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This study aimed to analyze the effect of carvacrol oil on biofilm formation of Enterobacter cloacae. The minimum inhibitory concentration of carvacrol oil against E. cloacae was 256 μg/mL. The biofilm formation was inhibited by carvacrol oil at sub-inhibitory concentrations of 64 and 128 μg/mL. The bacterial cell motilities and biofilm exopolysaccharide (EPS) production were both inhibited by carvacrol oil. Correlation analysis indicated that the inhibition of biofilms in mature stages was related to the decrease of EPS production. The biofilm thickness and excretion of extrapolymeric matrix were reduced by carvacrol oil, as shown by scanning electron microscopy and confocal laser scanning microscopy analyses. Transcriptional analysis showed that carvacrol oil down-regulated 24 of 29 selected genes, including curli fimbriae genes (csgABCEFG) and colonic acid polysaccharide genes (wcaABCDEFKLM, wza, wzb, wzc, gmd, manB, manC) which are required for biofilm formation of E. cloacae. In addition, different antimicrobial agents, including 1% lactic acid, 1% phenyllactic acid, and 1024 μg/mL of carvacrol oil considerably inactivated the bacterial cells in carvacrol-treated biofilms. Hence, carvacrol oil effectively inhibited biofilm formation and EPS production of E. cloacae, which is useful for further inactivating E. cloacae biofilms by other antimicrobial agents.

Catalyst effectiveness for metal carbene formation and reactions has been surveyed using N-(tert-butyl)-3-[(tert-butyldimethylsilyl)oxy]-2-diazo-N-(4-chlorobenzyl)but-3-enamide in the formation of the products from both intramolecular C–H insertion and aromatic cycloaddition. Both products are indicators of metal carbene intermediates, and this system provides a means to assess catalysts for metal carbene formation. Donor–acceptor cyclopropene production from the reactant enoldiazoacetamide has been monitored to assess its formation, and the independently formed cyclopropene has also been used to assess metal carbene formation. Catalysts of rhodium(II), copper(I), silver(I), Pd(II), cationic Au(I), and Zn(II) convert the enoldiazoacetamide to the donor–acceptor cyclopropene which serves as the resting state for the intermediate metal carbene, and both the enoldiazoacetamide and its derivative cyclopropene give the same ratios of insertion to cycloaddition products. Catalysts of copper(II) and ruthenium(II...

Phenylpropargyl diazoacetates exist in equilibrium with 1-phenyl-1,2-dien-1-yl diazoacetate - allenes that are rapidly formed at room temperature through 1,3-acyloxy migration catalyzed by gold(I) or gold(III) compounds, and these catalysts react solely with the π-donor rather than with the diazo group. The product allene of the aryldiazoacetates undergoes rearrangement that is not catalyzed by gold in which the terminal nitrogen of the diazo functional group adds to the central carbon of the allene, initiating a sequence of bond-forming reactions, resulting in the production of 1,5-dihydro-4H-pyrazol-4-ones in good yields. These 1,5-dihydro-4H-pyrazol-4-ones undergo intramolecular 1,3-acyl migration to form an equilibrium mixture and can quantitatively transfer the acyl group to an external nucleophile with formation of 4-hydroxypyrazoles. Reactions of phenylpropargyl phenyldiazoacetates catalyzed by cationic gold complexes are initiated at the diazo functional group to form a gold carbene whose subsequent cascade process (intramolecular addition, then aromatic substitution) results in the formation of a product that is uniquely characteristic of this pathway.

Chiral cyclopentane-fused indolines are synthesized with high regio- and enantiocontrol by formal [3 + 2]-annulation reactions of indoles and electrophilic enol carbenes. High enantioselectivity and exclusive regiocontrol occurred with enoldiazoacetamides using a less sterically encumbered prolinate-ligated dirhodium(II) catalyst in reactions with N-substituted indoles without substituents at the 2- or 3-positions via a selective vinylogous addition process. In this transformation, donor-acceptor cyclopropenes generated from enoldiazoacetamides serve as the carbene precursors to form metal carbene intermediates.

Chiral cyclopentyl β-amino esters are formed catalytically by [3+2] cycloaddition reactions of enecarbamates with electrophilic metalloenolcarbenes in high yield with up to 98 % ee and excellent diastereocontrol. Use of β-silyl-substituted enoldiazoacetates with a chiral dirhodium catalyst and trans-β-arylvinylcarbamates are optimal for this transformation, which occurs with hydrogen-bond association between the vinylcarbamate and the intermediate metalloenolcarbene. Reductive conversion of the protected amino esters forms highly functionalized cyclopentyl β-amino acids and 3-aminocyclopentanones.

Diazo compounds that are well-known to undergo [3 + 2]-cycloaddition provide the first examples of the previously unknown [4 + 2]-cycloaddition with dienes that occur thermally under mild conditions and in high yields. Reactions are initiated from reactants prepared from propargyl aryldiazoacetates that undergo gold(I)-catalyzed rearrangement to activated 1,3-dienyl aryldiazoacetates. These reactions proceed to mixtures of both [4 + 2]-cycloaddition and the 1,3-dienyl aryldiazoacetate after long reaction times. At short reaction times, however, both E- and Z-1,3-dienyl aryldiazoacetates are formed and, after isolation, thermal reactions with the E-isomers form the products from [4 + 2]-cycloaddition with ΔH(‡)298 = 15.6 kcal/mol and ΔS(‡)298 = -27.3 cal/(mol·°C). The Z-isomer is inert to [4 + 2]-cycloaddition under these conditions. The Hammett relationships from aryl-substituted diazo esters (ρ = +0.89) and aryl-substituted dienes (ρ = -1.65) are consistent with the dipolar nature of this transformation.

