Live and inactivated vaccine regimens against caecal Salmonella Typhimurium colonisation in laying hens.

OBJECTIVE: In Australia, Salmonella serovar Typhimurium (S. Typhimurium) is the predominant zoonotic serovar in humans and is frequently isolated from layer hens. Vaccination against this serovar has been previously shown to be effective in broilers and the aim of this current study was to assess and determine the best vaccination strategy (live or inactivated) to minimise caecal colonisation by S. Typhimurium. METHODS: A long-term experiment (56 weeks) was conducted on ISABROWN pullets using a commercial live aroA deleted mutant S. Typhimurium vaccine and an autogenous inactivated multivalent Salmonella vaccine (containing serovars Typhimurium, Infantis, Montevideo and Zanzibar). These vaccines were administered PO or by SC or IM injection, either alone or in combination. Pullets were vaccinated throughout rearing (to 18 weeks of age) and sequentially bled for antibody titre levels. The birds, vaccinated and controls, were challenged orally with a field isolate of S. Typhimurium at different ages, held for 21 days post-challenge, then euthanased and their caeca cultured for the presence of Salmonella. RESULTS: None of the oral live-vaccinated groups exhibited lasting protection. When administered twice, the inactivated vaccine gave significant protection at 17 weeks of age and the live vaccine given by SC injection given twice produced significant protection at 17, 25 and 34 weeks. CONCLUSIONS: Vaccination regimens that included parenteral administration of live or inactivated vaccines and thus achieved positive serum antibody levels were able to provide protection against challenge. Hence, vaccination may play a useful role in a management strategy for Salmonella carriage in layer flocks.

Intradiscal Pressure Changes during Manual Cervical Distraction: A Cadaveric Study.

The objective of this study was to measure intradiscal pressure (IDP) changes in the lower cervical spine during a manual cervical distraction (MCD) procedure. Incisions were made anteriorly, and pressure transducers were inserted into each nucleus at lower cervical discs. Four skilled doctors of chiropractic (DCs) performed MCD procedure on nine specimens in prone position with contacts at C5 or at C6 vertebrae with the headpiece in different positions. IDP changes, traction forces, and manually applied posterior-to-anterior forces were analyzed using descriptive statistics. IDP decreases were observed during MCD procedure at all lower cervical levels C4-C5, C5-C6, and C6-C7. The mean IDP decreases were as high as 168.7 KPa. Mean traction forces were as high as 119.2 N. Posterior-to-anterior forces applied during manual traction were as high as 82.6 N. Intraclinician reliability for IDP decrease was high for all four DCs. While two DCs had high intraclinician reliability for applied traction force, the other two DCs demonstrated only moderate reliability. IDP decreases were greatest during moving flexion and traction. They were progressevely less pronouced with neutral traction, fixed flexion and traction, and generalized traction.

Classification of Salmonella enterica serotypes from Australian poultry using repetitive sequence-based PCR.

AIMS: To evaluate a semi-automated repetitive extragenic palindromic sequence-based PCR (rep-PCR) system for the classification of Salmonella serotypes from Australian poultry. METHODS AND RESULTS: Using a DNA fingerprint library within the DiversiLab(®) System, four separate databases were constructed (serogroup B, C, E and Other). These databases contained 483 serologically confirmed (reference laboratory) Salmonella isolates. A blinded set of Salmonella cultures (n = 155) were typed by rep-PCR, matched against the internal library and compared with traditional serotyping. The predicted (Kullback-Leibler) serotype of 143 (92·3%) isolates matched traditional typing (P < 0·05). Of the 12 (7·7%) remaining isolates, ten (6·5%) resulted in 'No Match', one (0·65%) was incorrectly matched to the library (Salm. subsp 1 ser 4,12:-:-), and the other (0·65%) was referenced as Salm. ser. Sofia, whereas rep-PCR and in-house serotyping concurred as Salmonella serovar Typhimurium. Financial analysis showed higher material cost (215%) and a lower labour component (47·5%) for rep-PCR compared with serotyping. CONCLUSION: The DiversiLab(®) System, with serogroup databases, was successfully implemented as an adjunct for reference serotyping of Salmonella enterica. SIGNIFICANCE AND IMPACT OF THE STUDY: The DiversiLab(®) System platform is a cost-effective and easy-to-use system, which can putatively determine Salmonella enterica serotypes within a few hours.

