IS431, a staphylococcal insertion sequence-like element related to IS26 from Proteus vulgaris.

We present the nucleotide sequence of IS431, a new staphylococcal insertion sequence-like element flanking the mercury-resistance determinant of pI524 and associated with the methicillin-resistance determinant. IS431 left is 800 bp long and has a perfect terminal inverted repeat (IR) of 22 bp; IS431 right is 786 bp long and has a terminal IR homologous to the IR of IS431 left except that the terminal 8 bp are absent. Both IRs share a 10-bp homology with the IR of IS26 from Proteus vulgaris. No directly repeated sequences were detected immediately adjacent to the IRs. An open reading frame (ORF) of 675 bp spans most of the IS431 sequence. Its deduced amino acid (aa) sequence shows 40% homology to the 234-aa-long putative transposase coded by ORFI of IS26.

In vitro activity of ciprofloxacin against gram-positive bacteria. An overview.

A review of European data published before July 1986 and data from the authors' laboratory showed very similar activity of ciprofloxacin against gram-positive bacteria in all studies. Mean minimal inhibitory concentrations against 50 percent (MIC50) and 90 percent (MIC90) of staphylococcal strains were 0.32 and 0.59 mg/liter, respectively. The drug was equally active against staphylococcal strains resistant and susceptible to methicillin and/or gentamicin. The range of MIC50 and MIC90 values for Streptococcus faecalis and Streptococcus faecium was between 0.25 and 1 mg/liter and 1 to 8 mg/liter, respectively. Ciprofloxacin inhibited group A and group B streptococci at concentrations of 0.5 to 2 mg/liter. Group C and G strains were less susceptible (MICs, 2 to 16 mg/liter). Penicillin-resistant and penicillin-susceptible pneumococci and the Viridans streptococci were inhibited by 0.5 to 4 mg/liter. MIC50 and MIC90 values of ciprofloxacin against Corynebacterium, including group JK, and anaerobic gram-positive cocci were 1 and 8 mg/liter, respectively, and against anaerobic gram-positive rods, were 2 and 16 mg/liter, respectively. Listeria monocytogenes strains were inhibited by 0.12 to 2 mg/liter. Ciprofloxacin showed bactericidal activity against staphylococci and streptococci in minimal bactericidal concentration tests and in killing kinetic studies. The in vitro activity of the drug was influenced neither by the method employed, by the medium used, by the pH of the medium, nor by the size of the inoculum. Resistance to ciprofloxacin developed in staphylococci in a step-wise manner. First-step mutants usually showed a fourfold to eightfold decrease in susceptibility. Contrary to the situation in gram-negative organisms, resistant mutants did not show reduced growth.

Prospective evaluation of a semiquantitative dip slide method compared with quantitative bacterial cultures of BAL fluid.

BACKGROUND: Quantitative bacteriologic workup of BAL fluid (BALF) has evolved as a sensitive and specific technique for the diagnosis of bacterial pneumonia. Conventional quantitative cultures are expensive, time-consuming, and often unavailable on a 24-h basis. Therefore, we evaluated a dip slide method for the semiquantitative measurement of bacterial cultures in BALF specimens and compared the results with those from conventional quantitative cultures. METHODS: Fifty BALF specimens from 45 patients with suspected pulmonary infection were examined prospectively with both methods. We compared the microbiologic results of conventional quantitative cultures with those of the dip slide method that is commercially available for blood cultures. Cost-effectiveness analysis of both methods was performed. RESULTS: In 37 BALF specimens, 64 bacterial strains were detected with both techniques. The dip slide method and conventional cultures showed a high correlation with respect to the colony counts of the individual organisms per milliliter BALF (r=0.935; p= 0.0001) and the sum of colony counts in individual patients (r=0.947; p=0.0001). Although five strains were not detected by the dip slide technique, the diagnostic accuracy was not influenced. In 13 BALF samples, there was no growth of bacteria with both techniques. While the diagnostic yield of both methods was similar, the dip slide technique was 44 to 66% less expensive than conventional cultures. CONCLUSIONS: The examination of BALF with a clip slide method is highly comparable to conventional quantitative culture techniques, less expensive, and can be used independently of a specialized microbiology laboratory on a 24-h basis.

Impact of sar and agr on methicillin resistance in Staphylococcus aureus.

The global regulators agr and sar control expression of cell wall and extracellular proteins. Inactivation of either sar and/or agr in a typical heterogeneously methicillin-resistant Staphylococcus aureus resulted in a small but reproducible decrease in the number of cells in the subpopulation expressing high methicillin resistance. The amount of low affinity penicillin-binding protein PBP2', the prerequisite for methicillin resistance, was apparently not affected, however, a reduction in PBP1 and PBP3 production was observed, suggesting that these resident PBPs of the cells might be involved somehow together with PBP2' in high level methicillin resistance.

Genetic and molecular characterisation of resistance determinants in methicillin-resistant Staphylococcus-aureus.

A genetic analysis of resistance to antibiotics in methicillin-resistant Staphylococcus aureus was performed. Demonstration of plasmid-specific DNA either in transductants that had received antibiotic-resistance markers from multiply-resistant strains, or in segregants of methicillin-resistant strains that had lost unstable determinants except the one under study, indicated that markers of resistance to penicillin, chloramphenicol and neomycin are present on separate, mutually compatible plasmids. Absence of covalently closed circular DNA was demonstrated in transductants that were resistant to methicillin, tetracycline, erythromycin and streptomycin, as well as in segregants that had lost the penicillinase, chloramphenicol and neomycin plasmid, but were still resistant to methicillin, tetracycline, erythromycin, streptomycin and the sulphonamides. Analysis of plasmid DNA either in a 5-20% neutral sucrose gradient or by electron microscopy revealed the presence of three readily distinguishable plasmids. The molecular weights of these plasmids were estimated by comparing the sedimentation rate constants with those of known reference plasmids and by contour-length measurements. The molecular weight of the penicillinase plasmid was estimated to be 20 X 10(6) daltons, that of the chloramphenicol plasmid 3 X 10(6) daltons and that of the plasmid carrying the neomycin resistance marker 37 X 10(6) daltons.