Select transition metal compounds catalyze metal vinylcarbene formation from cyclopropenes, and their documented reactions include both intermolecular and intramolecular CH insertion and cyclopropanation, as well as [3+3]-cycloaddition. Although known to undergo carbene-like transformations for decades, the uses of cyclopropenes as reactive alternatives to diazo compounds under mild conditions has been limited. However, recently developed donor-acceptor cyclopropenes that are conveniently accessed from enoldiazoacetates and enoldiazoacetamides are effective metallo-vinylcarbene precursors. They provide entry to highly stereoselective metal carbene transformations under reaction conditions that are milder than those required for dinitrogen extrusion from diazo compounds.

A highly efficient two-step asymmetric synthesis of 1H-pyrrol-3(2H)-one derivatives from conveniently accessed 2,3-diketoesters. Aldol products are obtained with up to 89/11 d.r. and 99% ee. Benzilic acid rearrangement produces 1H-pyrrol-3(2H)-ones in good yields without loss of enantioselectivity.

Shelf life of fish packaged under modified atmosphere (MA) is extended, but within the United States, commercial application of MA with impermeable packaging films is restricted due to concerns that botulinum toxin production would precede spoilage when contaminated fish are held at abusive storage temperatures. Use of semipermeable packaging films has been advocated; however, previous studies are inconclusive in determining the oxygen transmission rate (OTR) of a film that is needed to achieve an acceptable margin of safety (i.e., toxin production occurs only after spoilage). This study was conducted to determine the influence of OTR (target OTRs of 3 to 15,000) on the development of spoilage volatiles and toxin in salmon inoculated with type E Clostridium botulinum and subjected to air, vacuum, or 75:25 CO2:N2 MA and storage temperatures of 4, 8, 12, or 16°C. The most dominant headspace volatile peak that was produced during spoilage of samples at 4, 8 or 12°C was a peak, having a Kovats retention index (KI) of 753, and at which external standards of 2- or 3-methyl 1-butanol also eluted. Under anaerobic conditions, both the aerobic microbial populations and the size of the KI 753 spoilage peak were less in inoculated samples compared with uninoculated samples. C. botulinum-inoculated samples that were stored at 12 or 16°C under conditions favorable for anaerobic growth were also characterized by a KI 688 peak. Using a previously developed model that related the percentage of elderly consumers who would prepare a sample having the KI 753 spoilage peak of a specific size, it was determined that for salmon packaged with 3 or 3,000 OTR films under any atmosphere and stored at 12 or 16°C, 2 to 61% of the consumers could potentially prepare toxin-contaminated samples. Hence, when abusive storage conditions are suspected, the fish should not be consumed.

The development of single- and mixed-species biofilms formed by Escherichia coli O157:H7 and Salmonella was observed, and the antimicrobial effectiveness of levulinic acid (LVA) plus sodium dodecyl sulfate (SDS) on the cells in single- and dual-species biofilms was determined. Biofilm-forming ability of single- and mixed-species cultures was observed by crystal violet staining and their resistance to levulinic acid plus SDS was determined by enumeration. Fluorescent protein-labeled E. coli O157:H7 and Salmonella were constructed and the bacterial composition of the biofilms after treatment with levulinic acid plus SDS was visualized by confocal laser scanning microscopy (CLSM). E. coli O157:H7 and Salmonella were antagonistic to each other, being more sensitive to levulinic acid plus SDS in mixed-species biofilms. Images captured by CLSM revealed that E. coli O157:H7 and Salmonella were distributed evenly in the single- and dual-species biofilms, and confirmed that the combination of levulinic acid and SDS was effective in inactivating bacterial cells in biofilms. Results revealed that levulinic acid with SDS may be used as a potential biofilm control intervention.

Conjugated cycloheptene-1,4-dione-enamines are cycloaddition products from a surprising rearrangement in a Au(iii)-catalysed reaction between propargyl aryldiazoacetates and cinnamyl imines. This complex transformation occurs through an initial rapid Au(iii)-catalysed [4+3]-cycloaddition to form dihydroazepinyl aryldiazoacetates followed by a subsequent uncatalyzed domino transformation that occurs by sequential [3+2]-cycloaddition-/nitrogen extrusion and acyloxy migration/retro-Michael addition/tautomerization.

Compost is organic material that has been degraded into a nutrient-stabilized humus-like substance through intense microbial activity, which can provide essential plant nutrients (nitrogen, phosphorus) to aid in the growth of fruits and vegetables. Compost can be generated from animal waste feedstocks; these can contain human pathogens, which can be inactivated through the heat and microbial competition promoted during the composting process. Outbreaks of infections caused by bacterial pathogens such as Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes on fruit and vegetable commodities consumed raw emphasize the importance of minimizing the risk of pathogenic contamination on produce commodities. This review article investigates factors that affect the reduction and survival of bacterial foodborne pathogens during the composting process. Interactions with indigenous microorganisms, carbon:nitrogen ratios, and temperature changes influence pathogen survival, growth, and persistence in finished compost. Understanding the mechanisms of pathogen survival during the composting process and mechanisms that reduce pathogen populations can minimize the risk of pathogen contamination in the cultivation of fruits and vegetables.