A validated miniaturized MPN method, based on ISO 6579:2002, for the enumeration of Salmonella from poultry matrices.

AIMS: To validate the effectiveness of a miniaturized most probable number method (mMPN) in enumerating Salmonella from poultry matrices. METHODS AND RESULTS: A MPN was developed, based on the ISO 6579:2002 method using modified semi-solid Rappaport-Vassiliadis media as the sole selective medium. The validation of the mMPN was shown to not differ significantly from, at the 95% confidence level (Student's t-test P = 0·357) to, the traditional 9-tube MPN (tMPN) using pure cultures of Salmonella ser. Typhimurium, Infantis, Montevideo, Muenster and Salmonella subsp II 1,4,12,27:b:[e,n,x] (Sofia). The validation of naturally and artificially contaminated poultry matrices (carcasses, scald tank water, faeces, caeca and feed) showed that detection using the mMPN compared well to the ISO 6572:2002; sensitivity (92%), specificity (97%) and agreement (KAPPA 0·72). The quantitative comparison between the tMPN and mMPN methods showed that 92% of enumerations were less than ± 1 log different (Student's t-test = 0·13). Financial analysis showed that the mMPN required 64% less media and 56% less labour than the tMPN. CONCLUSION: The mMPN is a consistent, easy to automate method for the enumeration of Salmonella from different poultry matrices. SIGNIFICANCE AND IMPACT OF STUDY: The miniaturized MPN reduces the material and labour cost of the method and enables the uniform and accurate measurement of the effectiveness of intervention strategies in the control of Salmonella colonization of poultry.

Advances in enteropathogen control in poultry production.

Poultry meat has been associated frequently and consistently with the transmission of enteric pathogens, including Salmonella and Campylobacter. This association has resulted in the development of HACCP-based intervention strategies. These strategies (hurdles) begin with elite breeder flocks and filter down the production pyramid. These hurdles include those already established, such as biosecurity, vaccination, competitive exclusion, pre- and probiotics, feed and water control, and those more experimental, such as bacteriophage or immunoglobulin therapy. The reduction in enteropathogens entering the processing plant, which employs critical control points, further reduce the exposure of consumers to these organisms. The synergistic application of hurdles will result in an environment that is restrictive and detrimental to enteropathogen colonization and contamination.

Evaluation of TECRA broth, Bolton broth, and direct plating for recovery of Campylobacter spp, from broiler carcass rinsates from commercial processing plants.

The purpose of this study was to compare a conventional culture broth method (Bolton enrichment), a newly developed proprietary broth method (TECRA Campylobacter enrichment), and direct plating for recovery of Campylobacter spp. from chicken carcass rinsates. Whole carcass rinses were taken from 140 carcasses at rehang (immediately after defeathering but before evisceration) and from 140 carcasses at postchill from eight different processing plants in the United States. The rinsate samples were packed in ice and shipped overnight to the laboratory. Aliquots of the rinsate were transferred into Bolton and TECRA enrichment broths and were direct plated. Standard laboratory procedures with Campy-cefex plates were followed for recovery of Campylobacter spp. For rehang carcasses, 94% were positive for Campylobacter spp. with the TECRA enrichment broth and 74% were positive with the Bolton enrichment broth. For postchill carcasses, 74% were positive for Campylobacter spp. with the TECRA enrichment broth and 71% were positive with the Bolton enrichment broth. Compared with the Bolton enrichment broth, TECRA enrichment broth significantly suppressed non-Campylobacter microflora (P < 0.05). Overall, TECRA enrichment broth yielded an 11% higher total number of Campylobacter-positive samples compared with the Bolton enrichment broth. Campylobacter spp. detection in postchill samples was significantly greater (P < 0.05) by enrichment (84%) than by direct plating (19%). The high number of Campylobacter-positive samples obtained with all procedures indicated that 99% of the carcass rinsates obtained at rehang and 84% obtained at postchill contained Campylobacter spp.

