Evaluation of the treatment of vancomycin-resistant enterococcal urinary tract infections in a large academic medical center.

BACKGROUND: Vancomycin-resistant enterococci (VRE) are a common cause of nosocomial urinary tract infections (UTIs) among hospitalized patients. Clinicians need to differentiate between VRE-associated urinary colonization, asymptomatic bacteriuria, and UTIs to determine the need for treatment and length of therapy. OBJECTIVE: To characterize the diagnosis and management of VRE from urinary sources, including compliance with institutional treatment guidelines, and identify risk factors associated with clinical failure. METHODS: We performed a retrospective, single-center, cohort study among patients with VRE-positive cultures from urinary sources over a 3-year study period (July 2008-September 2011). Descriptive statistics were used to evaluate demographics, diagnostics, guideline compliance, pharmacotherapy, and outcomes. Risk factors associated with clinical failure were identified by multivariate logistic regression analysis. RESULTS: Two hundred sixty-nine distinct episodes of VRE met inclusion criteria among 252 patients. Forty-seven percent and 77% of episodes occurred in patients admitted to an intensive care unit and hospitalized for 7 or more days, respectively. Fifty-eight percent of the episodes were classified as asymptomatic bacteriuria or colonization. Compliance with institutional treatment guidelines for the appropriate drug, dose, and duration occurred in approximately 70% of the cases. Among noncompliant cases (n = 83), 48 (58%) were overtreated, and 35 (42%) were undertreated. Clinical failure among all cases was common, including mortality (17.1%). Factors independently associated with clinical failure determined on multivariate analysis included weight 100 kg or more (OR 5.30; 95% CI 1.42-12.21; p = 0.014), renal disease (OR 2.57; 95% CI 1.02-6.47; p = 0.048), indwelling catheter (OR 4.62; 95% CI 1.05-18.24; p = 0.046), and VRE bloodstream infection (OR 15.71; 95% CI 2.9-128.7; p < 0 .001). CONCLUSIONS: Improved education is needed to minimize cases of overtreatment and undertreatment of VRE-associated UTIs and decrease inappropriate drug-related costs and clinical failure rates. Risk factors for clinical failure can be used to risk stratify VRE-associated UTIs and further guide treatment decisions.

Alp7R regulates expression of the actin-like protein Alp7A in Bacillus subtilis.

Alp7A is a bacterial actin from Bacillus subtilis plasmid pLS20 that functions in plasmid segregation. Alp7A's function requires that it assemble into filaments that treadmill and exhibit dynamic instability. These dynamic properties require the two other components of the alp7A operon, the downstream alp7R gene and the upstream alp7C sequence, as does the ability of Alp7A to form filaments at its physiological concentration in the cell. Here, we show that these two other components of the operon also determine the amount of Alp7A that is produced in the cell. The deletion of alp7R leads to overproduction of Alp7A, which assembles into large, amorphous, static filaments that disrupt chromosome segregation and cell division. The product of the alp7R gene is a DNA-binding protein that represses transcription of the alp7A operon. Purified Alp7R protein binds specifically to alp7C, which contains two σ(A) promoters embedded within a series of near-repeats of a 10-mer. Alp7R also shows the typical nonspecific binding activity of a DNA-binding protein: Alp7R-GFP (green fluorescent protein) associates with the chromosomes of cells that lack alp7C. When Alp7A-GFP is produced in B. subtilis along with untagged Alp7R, Alp7A-GFP also colocalizes with the chromosome, indicating that Alp7R associates with Alp7A. Hence Alp7R, determines both the activity and the cellular concentration of Alp7A, and it can associate with Alp7A even if it is not bound to alp7C.

Phylogenetic analysis identifies many uncharacterized actin-like proteins (Alps) in bacteria: regulated polymerization, dynamic instability and treadmilling in Alp7A.

Actin, one of the most abundant proteins in the eukaryotic cell, also has an abundance of relatives in the eukaryotic proteome. To date though, only five families of actins have been characterized in bacteria. We have conducted a phylogenetic search and uncovered more than 35 highly divergent families of actin-like proteins (Alps) in bacteria. Their genes are found primarily on phage genomes, on plasmids and on integrating conjugative elements, and are likely to be involved in a variety of functions. We characterize three Alps and find that all form filaments in the cell. The filaments of Alp7A, a plasmid partitioning protein and one of the most divergent of the Alps, display dynamic instability and also treadmill. Alp7A requires other elements from the plasmid to assemble into dynamic polymers in the cell. Our findings suggest that most if not all of the Alps are indeed actin relatives, and that actin is very well represented in bacteria.

Treadmilling of a prokaryotic tubulin-like protein, TubZ, required for plasmid stability in Bacillus thuringiensis.

Prokaryotes rely on a distant tubulin homolog, FtsZ, for assembling the cytokinetic ring essential for cell division, but are otherwise generally thought to lack tubulin-like polymers that participate in processes such as DNA segregation. Here we characterize a protein (TubZ) from the Bacillus thuringiensis virulence plasmid pBtoxis, which is a member of the tubulin/FtsZ GTPase superfamily but is only distantly related to both FtsZ and tubulin. TubZ assembles dynamic, linear polymers that exhibit directional polymerization with plus and minus ends, movement by treadmilling, and a critical concentration for assembly. A point mutation (D269A) that alters a highly conserved catalytic residue within the T7 loop completely eliminates treadmilling and allows the formation of stable polymers at a much lower protein concentration than the wild-type protein. When expressed in trans, TubZ(D269A) coassembles with wild-type TubZ and significantly reduces the stability of pBtoxis, demonstrating a direct correlation between TubZ dynamics and plasmid maintenance. The tubZ gene is in an operon with tubR, which encodes a putative DNA-binding protein that regulates TubZ levels. Our results suggest that TubZ is representative of a novel class of prokaryotic cytoskeletal proteins important for plasmid stability that diverged long ago from the ancient tubulin/FtsZ ancestor.