An efficient one-pot, three-component reaction of 2,3-diketo esters with amines and ketones has been developed for the synthesis of 1 H-pyrrol-3(2 H)-ones. By using trifluoroacetic acid (TFA) as the additive and acetonitrile (MeCN) as the solvent, this convenient method provides a library of 1 H-pyrrol-3(2 H)-ones in moderate to good yields. The simple protocol features readily available starting materials, a straightforward process, good functional group tolerance, and broad substrate scope.

Dirhodium(II) catalyzed dinitrogen extrusion from diazocarbonyl compounds by 2,6-dichloropyridine-N-oxide forms ketocarbonyl compounds in near-quantitative yields. Reactions occur at room temperature, and the pyridine product does not coordinate with dirhodium(II) to inhibit catalysis. Anhydrous tricarbonyl compounds, as well as dicarbonyl compounds, are conveniently prepared by this methodology, and they have been used in situ for catalytic ene and aldol transformations.

The objective of this study was to determine Salmonella numbers on retail raw chicken carcasses in Guatemala and to phenotypically characterize the isolates (serotyping and antibiotic susceptibility). In total, 300 chicken carcasses were collected from seven departments in Guatemala. Salmonella numbers were determined using the most-probable-number method following the U. S. Department of Agriculture's Food Safety and Inspection Service protocol. In total, 103 isolates were obtained, all of which were tested for antibiotic susceptibility, whereas 46 isolates were serotyped. Overall, Salmonella prevalence and mean number (mean log most probable number per carcass) was 34.3% and 2.3 (95% confidence interval: 2.1 to 2.5), respectively. Significant differences (P0.05) in Salmonella prevalence were found by storage condition (refrigerated or ambient temperature), market type (wet markets, supermarkets, and independent poultry stores), chicken production system (integrated or nonintegrated production company), and chicken skin color (white or yellow). Chickens produced by integrated companies had lower Salmonella numbers (P0.05) than nonintegrated companies, and white-skin carcasses had lower numbers (P0.05) than yellow-skin carcasses. Among 13 different Salmonella serovars identified, Paratyphi B (34.8%) was most prevalent, followed by Heidelberg (16.3%) and Derby (11.6%). Of all the Salmonella isolates, 59.2% were resistant to one to three antibiotics and 13.6% to four or more antibiotics. Among all the serovars obtained, Salmonella Paratyphi B and Heidelberg were the most resistant to the antibiotics tested. Salmonella levels and antibiotic resistant profiles among isolates from raw poultry at the retail market level were high relative to other reports from North and South America. These data can be used by Guatemalan stakeholders to develop risk assessment models and support further research opportunities to control transmission of Salmonella spp. and antibiotic-resistant isolates from chicken meat to humans.

5-Vinyl-pyrrole and 4-hydroxy-indole derivatives are synthesized by metal-free aldol/cyclization/aromatization cascade reactions of in situ generated enamines with 2,3-diketoesters. This convenient atom-economical process produces multifunctional pyrrole and indole products in moderate to good yields.

A formal [3 + 3]-cycloaddition of enoldiazoacetates with donor-acceptor cyclopropanes was realized by the combination of a Lewis acid-catalyzed diastereoselective [3 + 2]-cycloaddition and a subsequent rhodium-catalyzed chemoselective ring expansion. This tandem transformation provides an efficient approach to highly functionalized cyclohexenes.

A one-pot, three-component reaction between a 2,3-diketoester, an imine, and an aldehyde, catalyzed by p-toluenesulfonic acid, produces fully-substituted 3-aminopyrroles that can be converted to pyrrolo[3,2-d]pyrimidine-2,4-dione.

A series of botulism challenge studies were conducted to determine if botulinum toxin would be produced in mussels ( Mytilus edulis ) inoculated with non-proteolytic Clostridium botulinum spores and held under modified atmosphere (MA) packaging conditions at normal (4 °C) and abusive (12 °C) temperatures. Spore mixtures of six strains of non-proteolytic C. botulinum were introduced into live mussels through immersion in a seawater solution with cultured algae. Mussels were packed in a commercial high-oxygen (60–65% O 2 ) MA-package with a buffer, and also packed under a vacuum. Feeding live mussels cultured algae (10 4 cells/ml) with a C. botulinum spore suspension (10 3 spores/ml) in seawater at 4 °C for 6 h resulted in the uptake of spores into mussel tissue (500/g) and the mussel GI tract (100/g). Under all of the experimental conditions evaluated, none of the fresh mussels became toxic, even after spoilage and in the absence of oxygen. However, control samples using tuna or cooked mussel meats became toxic in the absence of oxygen. Botulinum toxin was not produced in fresh mussels packaged under the MA-packaging conditions evaluated, even at an abusive storage temperature (12 °C) for at least 12 days or at normal storage temperate (4 °C) for at least 21 days, which is beyond their shelf life.

Listeria monocytogenes contamination of cantaloupes has become a serious concern as contaminated cantaloupes led to a deadly outbreak in the United States in 2011. To reduce cross-contamination between cantaloupes and to reduce resident populations on contaminated melons, application of sanitizers in packing shed wash water is recommended. The sanitizing agent of 5% levulinic acid and 2% sodium dodecyl sulfate (SDS) applied as a single hurdle in either a simulated dump or dip treatment significantly reduced L. monocytogenes to lower levels at the stem scar compared to a simulated dump treatment employing 200 ppm chlorine, however pathogen reductions on the rind tissue were not significantly different. Double hurdle approaches employing two sequential packing plant treatments with different sanitizers revealed decreased reduction of L. monocytogenes at the stem scar. In contrast, application of sanitizers both in the field and at the packing plant led to greater L. monocytogenes population reductions than if sanitizers were only applied at the packing plant.