Cronobacter ('Enterobacter sakazakii'): current status and future prospects.

The genus Cronobacter accommodates the 16 biogroups of the emerging opportunistic pathogen known formerly as Enterobacter sakazakii. Cronobacter spp. are occasional contaminants of milk powder and, consequently, powdered infant formula and represent a significant health risk to neonates. This review presents current knowledge of the food safety aspects of Cronobacter, particularly in infant formula milk powder. Sources of contamination, ecology, disease characteristics and risk management strategies are discussed. Future directions for research are indicated, with a particular focus on the management of this increasingly important bacterium in the production environment.

Cereulide, the emetic toxin of Bacillus cereus, is putatively a product of nonribosomal peptide synthesis.

AIMS: To determine if cereulide, the emetic toxin produced by Bacillus cereus, is produced by a nonribosomal peptide synthetase (NRPS). METHODS AND RESULTS: NC Y, an emetic strain of Bacillus cereus, was examined for a NRPS gene using PCR with primers recognizing a fragment of a NRPS gene from the cyanobacterium Microcystis. The amplicon was sequenced and compared with other gene sequences using BLAST analysis, which showed that the amplicon from strain NC Y was similar in sequence to peptide synthetase genes in other micro-organisms, including Bacillus subtilis and B. brevis, while no such sequence was found in the complete genome sequence of a nonemetic strain of B. cereus. Specific PCR primers were then designed and used to screen 40 B. cereus isolates previously implicated in outbreaks of foodborne illness. The isolates were also screened for toxin production using the MTT cell cytotoxicity assay. PCR and MTT assay screening of the B. cereus isolates revealed a high correlation between the presence of the NRPS gene and cereulide production. CONCLUSIONS: The results indicate that cereulide is produced by a NRPS complex. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study to provide evidence identifying the mechanism of production of cereulide, the emetic toxin of B. cereus. The PCR primers developed in the study allow determination of the potential for cereulide production among isolates of B. cereus.

Modafinil: a gift to portmanteau.

This article reviews the use of modafinil in palliative care, and its use in promoting vigilance and wakefulness. Modafinil's therapeutic and chemical class, mechanism of action, pharmacokinetics, indications, dosage, side effects, and drug interactions are also discussed. Modafinil seems to fit the requirements of portmanteau, using one drug to treat multiple symptoms.

The growth, properties and interactions of yeasts and bacteria associated with the maturation of Camembert and blue-veined cheeses.

The growth of yeasts and bacteria were monitored during the maturation of Camembert and blue-veined cheese produced in Australia. Yeasts were prominent throughout maturation, growing to 10(5)-10(9)/g, depending on the manufacturer. Debaryomyces hansenii predominated, but there were lesser, inconsistent contributions from Yarrowia lipolytica. Of the non-lactic acid bacteria, Acinetobacter species were significant during the maturation of Camembert but not blue-veined cheeses, and grew to 10(6)-10(8) cfu/g. Staphylococcus and Micrococcus species were consistently isolated from the cheeses with Staphylococcus xylosus growing to 10(5)-10(9) cfu/g, depending on the product. Lactic acid bacteria (10(7)-10(9) cfu/g) were present throughout maturation but were not identified. Interactions between the various yeasts and bacterial isolates were examined. Several strains of D. hansenii exhibited killer activity but not against Y. lipolytica. None of the yeasts were antagonistic towards the bacteria but some strains of D. hansenii enhanced the growth of Y. lipolytica and S. xylosus. The yeast and bacterial isolates exhibited various degrees of extracellular proteolytic and lipolytic activities.

