Staphylococcus aureus is a versatile pathogen capable of causing life-threatening infections. Many of its cell wall and exoproduct virulence determinants are controlled via the accessory gene regulator (agr). Although considered primarily as an extracellular pathogen, it is now recognized that S. aureus can be internalized by epithelial and endothelial cells. Traditional experimental approaches to investigate bacterial internalization are extremely time-consuming and notoriously irreproducible. We present here a new reporter gene method to assess intracellular growth of S. aureus in MAC-T cells that utilizes a gfp-luxABCDE reporter operon under the control of the Bacillus megaterium xylA promoter, which in S. aureus is expressed in a growth-dependent manner. This facilitates assessment of the growth of internalized bacteria in a nondestructive assay. The dual gfplux reporter cassette was also evaluated as a reporter of agr expression and used to monitor the temporal induction of agr during the MAC-T internalization process. The data obtained suggest that agr induction occurs prior to endosomal lysis and that agr-regulated exoproteins appear to be required prior to the release and replication of S. aureus within the infected MAC-T cells.Staphylococcus aureus is the etiologic agent of numerous infections in humans and domesticated animals and has been implicated in a multitude of diseases, ranging from minor wound infections to more serious diseases, including endocarditis, osteomyelitis, and septic shock (reviewed by Projan and Novick [34]). The expression of many S. aureus virulence factors is under the control of the accessory gene regulator (agr) which, on entering post-exponential phase, downregulates the production of cell surface-associated proteins and upregulates the expression of secreted toxins and extracellular enzymes (28,33,38). The role of the agr regulon is supported by in vivo studies, which show that agr mutants are greatly attenuated in several animal models, including intramammary infections (13), arthritis in mice (1), and endocarditis in rabbits (7). The agr locus is a quorum-sensing-regulated system activated by autoinducing peptide pheromone (AIP) (21, 25). The agr locus consists of two divergent transcriptional units, RNAII and RNAIII, which are under the control of the P2 and P3 promoters, respectively (reviewed by Novick and Muir [30]). RNAII is a polycistronic mRNA that encodes the agrB and agrD genes required for the synthesis of the AIP and also the two component signal transduction proteins, AgrA and AgrC, which are responsible for sensing and responding to the AIP. RNAIII is the effector molecule in the agr regulon acting primarily at the level of gene transcription. Different S. aureus strains produce AIPs with distinct structures, and strains can be grouped on this basis since they will activate the agr response of strains within the same group and inhibit the agr response of strains from different groups by competitive inhibition (21,30). This inhibitory action of AIPs has identi...

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Retail poultry products are widely purported as the major infection vehicle for human campylobacteriosis. Numerous intervention strategies have sought to reduce Campylobacter contamination on broiler carcasses in the abattoir. This study reports the efficacy of bacteriophage in reducing the number of recoverable Campylobacter jejuni cells on artificially contaminated chicken skin

The ability of phages to survive processing is an important aspect of their potential use in the biocontrol of Campylobacter in poultry production. To this end, we have developed a procedure to recover Campylobacter bacteriophages from chilled and frozen retail poultry and have validated the sensitivity of the method by using a characterized Campylobacter phage (i.e., NCTC 12674). By using this method, we have shown that Campylobacter phages can survive on retail chicken under commercial storage conditions. Retail chicken portions purchased in the United Kingdom were screened for the presence of endogenous Campylobacter phages. Thirty-four Campylobacter bacteriophages were isolated from 300 chilled retail chicken portions, but none could be recovered from 150 frozen chicken portions. The phage isolates were characterized according to their lytic profiles, morphology, and genome size. The free-range products were significantly more likely to harbor phages (P < 0.001 by single-factor analysis of variance) than were standard or economy products. This study demonstrates that Campylobacter bacteriophages, along with their hosts, can survive commercial poultry processing procedures and that the phages exhibited a wide range of recovery rates from chicken skin stored at 4°C.