β,γ-Unsaturated α-diazocarbonyl compounds possess two reactive sites for electrophilic addition – one at the diazo carbon and the other at the vinylogous γ-position. Controlled by catalyst, divergent transformations are achieved starting from the same starting materials, either by Lewis acid-catalyzed addition or by dirhodium-catalyzed metal carbene reactions. In select cases two catalysts working in combination or in sequence provide a relay for cascade transformations. In this review, we summarize advances in catalyst-dependent divergent transformations of β,γ-unsaturated α-diazocarbonyl compounds and highlight the potential of this exciting research area and the many challenges that remain.

Bis-sigma-(phenyl)-tetrakis-mu-(capralactamato)dirhodium(III), formed by oxidative reactions of dirhodium(II) carboxamidates with arylboronic acids promoted by copper(II), exists in two non-interconvertable conformations. In these conformational isomers the caprolactamate ligands are oriented around dirhodium( III) as a propeller with apparent C4 symmetry and in a biplanar array. Cyclic voltammetry and spectroelectrochemistry as well as Raman and resonance Raman spectroscopy of the conformationally distinct Rh-2(6+) carboxamidates have been obtained in order to explore the origin of their non-interconversion. Differences in their vibrational spectra occur only in the low wavenumber spectral region, but there is a 250 mV difference in oxidation potential between conformational isomers consistent with a substantial difference ground state energies. Notably, spectroelectrochemical data show that oxidation does not result in isomerization. (C) 2014 Elsevier B.V. All rights reserved.

β-Lactam derivatives are produced through intermediate donor–acceptor cyclopropene intermediates in high yield, exclusive cis-diastereoselectivity, and high enantiocontrol in a chiral dirhodium carboxylate catalyzed intramolecular C–H functionalization reaction of enoldiazoacetamides.

Escherichia coli O157:H7 has been the causative agent of many outbreaks associated with leafy green produce consumption. Elucidating the mechanism by which contamination occurs requires monitoring interactions between the pathogen and the plant under typical production conditions. Intentional introduction of virulent strains into fields is not an acceptable practice. As an alternative, attenuated strains of natural isolates have been used as surrogates of the virulent strains; however, the attachment properties and environmental stabilities of these attenuated isolates may differ from the unattenuated outbreak strains. In this study, the Shiga toxin (stx1, stx2, and/or stx2c) genes as well as the eae gene encoding intimin of two E. coli O157:H7 outbreak isolates, F4546 (1997 alfalfa sprout) and K4492 (2006 lettuce), were deleted. Individual gene deletions were confirmed by polymerase chain reaction (PCR) and DNA sequencing. The mutant strains did not produce Shiga toxin. The growth kinetics of these mutant strains under nutrient-rich and minimal conditions were identical to those of their wild-type strains. Attachment to the surface of lettuce leaves was comparable between wild-type/mutant pairs F4546/MD46 and K4492/MD47. Adherence to soil particles was also comparable between the virulent and surrogate pairs, although the F4546/MD46 pair exhibited statistically greater attachment than the K4492/MD47 pair (p≤0.05). Wild-type and mutant pairs F4546/MD46 and K4492/MD47 inoculated into wet or dry soils had statistically similar survival rates over the 7-day storage period at 20°C. A plasmid, pGFPuv, containing green fluorescent protein was transformed into each of the mutant strains, allowing for ease of identification and detection of surrogate strains on plant material or soil. These pGFPuv-containing surrogate strains will enable the investigation of pathogen interaction with plants and soil in the farm production environment where the virulent pathogen cannot be used.

Intra-abdominal infections (IAI) are an important cause of morbidity and are frequently associated with poor prognosis, particularly in high-risk patients. The cornerstones in the management of complicated IAIs are timely effective source control with appropriate antimicrobial therapy. Empiric antimicrobial therapy is important in the management of intra-abdominal infections and must be broad enough to cover all likely organisms because inappropriate initial antimicrobial therapy is associated with poor patient outcomes and the development of bacterial resistance. The overuse of antimicrobials is widely accepted as a major driver of some emerging infections (such as C. difficile), the selection of resistant pathogens in individual patients, and for the continued development of antimicrobial resistance globally. The growing emergence of multi-drug resistant organisms and the limited development of new agents available to counteract them have caused an impending crisis with alarming implications, especially with regards to Gram-negative bacteria. An international task force from 79 different countries has joined this project by sharing a document on the rational use of antimicrobials for patients with IAIs. The project has been termed AGORA (Antimicrobials: A Global Alliance for Optimizing their Rational Use in Intra-Abdominal Infections). The authors hope that AGORA, involving many of the world's leading experts, can actively raise awareness in health workers and can improve prescribing behavior in treating IAIs.