Schwannoma: challenging diagnosis.

OBJECTIVE: To discuss the presentation of a schwannoma in a 30-year-old man and to discuss the clincial features of this tumor. CLINICAL FEATURES: The patient had lower right back and abdominal pain that was made worse by any jarring motion. Magnetic resonance imaging showed an intradural extramedullary mass of the thoracic spine behind the T10 vertebral body, which was found to be a schwannoma. INTERVENTION AND OUTCOME: A full laminectomy of T10 and partial laminectomies of T9 and T11 allowed removal of the tumor. CONCLUSION: When undiagnosed abdominal pain is present, spinal tumor should be considered one possible diagnosis.

cDNA array analysis of cag pathogenicity island-associated Helicobacter pylori epithelial cell response genes.

Helicobacter pylori strains containing the cag pathogenicity island (PAI) induce NF-kappaB activation and interleukin-8 secretion in gastric epithelial cells. The aim of this study was to investigate changes in epithelial gene expression induced by cag PAI-positive and -negative strains of H. pylori using high-density cDNA array hybridization technology. Radio-labeled cDNA prepared from H. pylori-infected Kato 3 gastric epithelial cells was hybridized to high-density cDNA arrays to identify changes in epithelial gene expression compared to noninfected controls. In vivo expression of selected, differentially expressed genes was examined by reverse transcription-PCR analysis of H. pylori-positive and -negative gastric mucosa. Screening of ca. 57,800 cDNAs identified 208 known genes and 48 novel genes and/or expressed sequence tags of unknown function to be differentially expressed in Kato 3 cells following H. pylori infection. Marked differences in gene expression profiles were observed following cag PAI-positive and cag PAI-negative infection with 15 novel cDNAs and 92 known genes being differentially expressed. H. pylori was found to change the expression of genes encoding growth factors and cytokine/chemokines and their receptors, apoptosis proteins, transcription factors and metalloprotease-disintegrin proteins (ADAMs), and tissue inhibitors of metalloproteinases. Gastric differential expression of selected known genes (amphiregulin and ADAM 10) and a novel gene (HPYR1) was confirmed in vivo in patients with H. pylori infection. Confirmation of the in vivo expression of selected genes demonstrates the usefulness of this approach for investigating pathogen-induced changes in host gene expression.

Dynamic two-dimensional fluorescence correlation spectroscopy. Generalized correlation and experimental factors.

Dynamic two-dimensional fluorescence correlation spectroscopy (2D FCS) is presented in the general form. Dynamic 2D FCS evaluates the time correlation function between two wavelength axes when an external perturbation is applied to the sample. It displays the vibronic features with similar time response functions in the synchronous correlation spectrum and the features with different time responses in the asynchronous correlation spectrum. The correlation analysis allows detailed assignments of the vibronic spectra of multicomponent samples. The emission-emission 2D FCS has proven to be able to resolve spectra with substantial overlaps, of species in equilibrium with each other, and of reacting species whose kinetic constants are linked and multiexponential. Similarly, the correlation analysis between excitation wavelengths allows the assignment of the excitation bands to fluorescent components. When a sinusoidal light source is used to excite the sample, the excitation-emission correlation requires the collection of only four spectra, two in-phase and two quadrature. The two-dimensional excitation-emission correlation analysis uncovers the association between the excitation and the emission vibronic features, enabling the complete assignment of the component spectra. The band associations and spectral assignments are facilitated by the two-dimensional phase map that is constructed from the synchronous and asynchronous correlation spectra. Spectral resolution can be optimized by varying the frequency of excitation and is not influenced by the detector phase angle used to collect the spectra. The resolution power of the 2D FCS is demonstrated with the retrieval of the anthracene emission spectrum from a pyrene-anthracene mixture when it contributes only 4% to the total fluorescence intensity.

