin and 2,3-butanediol production in order to avoid lethal acidification of their environment, although the regulation of this switch is only poorly understood. In this study, we report the identification of the
budAB
structural operon, involved in acetoin production in
Serratia plymuthica
RVH1, and its activation by a LysR-type regulator encoded by
budR
, immediately upstream of this operon. In addition, the regulation of
budR
transcription was elucidated and found to be subject to negative control by BudR itself and to positive control by external stimuli such as
N
-(3-oxohexanoyl)-
l
-homoserine lactone (OHHL) quorum sensing signaling molecules and acetate. Interestingly, however, we observed that induction of
budR
transcription by OHHL or acetate did not require BudR, indicating the involvement of additional regulatory factors in relaying these environmental signals to the
budR
promoter.
Binding Sites for Bacillus thuringiensis Cry2Ae Toxin on Heliothine Brush Border Membrane Vesicles Are Not Shared with Cry1A, Cry1F, or Vip3A Toxin
Authors: Gouffon C. , Van Vliet A. , Van Rie J. , Jansens S. , Jurat-Fuentes J. L.
Published: March 2011

The use of combinations of
Bacillus thuringiensis
(Bt) toxins with diverse modes of action for insect pest control has been proposed as the most efficient strategy to increase target range and delay the onset of insect resistance. Considering that most cases of cross-resistance to Bt toxins in laboratory-selected insect colonies are due to alteration of common toxin binding sites, independent modes of action can be defined as toxins sharing limited or no binding sites in brush border membrane vesicles (BBMV) prepared from the target insect larvae. In this paper, we report on the specific binding of Cry2Ae toxin to binding sites on BBMV from larvae of the three most commercially relevant heliothine species,
Heliothis virescens
,
Helicoverpa zea
, and
Helicoverpa armigera
. Using chromatographic purification under reducing conditions before labeling, we detected specific binding of radiolabeled Cry2Ae, which allowed us to perform competition assays using Cry1Ab, Cry1Ac, Cry1Fa, Vip3A, Cry2Ae, and Cry2Ab toxins as competitors. In these assays, Cry2Ae binding sites were shared with Cry2Ab but not with the tested Cry1 or Vip3A toxins. Our data support the use of Cry2Ae toxin in combination with Cry1 or Vip3A toxins in strategies to increase target range and delay the onset of heliothine resistance.
Genetic Typing, Based on the 56-Kilodalton Type-Specific Antigen Gene, of Orientia tsutsugamushi Strains Isolated from Chiggers Collected from Wild-Caught Rodents in Taiwan
Authors: Lin Pey-Ru , Tsai Hui-Ping , Tsui Pei-Yi , Weng Ming-Hui , Kuo Ming-Der , Lin Hung-Chi , Chen Kuo-Ching , Ji Dar-Der , Chu Der-Ming , Liu Wen-Tssann
Published: March 2011

Orientia tsutsugamushiis the etiological agent of scrub typhus, a mite-borne, febrile illness that occurs in the Asia-Pacific region. We conducted strain characterization ofO. tsutsugamushiisolates from chiggers obtained from rodents based the nucleotide sequence of the 56-kDa outer membrane protein gene. With the use of PCR, a total of 68 DNA sequences of 56-kDa antigen genes were amplified. Phylogenetic analysis revealed that there were at least six definable clusters among the 68 isolates: 37% Karp-related strains (25/68), 27% TA763 strains (18/68), 12% JG-related strains (8/68), 19% Kato-related strains (13/68), 4% divergent strains (3/68), and 1% representing a Gilliam prototype strain (1/68). Overall, theO. tsutsugamushigenotypes exhibited a high degree of diversity, similar to that seen in strains from the rest of the areas where scrub typhus is endemic. Moreover, the 56-kDa protein sequence similarity betweenO. tsutsugamushiisolates from mites and those from human patients (H. Y. Lu et al., Am. J. Trop. Med. Hyg. 83:658-663, 2010) were striking, thus highlighting potential risk factors for this emerging zoonotic disease.
Corynebacterium glutamicum Tailored for Efficient Isobutanol Production
Authors: Blombach Bastian , Riester Tanja , Wieschalka Stefan , Ziert Christian , Youn Jung-Won , Wendisch Volker F. , Eikmanns Bernhard J.
Published: March 2011

We recently engineered
Corynebacterium glutamicum
for aerobic production of 2-ketoisovalerate by inactivation of the pyruvate dehydrogenase complex, pyruvate:quinone oxidoreductase, transaminase B, and additional overexpression of the
ilvBNCD
genes, encoding acetohydroxyacid synthase, acetohydroxyacid isomeroreductase, and dihydroxyacid dehydratase. Based on this strain, we engineered
C. glutamicum
for the production of isobutanol from glucose under oxygen deprivation conditions by inactivation of
l
-lactate and malate dehydrogenases, implementation of ketoacid decarboxylase from
Lactococcus lactis
, alcohol dehydrogenase 2 (ADH2) from
Saccharomyces cerevisiae
, and expression of the
pntAB
transhydrogenase genes from
Escherichia coli
. The resulting strain produced isobutanol with a substrate-specific yield (Y
P/S
) of 0.60 ± 0.02 mol per mol of glucose. Interestingly, a chromosomally encoded alcohol dehydrogenase rather than the plasmid-encoded ADH2 from
S. cerevisiae
was involved in isobutanol formation with
C. glutamicum
, and overexpression of the corresponding
adhA
gene increased the Y
P/S
to 0.77 ± 0.01 mol of isobutanol per mol of glucose. Inactivation of the malic enzyme significantly reduced the Y
P/S
, indicating that the metabolic cycle consisting of pyruvate and/or phosphoenolpyruvate carboxylase, malate dehydrogenase, and malic enzyme is responsible for the conversion of NADH+H
+
to NADPH+H
+
. In fed-batch fermentations with an aerobic growth phase and an oxygen-depleted production phase, the most promising strain,
C. glutamicum
Δ
aceE
Δ
pqo
Δ
ilvE
Δ
ldhA
Δ
mdh
(pJC4
ilvBNCD-pntAB
)(pBB1
kivd-adhA
), produced about 175 mM isobutanol, with a volumetric productivity of 4.4 mM h
−1
, and showed an overall Y
P/S
of about 0.48 mol per mol of glucose in the production phase.