Allylic oxidation of heteroatom substituted cyclic alkenes by tert-butyl hydroperoxide (70% TBHP in water) using catalytic dirhodium caprolactamate [Rh2(cap)4] forms enone products with a variety of 2-substituted cyclic enamides and 3,4-dihyro-2H-pyrans. These reactions occur under mild reaction conditions, are operationally convenient to execute, and are effective for product formation with as low as 0.25 mol% catalyst loading. With heteroatom stabilization of the intermediate allylic free radical two sites for oxidative product formation are possible, and the selectivity of the oxidative process varies with the heteroatom when R = H. Cyclic enamides produce 4-piperidones in good yields when R = alkyl or aryl, but oxidation of 2H-pyrans also gives alkyl cleavage products. Alternative catalysts for TBHP oxidations show comparable selectivities but give lower product yields.

Many types of fermented foods, known throughout the world, have been prepared and consumed for centuries for their nutritional value, flavor and stability. This chapter addresses fermented foods commonly known around the world as well as many that are locally produced. It highlights microbiological diversity of these foods. It also addresses the types of foods include dairy products, cereal-based products, fruit and vegetable products, meat products, locally produced traditional fermented foods and innovative functional foods. Lactic acid bacteria play an important role in fermentation of many commonly consumed foods, such as yogurt, cheese, bread, sausage, pickles and olives. Yogurt is one of the most important cultured dairy products consumed throughout the world. Acetic acid bacteria are another important group of bacteria which are essential in vinegar and cocoa fermentations. Yeasts play an important role in the production of bread and many alcoholic beverages, such as beer and wine.

Listeria monocytogenes is a nonfastidious, widely occurring foodborne pathogen that is a major challenge to the food industry. Enterococcus durans 152, a confirmed L. monocytogenes-control microorganism, was isolated from floor drain samples from a food processing facility. In this study, the two bacteriocins produced by E. durans 152 were characterized and identified as Dur 152A (an enterocin L50A derivative with two amino acid substitutions of I→M) and enterocin L50B. The bacteriocins were then partially purified and evaluated for inhibitory activity to L. monocytogenes in deli ham. Results revealed that at 400 AU/ml, the bacteriocins prevented growth of listeria in deli ham for at least 10 weeks at 8 °C and at least 30 days at 15 °C. For comparison, 500 ppm Nisin controlled listeria growth for up to 6 weeks at 8 °C and up to 18 days at 15 °C. These findings reveal the potential for the bacteriocins of E. durans 152 to serve as anti-listerial agents in deli meat.

The first asymmetric [3+1]-cycloaddition was successfully achieved by copper(I) triflate/double-sidearmed bisoxazoline complex catalyzed reactions of β-triisopropyl-silyl-substituted enoldiazo compounds with sulfur ylides. This methodology delivered a series of chiral cyclobutenes in good yields with high enantio- and diastereoselectivities (up to 99% ee, and >20:1 d.r.). Additionally, the [3+1]-cycloaddition of catalytically generated metallo-enolcarbenes was successfully extended to reaction with a stable benzylidene dichlororuthenium complex.

This workshop, jointly presented by the ILSI North America Technical Committees on Food Microbiology and Sodium, aimed to provide greater knowledge and appreciation of the opportunities and challenges facing the food industry in answering the public health community's call to reduce sodium levels in the food supply. One major challenge is finding effective substitutes for the various antimicrobial and functional roles that sodium plays across different food categories. Sodium plays a critical role in retarding the growth of pathogens and food spoilage bacteria. Moreover, taste is an important factor for consumers when they choose food products, and the flavor changes that occur when salt is reduced or replaced must be considered and ingredients and processes adjusted accordingly. The workshop provided a platform for a multidisciplinary discussion among the microbiology, food science, nutrition, and public health communities to share progress and propose solutions, including the formation of public-private partnerships, to develop coordinated and comprehensive strategies. This paper provides an overview of the issues raised, rather than a specific summary of workshop discussions. The Food and Drug Administration's Draft Guidance for Industry on voluntary sodium reduction and the 2015 US Dietary Guidelines for Americans were released subsequent to the workshop and are also discussed.

Chiral cyclopenta[2,3]cyclopropa[1,2-c]pyran-4-ones have been synthesized via dirhodium(II)-catalyzed intramolecular cyclopropanation of (Z)-1,3-dienyl aryldiazoacetates. High regio-, diastereo-, and enantiocontrol were achieved using chiral dirhodium 2-phthalimide carboxylates. Preferential addition occurs at the 3,4- rather than the 1,2-double bond with the chiral dirhodium catalysts, although both outcomes occur with other transition-metal catalysts.

This chapter provides an overview regarding the advantages of virulotyping over historic serology-based, PCR, based on genes that identify an organism, or enzymatic and biochemical-based analyses of foodborne pathogens in clinical diagnostics and food industry microbiology testing. Traditional identification and characterization methods are designed to detect a given genus, species, serovar or genetic variant of bacteria via enzymatic, genetic or biochemical characterization. Virulotyping is a technique that identifies specific virulence genes within the pathogen’s genome; thus having the discriminatory power to differentiate between a virulent and an avirulent (nonpathogenic) strain of bacterium within a species. As an example, the United States Department of Agriculture, Food Safety and Inspection Service (USDA-FSIS) methods for detection, isolation and identification of Shiga toxin producing E. coli (STEC) use the presence of the eae and stx genes as an initial screen after selective culture enrichment to determine if an adulterant STEC may be present in a raw beef verification sample and as a confirmatory screen of STEC isolates. While virulence testing for STEC is just one example of potential advantages of virulotyping, further benefits may also be realized from virulotyping other common foodborne pathogens (e.g., Campylobacter spp., Salmonella enterica, and Listeria monocytogenes), should avirulent subgroups one day be identified. While PCR and microarray analyses are currently used to perform virulotyping, genome sequencing-based methods may be a more robust and discriminatory technique, when they become practicable for food safety testing and clinical diagnostics. A potential future virulotyping-based medical diagnostics and food testing system might be envisioned in the United States, which could prevent illnesses.

