Clarithromycin accelerated the resolution of VAP and weaning from mechanical ventilation in surviving patients and delayed death in those who died of sepsis. The mortality rate at day 28 was not altered. Results are encouraging and render new perspectives on the management of sepsis and VAP.

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The maximum parsimony method was used to reconstruct the genealogical history of the family of intracellular calcium-binding proteins represented by six major present-day lineages, three of which--calcium dependent modulator protein, heart and skeletal muscle troponin Cs, and alkali light chains of myosin--were found to share a closer kinship with one another than with the other lineages. Similarly, parvalbumins and regulatory light chains of myosin were depicted as more closely related, whereas the branch of intestinal calcium-binding protein proved to have the most distant separation. The computer-generated amino acid sequence for the common ancestor of these six lineages described a four domain protein in which each domain of approximately 40 amino acid residues had a mid-region. 12 residue segment that bound calcium and had properties most resembling those of the calcium dependent modulator protein. It could then be deduced that parvalbumins evolved by deletion of domain I, inactivation of calcium-binding properties in domain II, and acquisition of increased affinity for Ca++ and Mg++ in domains III and IV. Regulatory light chains of myosin lost the cation binding property from three domains, retaining it in I, whereas alkali light chains of myosin lost this ability from each of the four domains. In skeletal muscle troponin C all domains retained their calcium-binding activity; however, like parvalbumins, domains III and IV acquired high affinity properties. Cardiac troponin C lost its binding activity from domain I but otherwise resembled the skeletal muscle form. Finally, intestinal calcium-binding protein evolved by deletion of domains III and IV. Positive selection could be implicated in these evolutionary changes in that the rate of fixation of mutations substantially increased in the mid portions of those domains which were loosing calcium-binding activity. Likewise, when the cation binding sites were changing from low to high affinity, an accelerated rate of fixed mutations was observed. Once this new functional parameter was selected these regions showed a remarkable conservatism, as did those binding sites which were maintaining the lower affinity. Moreover even in sequence regions not directly involved in cation binding, the lineage of troponin C because very conservative over the past 300 million years, perhaps becuase of the necessity for maintaining specific interfaces in order for the molecule to interact with troponin I and T in a functional thin myofilament. A similar phenomenon was observed in domain II of the regulatory light chains of the myosin lineage suggesting a possible binding site with the heavy chain of myosin.

Purpose: Anti-virulence strategies have not been evaluated for the prevention of bacterial infections. Prolonged colonization of intubated patients with Pseudomonas aeruginosa isolates producing high-levels of the quorum sensing (QS)-regulated virulence factor rhamnolipids has been associated with ventilator-associated pneumonia (VAP). In this pathogen, azithromycin reduces QSregulated virulence. We aimed to assess whether azithromycin could prevent VAP in patients colonized by rhamnolipids producing isolates. Methods: In a randomized, doubleblind, multicenter trial, intubated colonized patients received either 300 mg/day azithromycin or placebo. Primary endpoint was the occurrence of P. aeruginosa VAP. We further identified those patients persistently colonized by isolates producing highlevels of rhamnolipids and therefore at the highest risk to develop VAP linked to this QS-dependent virulence factor. Results: Ninety-two patients were enrolled; 43 azithromycin-treated and 42 placebo patients were eligible for the per-protocol analysis. In the per-protocol population, the occurrence of P. aeruginosa VAP was reduced in the azithromycin group but without reaching statistical significance (4.7 vs. 14.3 % VAP, p = 0.156). QS-dependent virulence of colonizing isolates was similarly low in both study groups, and only five patients in each arm were persistently colonized by high-level rhamnolipids producing isolates. In this high-risk subgroup, the incidence of VAP was reduced fivefold in azithromycin versus placebo patients (1/5 vs. 5/5 VAP, p = 0.048). Conclusions: There was a trend towards reduced incidence of VAP in colonized azithromycin-treated patients. In addition, azithromycin significantly prevented VAP in those patients at high risk of rhamnolipiddependent VAP, suggesting that virulence inhibition is a promising antimicrobial strategy.

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1. The three main components of the 1.5-2s ultracentrifugal peak of carp myogen (white muscle) have been isolated by ammonium sulphate fractionation and zone electrophoresis, and crystallized. 2. The molecular weights of these three proteins were determined by sedimentation and diffusion, by the Archibald method and by amino acid analysis, and found to lie between 9000 and 13000. 3. Their complete amino acid compositions were determined by column chromatography and by their ultraviolet spectra. Both methods revealed abnormal compositions, including the absence of tryptophan and methionine and the presence of large amounts of phenylalanine. At most 1 residue each of tyrosine, cysteine, proline, arginine and histidine was found/molecule. 4. The specific viscosity of component 3 was lower than that of other small globular proteins described so far, a fact that suggests that these proteins approximate more closely to the ideal case of the spherical protein molecule. Also, the presence of a single residue of several amino acids, the absence of disulphide bonds, and the apparent reversibility of denaturation by urea of component 3 suggest that the study of these molecules could provide new information on the structure of proteins.

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