C-glycosides to fused polycyclic ethers. An efficient entry into the A-D ring system of gambierol.

[reaction: see text]. This Letter describes our use of C-glycosides to synthesize the A-D ring system of the marine ladder toxin gambierol in 20 steps.

C-glycosides to fused polycyclic ethers. A formal synthesis of (+/-)-hemibrevetoxin B.

This paper describes a formal total synthesis of the marine ladder toxin hemibrevetoxin B from Danishefsky's dienes. This approach couples the generation of C-glycosides from cyclic enol ethers with metathesis or acid-mediated annulation reactions. The result is a highly efficient synthesis of the tetracyclic ring system of hemibrevetoxin B.

Applications of nylon membrane arrays to gene expression analysis.

Gene expression analyses by hybridization of probes derived from mRNA to cDNA targets arrayed on a nylon membranes have been performed with increasing frequency and success over the last decade. While the initial costs of generating arrays are moderately high, they are now available commercially as complete packages which include the membranes and associated image analysis software for acquisition and processing of the data. Arrays can be used to generate information concerning the expression of mRNA from cells treated with various agents or from different tissues, e.g. comparing diseased with normal controls. To date, many groups, including immunologists, have used this technology to examine gene expression within their area of biological interest. The main characteristic of these systems is the large amount of data generated, since the expression of many thousands of genes are measured in parallel. The main practical issues are sensitivity of detection, reproducibility, comparability with other systems (e.g. Northern blots) and processing of data. Some of the significant applications of nylon array technology to date are reviewed in this chapter, and with these issues in mind, we include a discussion of our own experiences in this area.

Fluorescent detection of Zn(2+)-rich vesicles with Zinquin: mechanism of action in lipid environments.

High concentrations of free Zn2+ ions are found in certain glutamatergic synaptic vesicles in the mammalian brain. These terminals can be visualized histochemically with quinoline sulfonamide compounds that form fluorescent complexes with Zn2+. The present study was undertaken to examine the interaction of the water-soluble quinoline sulfonamide probe, Zinquin (2-methyl-8-(toluene-p-sulfonamido)-6-quinolyloxyacetic acid) with the complex heterogeneous cellular environment. Experiments on rat hippocampal and neocortical slices gave indications that Zinquin in its free acid form was able to diffuse across the plasma and synaptic vesicle membranes. Further experiments were undertaken on unilamellar liposomes to study the interaction of Zinquin and its metal complexes in membranes. These experiments confirmed that Zinquin is able to diffuse across lipid bilayers. Steady-state and time-resolved fluorimetric studies showed that Zinquin in aqueous solution mainly forms a 1:2 (metal:ligand) complex with small amounts of a 1:1 complex. Formation of the 1:1 complex was favored by the presence of lipid, suggesting that it partitions into membranes. Evidence is presented that Zinquin can act as a Zn(2+)-ionophore, exchanging Zn2+ for two protons. The presence of a pH gradient across vesicles traps the Zn(2+)-probe complex within the vesicles. Zinquin is useful as a qualitative probe for detecting the presence of vesicular Zn2+; however, its tendency to partition into membranes and to serve as an ionophore should be borne in mind.

Contemporary and emergent pharmacological therapies for chronic pain: nonopioid analgesia.

The American Pain Society estimates that 50 million Americans are partially or totally disabled by pain. This striking statistic is certain to increase as our population continues to age. In order to combat this growing problem, healthcare professionals must arm themselves with information. By developing the appropriate pain assessment skills and by staying abreast of the rapidly-changing therapies used in pain management, clinicians can dramatically impact the quality of life of those living with pain.

Aluminum- and boron-mediated c-glycoside synthesis from 1,2-anhydroglycosides.

[reaction: see text]This letter describes a single flask strategy to the synthesis of alpha-C-glycosides from glycals. This protocol couples a glycal epoxidation reaction with a C-2 alkoxy-directed carbon-carbon bond-forming reaction.