For many years, the U.S. has underutilized its human resources, as evidenced by the pervasive underrepresentation of several racial and ethnic groups within academia in general and the science, technology, engineering, and mathematics (STEM) disciplines, in particular. To address this underutilization, academic departments within U.S. universities must develop grassroots efforts that both recruit and retain students from all segments of the U.S. population. While many departments have noted this need and are earnestly trying to increase the diversity of their programs, very few have successfully achieved this laudable goal. This inquiry into the salient features of successful diversity agendas within STEM academic departments provides key insights into how departments can increase and sustain the doctoral degree attainment of students from underrepresented racial and ethnic minority groups.

Reduction of enteric pathogens in animal manures before field application is essential for mitigating the risk of foodborne illness associated with produce. Aerobic composting of manures has been advocated as an effective treatment for reducing pathogen populations, and heat is a major factor contributing to pathogen inactivation. This study was initiated to determine the potential contribution of both thermal and nonthermal (pH, volatile acids, and ammonia) factors to pathogen inactivation during aerobic composting in bioreactors for mixtures containing manure from various sources (dairy, chicken, and swine). The test mixtures were formulated with an initial moisture content of 60% and a C:N ratio of 20:1, using straw and cottonseed meal as amendments. Mixtures were then inoculated with Salmonella and Listeria monocytogenes labeled with green fluorescent protein at initial populations of ca. 10(7) CFU/g. Three replicate trials of each treatment were conducted. Temperatures within the bioreactors were recorded at 30-min intervals, and duplicate samples were withdrawn daily from two sampling locations within the bioreactor. Significant regression models were derived relating decreases in pathogen populations to the degree of heat generated in the mixture (cumulative heat) and the pH of the mixture on the day before the pathogen losses were calculated (P < 0.0002). Although pathogens in swine manure compost mixtures were inactivated by the third day of composting, very little heat was generated in these mixtures, which were characterized by significantly higher levels of volatile acids compared with the other two compost mixtures. Therefore, volatile acids could help achieve pathogen inactivation when temperatures are too low such as when heat is lost too quickly at the surface of static compost piles or during winter composting.

An enantioselective carbonyl-ene reaction of glyoxylate esters with 1,1-disubstituted alkenes catalyzed by chiral cationic dirhodium(II,III) carboxamidates is described. The paddlewheel dirhodium(II,III) carboxamidates having one open coordination site at each rhodium smoothly catalyze the carbonyl-ene reaction to afford homoallylic alcohol products in good isolated yields with high enantioselectivities.

Oxidations with tert -butyl hydroperoxide (TBHP) catalyzed by dirhodium caprolactamate [Rh 2 (cap) 4 ] provide mild and selective protocols for the oxidative Mannich reaction, conversion of secondary amines to imines, dearomatization of 4-substituted phenols, and oxyfunctionalization of allylic, benzylic, and propargylic positions. The dirhodium catalyst in combination with TBHP is a convenient platform for mechanistic investigations, and their uses point to a variety of directions for the future evolution of the field.

Carbonyl–ene reactions of 2,3-diketoesters catalyzed by [Cu{(S,S)-tBu-box}](SbF6)2 [box=bis(oxazoline)] generate chiral α-functionalized α-hydroxy-β-ketoesters in up to 94 % yield and 97 % ee. The 2,3-diketoesters are conveniently accessed from the corresponding α-diazo-β-ketoester, and a catalyst loading as low as 1.0 mol % can be achieved.

This study investigated the degree of cross-contamination between deli foods and slicers by Listeria monocytogenes, Salmonella, and Escherichia coli O157:H7, and their inactivation by levulinic acid (LA) plus sodium dodecyl sulfate (SDS) on slicers. The transfer rate of pathogens at 5 locations on the contaminated slicers (scenario I) and on food slices (scenario II) was determined. The antimicrobial efficacy of the LA + SDS sanitizers applied either as a liquid or as foam at three concentrations (0.5% LA + 0.05% SDS, 1% LA + 0.1% SDS, and 2% LA + 0.5% SDS) was determined for decontamination of the pathogens on the slicers at 21 °C. After slicing 10 slices, the pathogens recovered from slicer blades were significantly (P 0.05) less than the recovery from some other contact locations (scenario I). With an initial inoculum at approximately 8.5 log CFU/blade, the populations of the pathogens transferred from blades to slices decreased logarithmically (R(2) 0.9, scenario II). Contaminated slicer surfaces sprayed with 1% LA plus 0.1% SDS as a foam (45-55 psi) reduced within 1 min 6.0 to 8.0 log CFU/blade of the pathogens. Results revealed that cross-contamination can occur between deli foods and slicers. Also, LA-based sanitizer applied as foam can be a useful treatment to remove microbial contamination on the slicers.

