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Evolutionary and Functional Relationships among the Nontypeable Haemophilus influenzae HMW Family of Adhesins,. Amy Z. Buscher, 2004.Nontypeable Haemophilus influenzae (NTHi) is a common cause of localized respiratory tract disease and initiates infection by colonizing the nasopharynx . Approximately 75 to 80% of NTHi clinical isolates produce proteins that belong to the HMW family of adhesins, which are believed to facilitate colonization . The prototype HMW adhesins are designated HMW1 and HMW2 and were identified in NTHi strain 12 . HMW1 and HMW2 are 71% identical and 80% similar overall, yet display differing cellular binding specificities . In the present study we set out to define more clearly the relationships between HMW1 and HMW2 and other members of the HMW family of adhesins . PCR analysis of 49 epidemiologically distinct isolates revealed that all strains possessing hmw genes as determined by Southern analysis contain two hmw loci in conserved, unlinked physical locations on the chromosome . Functional analysis of the HMW adhesins produced by three unrelated strains demonstrated that each isolate possesses one protein with HMW1-like adherence properties and another with HMW2-like adherence properties . These findings suggest that the hmw1 and hmw2 loci may have arisen via a gene duplication event in an ancestral strain . In addition, they support the hypothesis that the distinct binding specificities of HMW1 and HMW2 emerged early and have persisted over time, suggesting an ongoing selective advantage . Acquired Bacitracin Resistance in Enterococcus faecalis Is Mediated by an ABC Transporter and a Novel Regulatory Protein, BcrR. Janet M. Manson, 2004.Bacitracin resistance (bacitracin MIC,  256 µg ml–1) has been reported in Enterococcus faecalis, and in the present study we report on the genetic basis for this resistance . Mutagenesis was carried out with transposon Tn917 to select for E . faecalis mutants with decreased resistance to bacitracin . Two bacitracin-sensitive mutants (MICs, 32 µg ml–1) were obtained and Tn917 insertions were mapped to genes designated bcrA and bcrB . The amino acid sequences of BcrA (ATP-binding domain) and BrcB (membrane-spanning domain) are predicted to constitute a homodimeric ATP-binding cassette (ABC) transporter, the function of which is essential for bacitracin resistance in E . faecalis . The bcrA and bcrB genes were organized in an operon with a third gene, bcrD, that had homology to undecaprenol kinases . Northern analysis demonstrated that bcrA, bcrB, and bcrD were transcribed as a polycistronic message that was induced by increasing concentrations of bacitracin but not by other cell wall-active antimicrobials (e.g., vancomycin) . Upstream of the bcrABD operon was a putative regulatory gene, bcrR . The bcrR gene was expressed constitutively, and deletion of bcrR resulted in a bacitracin-sensitive phenotype . No bcrABD expression was observed in a bcrR mutant, suggesting that BcrR is an activator of genes essential for bacitracin resistance (i.e., bcrABD) . The bacitracin resistance genes were found to be located on a plasmid that transferred at a high frequency to E . faecalis strain JH2-2 . This report represents the first description of genes that are essential for acquired bacitracin resistance in E . faecalis .
Extended-Spectrum-Cephalosporin Resistance in Salmonella enterica Isolates of Animal Origin. Jeffrey T. Gray, 2004.A total of 112 out of 5,709 Salmonella enterica isolates from domestic animal species exhibited decreased susceptibilities to ceftiofur and ceftriaxone, and each possessed the blaCMY gene . Ten Salmonella serotypes were significantly more likely to include resistant isolates . Isolates from turkeys, horses, cats, and dogs were significantly more likely to include resistant isolates . The C-Terminal Flexible Domain of the Heme Chaperone CcmE Is Important but Not Essential for Its Function. Elisabeth Enggist, 2003.CcmE is a heme chaperone active in the cytochrome c maturation pathway of Escherichia coli . It first binds heme covalently to strictly conserved histidine H130 and subsequently delivers it to apo-cytochrome c . The recently solved structure of soluble CcmE revealed a compact core consisting of a ß-barrel and a flexible C-terminal domain with a short  -helical turn . In order to elucidate the function of this poorly conserved domain, CcmE was truncated stepwise from the C terminus . Removal of all 29 amino acids up to crucial histidine 130 did not abolish heme binding completely . For detectable transfer of heme to type c cytochromes, only one additional residue, D131, was required, and for efficient cytochrome c maturation, the seven-residue sequence 131DENYTPP137 was required . When soluble forms of CcmE were expressed in the periplasm, the C-terminal domain had to be slightly longer to allow detection of holo-CcmE . Soluble full-length CcmE had low activity in cytochrome c maturation, indicating the importance of the N-terminal membrane anchor for the in vivo function of CcmE .
A Multifunction ABC Transporter (Opt) Contributes to Diversity of Peptide Uptake Specificity within the Genus Lactococcus. Mauld Lamarque, 2004.Growth of Lactococcus lactis in milk depends on the utilization of extracellular peptides . Up to now, oligopeptide uptake was thought to be due only to the ABC transporter Opp . Nevertheless, analysis of several Opp-deficient L . lactis strains revealed the implication of a second oligopeptide ABC transporter, the so-called Opt system . Both transporters are expressed in wild-type strains such as L . lactis SK11 and Wg2, whereas the plasmid-free strains MG1363 and IL-1403 synthesize only Opp and Opt, respectively . The Opt system displays significant differences from the lactococcal Opp system, which made Opt much more closely related to the oligopeptide transporters of streptococci than to the lactococcal Opp system: (i) genetic organization, (ii) peptide uptake specificity, and (iii) presence of two oligopeptide-binding proteins, OptS and OptA . The fact that only OptA is required for nutrition calls into question the function of the second oligopeptide binding protein (Opts) . Sequence analysis of oligopeptide-binding proteins from different bacteria prompted us to propose a classification of these proteins in three distinct groups, differentiated by the presence (or not) of precisely located extensions .
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