Studies were done at 21 °C to determine the bactericidal activity of lactic acid, levulinic acid, and sodium dodecyl sulfate (SDS) applied individually and in combination on Shiga toxin-producing Escherichia coli (STEC) in pure culture and to compare the efficacy of lactic acid and levulinic acid plus SDS treatments applied by spray or immersion to inactivate STEC and Salmonella (10(7) CFU/cm2) on beef trim pieces (10 by 10 by 7.5 cm). Application of 3% lactic acid for 2 min to pure cultures was shown to reduce E. coli O26:H11, O45:H2, O111:H8, O103:H2, O121:H2, O145:NM, and O157:H7 populations by 2.1, 0.4, 0.3, 1.4, 0.3, 2.1, and 1.7 log CFU/ml, respectively. Treatment with 0.5% levulinic acid plus 0.05% SDS for1 min reduced the populations of all STEC strains to undetectable levels (6 log/ml reduction). Beef surface temperature was found to affect the bactericidal activity of treatment with 3 % levulinic acid plus 2% SDS (LV-SDS). Treating cold (4 °C) beef trim with LV-SDS at 21, 62, or 81 °C for 30 s reduced E. coli O157:H7 by 1.0, 1.1, or 1.4 log CFU/cm2, respectively, whereas treating beef trim at 8 °C with LV-SDS at 12 °C for 0.1, 1, 3, or 5 min reduced E. coli O157:H7 by 1.4, 2.4, 2.5, or 3.3 log CFU/cm(2), respectively. Spray treatment of beef trim at 4 °C with 5 % lactic acid only reduced the E. coli O157:H7 population by 1.3 log CFU/cm2. Treating beef trim at 8 °C with LV-SDS for 1, 2, or 3 min reduced Salmonella Typhimurium by 2.1, 2.6, and5.0 log CFU/cm2, respectively. Hand massaging the treated beef trim substantially reduced contamination of both pathogens, with no detectable E. coli O157:H7 or Salmonella Typhimurium (5 CFU/cm2) on beef trim pieces treated with LV-SDS. Reduction of E. coli O157:H7 and Salmonella Typhimurium populations was enhanced, but bactericidal activity was affected by the meat temperature.

Conspectus The combination of two or more unsaturated structural units to form cyclic organic compounds is commonly referred to as cycloaddition, and the combination of two unsaturated structural units that forms a six-membered ring is formally either a [5 + 1]-, [4 + 2]-, [2 + 2 + 2]-, or [3 + 3]-cycloaddition. Occurring as concerted or stepwise processes, cycloaddition reactions are among the most useful synthetic constructions in organic chemistry. Of these transformations, the concerted [4 + 2]-cycloaddition, the Diels–Alder reaction, is by far the best known and most widely applied. However, although symmetry disallowed as a concerted process and lacking certifiable examples until recently, stepwise [3 + 3]-cycloadditions offer advantages for the synthesis of a substantial variety of heterocyclic compounds, and they are receiving considerable attention. In this Account, we present the development of stepwise [3 + 3]-cycloaddition reactions from virtual invisibility in the 1990s to a rapidly growing synthetic methodology today, involving organocatalysis or transition metal catalysis. With origins in organometallic or vinyliminium ion chemistry, this area has blossomed into a viable synthetic transformation for the construction of six-membered heterocyclic compounds containing one or more heteroatoms. The development of [3 + 3]-cycloaddition transformations has been achieved through identification of suitable and compatible reactive dipolar adducts and stable dipoles. The reactive dipolar species is an energetic dipolar intermediate that is optimally formed catalytically in the reaction. The stepwise process occurs with the reactive dipolar adduct reacting as an electrophile or as a nucleophile to form the first covalent bond, and this association provides entropic assistance for the construction of the second covalent bond and the overall formal [3 + 3]-cycloaddition. Organocatalysis is well developed for both inter- and intramolecular synthetic transformations, but the potential of transition metal catalysis for [3 + 3]-cycloaddition has only recently emerged. The key to the rapid development of the transition metal-based methodology has been recognition that certain catalytically generated vinylcarbenes are effective dipolar adducts for reactions with stable dipolar compounds, including aryl and vinyl ylides. In particular, metallo-enolcarbenes that are generated catalytically from conveniently prepared stable enoldiazoacetates or from donor–acceptor cyclopropenes are highly effective dipolar adducts for [3 + 3]-cycloaddition. The electron-donating oxygen of the silyl ether enhances electrophilic ring closure to the metal-bound carbon of the initial adduct from vinylogous addition, and this enhancement inhibits the alternative [3 + 2]-cycloaddition across the carbon–carbon double bond of the vinylcarbene. Catalytically generated metallo-enolcarbenes react under mild conditions with a broad spectrum of compatible stable dipoles, including nitrones, azomethine imines, ylides, and certain covalent precursors of stable dipoles, to form [3 + 3]-cycloaddition products having the β-ketoester functionality (in dihydrooxazines, tetrahydropyridazines, pyrazolidinone and pyraxole derivatives, dihydroquinolines, and quinolizidines, for example) in high yield. Two ways to access these metallo-enolcarbenes, either by dinitrogen extrusion from enoldiazoacetate esters or by rearrangement of donor–acceptor cyclopropenes, enhance the versatility of the process. The [3 + 3]-cycloaddition methodology is a complementary strategy to [4 + 2]-cycloaddition for the synthesis of heterocyclic compounds having six-membered rings. High levels of enantioselectivity are obtained with the use of chiral ligands on transition metal catalysts that include those on dirhodium(II) and silver(I).

The objectives of this study were to quantify Salmonella counts on retail raw poultry meat in Vietnam and to phenotypically characterize (serovars and antibiotic resistance) the isolates. A total of 300 chicken carcasses were collected from two cities and two provinces in Vietnam. Salmonella counts on the samples were determined according to the most-probable-number (MPN) method of the U.S. Department of Agriculture, Food Safety and Inspection Service (USDA-FSIS). A total of 457 isolates were serotyped and tested for antibiotic susceptibility. Overall, 48.7% of chicken samples were Salmonella positive with a count of 2.0 log MPN per carcass. There were no significant differences (P0.05) in log MPN per carcass by the study variables (market type, storage condition, and chicken production system). There was a significant difference (P0.05) in Salmonella-positive prevalence by chicken production system. Among the 22 Salmonella serovars identified, Albany was the most frequent (34.1%), followed by Agona (15.5%) and Dabou (8.8%). Resistance to at least one antibiotic was common (i.e., 73.3%), with high resistance to tetracycline (59.1%) and ampicillin (41.6%). Resistance to three antibiotics was the most frequently found multidrug resistance profile (17.7%, n = 81); the profile that was resistant to the highest number of drugs was resistant to nine antibiotics (0.7%, n = 3). Only Salmonella Albany posed phenotypic resistance to ceftriaxone (a drug of choice to treat severe cases of salmonellosis). The data revealed that, whereas Salmonella prevalence on raw poultry was high (48.7%), counts were low, which suggests that the exposure risk to Salmonella is low. However, improper storage of raw chicken meat and cross-contamination may increase Salmonella cell counts and pose a greater risk for infection. These data may be helpful in developing risk assessment models and preventing the transmission of foodborne Salmonella from poultry to humans in Vietnam.

Catalyst-dependent [4 + 2]-cycloaddition reactions of azoalkenes from α-halohydrazones with enol diazoacetates have been developed. A [4 + 2]-cycloaddition of enol diazoacetates with in situ formed azoalkenes produces tetrahydropyridazinyl-substituted diazoacetates promoted by only Cs2CO3. In contrast, donor–acceptor cyclopropenes, which are formed in situ from enol diazoacetates by Rh2(OAc)4-catalyzed dinitrogen extrusion, undergo [4 + 2]-cycloaddition with azoalkenes to yield bicyclo[4.1.0]tetrahydropyridazines. These stable cycloaddition products undergo subsequent one-step transformations to form 6-alkylidenetetrahydropyridazines and 4,5,6,7-tetrahydro-1,2-diazepine derivatives in good yields.

Chiral cyclopentyl β-amino esters are formed catalytically by [3+2] cycloaddition reactions of enecarbamates with electrophilic metalloenolcarbenes in high yield with up to 98 % ee and excellent diastereocontrol. Use of β-silyl-substituted enoldiazoacetates with a chiral dirhodium catalyst and trans-β-arylvinylcarbamates are optimal for this transformation, which occurs with hydrogen-bond association between the vinylcarbamate and the intermediate metalloenolcarbene. Reductive conversion of the protected amino esters forms highly functionalized cyclopentyl β-amino acids and 3-aminocyclopentanones.

Summary RNA-binding proteins play crucial roles in directing RNA translation to neuronal synapses. Staufen2 (Stau2) has been implicated in both dendritic RNA localization and synaptic plasticity in mammalian neurons. Here, we report the identification of functionally relevant Stau2 target mRNAs in neurons. The majority of Stau2-copurifying mRNAs expressed in the hippocampus are present in neuronal processes, further implicating Stau2 in dendritic mRNA regulation. Stau2 targets are enriched for secondary structures similar to those identified in the 3′ UTRs of Drosophila Staufen targets. Next, we show that Stau2 regulates steady-state levels of many neuronal RNAs and that its targets are predominantly downregulated in Stau2-deficient neurons. Detailed analysis confirms that Stau2 stabilizes the expression of one synaptic signaling component, the regulator of G protein signaling 4 ( Rgs4 ) mRNA, via its 3′ UTR. This study defines the global impact of Stau2 on mRNAs in neurons, revealing a role in stabilization of the levels of synaptic targets.

AIMS Three soils that varied in their physicochemical characteristics and microbial diversity were inoculated with Escherichia coli O157:H7 and Salmonella to determine the relative impact of abiotic and biotic factors on the pathogens' survival when the soil was held at 25°C. METHODS AND RESULTS Three soils that were classified as having low, medium and high microbial diversity were divided into two batches for adjustment to 20% of water-holding capacity and to 40% of water-holding capacity. Soils were inoculated with both green fluorescent-labelled E. coli O157:H7 and red fluorescent-labelled Salmonella (5 log CFU g(-1) dry weight) and held at 25°C. Pathogens inoculated into an acidic soil died off within 9 weeks, whereas they were still detected in the other two soils by enrichment culture after 18 weeks. Moisture did not affect inactivation of E. coli O157:H7, but did affect Salmonella inactivation in soil having the greatest organic load and microbial diversity. Using multiple linear regression analysis, 98.7% of the variability in the inactivation rate for E. coli O157:H7 was explained by a model that included the variables of initial pH and electrical conductivity. Salmonella's inactivation rate was predicted by a model that included pH and initial cell numbers of copiotrophic and oligotrophic bacteria. CONCLUSION This study provided evidence of specific properties that impact inactivation of E. coli O157:H7 and Salmonella in soils at 25°C. SIGNIFICANCE AND IMPACT OF THE STUDY Identification of factors influential in the die-off of enteric pathogens will assist in developing guidelines for safe intervals between field contamination events and planting or harvesting of fresh-cut produce crops.