Genetics of multiply-resistant Staphylococcus aureus.

A substantial portion of recently isolated, multiply-resistant Staphylococcus aureus was shown to carry R-determinants in plasmids. Some of these plasmids were small (3 Mdal) and carried only one R-marker. Others were 18-36 Mdal in size and carried two or more R-determinants. Several of the larger plasmids could be transferred by a conjugation-like process. The location of R-markers on transposable DNA sequences also was observed. Transposition as well as stable integration of R-plasmids into the chromosome can explain the frequent observation of chromosomal location of resistance. Chromosomal resistance might be an advantage for an organism frequently exposed to antibiotics. Molecular evidence suggests that methicillin resistance resides on additional chromosomal DNA. The organization of staphylococcal genomes as well as efficient transfer processes explain the genetic versatility of Staph. aureus, which has resulted in the development of multiply-resistant strains.

[Enzymatic assays of aminoglycoside antibiotics]. / Enzymatische Wertbestimmung von Aminoglykosid-Antibiotika.

Adenylylating or acetylating enzymes, which are produced by grampositive and gramnegative bacteria resistant to aminoglycoside antibiotics, are useful tools in determining concentrations of aminoglycosides in patients' body fluids. Due to their accuracy, stability, versatility, fastness and inexpensiveness, enzymatic assays are the tests of choice for the routine determination of aminoglycoside concentrations.

A comparative structural study on the steroids [1,2,5]oxadiazolo[3', 4':3,4]-5 alpha-pregn-16-en-20-one oxime (HS998), [1,2,5]oxadiazolo[3, 4':3,4]-5 beta-pregn-16-en-20-one (HS973) by NMR, molecular modeling, and X-ray investigations.

The molecular structure of the steroids [1,2,5]oxadiazolo[3', 4':3,4]-5 alpha-pregn-16-en-20-one oxime, [1,2,5]oxadiazolo[3',4':3, 4':3,4]-5 alpha-pregn-16-en-20-one and [1,2,5]oxadiazole]3',4':3,4]-5 beta-pregn-16-en-20-one has been determined. The proton-proton distances in the solid state from previous crystallographic studies are compared with the corresponding distances from novel and previous solution NMR as well as from novel in vacuo modeling studies.

Safety aspects of enterococci from the medical point of view.

Enterococci occur in a remarkable array of environments. They can be found in soil, food, and water, and make up a significant portion of the normal gut flora of humans (10(5)-10(7)/g of stool) and animals. As other bacteria of the gut flora, enterococci can also cause infectious diseases. Most clinical isolates are Enterocococus faecalis, which account for 80-90% of clinical strains. Enterocococus faecium accounts for 5-10% of such isolates. Typical enterococcal infections occur in hospitalised patients with underlying conditions representing a wide spectrum of severity of illness and immune modulation. Enterococci today rank second to third in frequency among bacteria isolated from hospitalised patients. They are isolated from urinary tract infections, intra-abdominal and pelvic infections, bacteremias, wound and tissue infections, and endocarditis--often as part of a polymicrobial flora. Surprisingly, little is known about the factors that contribute to the ability of enterococci to cause infections. Many strains of E. faecalis produce a cytolysin (haemolysin) exhibiting tissue-damaging capacity. Further extracellular products often observed in clinical isolates are a proteinase (gelatinase), hyaluronidase, and extracellular superoxide. Furthermore, many of the clinical isolates possess the aggregation substance on the surface and an extracellular surface protein, both contributing to the adherence to eucaryotic cells. Some strains of E. faecalis, and many E. faecium strains are resistant to multiple antimicrobials. The ultimate role of all these factors in enterococcal pathogenicity remains to be determined. It was previously thought that enterococcal infections were endogenously acquired from the patient's own gut flora. A rather new concept that has emerged is that enterococcal disease is a two-stage process. There is an initial colonisation of the gastrointestinal tract by enterococcal strains possessing virulence traits and/or antibiotic resistance. Subsequently, this population spreads, often facilitated by antibiotic elimination of competitors. For a selected number of patients, there is subsequent tissue invasion from the gastrointestinal tract reservoir. From this concept, it can be deduced that enterococcal strains without virulence traits and antibiotic resistances exogenously transferred into the human gut via food products or probiotics will not represent any risk for immunocompetent individuals. In very severely immunocompromised patients, however, a risk for enterococcal disease by such strains cannot completely be excluded.

Evolution of resistance in microorganisms of human origin.

Resistance to antimicrobials in bacteria results from either evolution of "new" DNA or from variation in existing DNA. Evidence suggests that new DNA did not originate since the use of antibiotics in medicine, but evolved long ago in soil bacteria. This evidence is based on functional and structural homologies of resistance proteins in human pathogens, and resistance proteins or physiological proteins of soil bacteria. Variation in existing DNA has been shown to comprise variations in structural or regulatory genes of the normal chromosome or mutations in already existing plasmid-mediated resistance genes modifying the resistance phenotype. The success of R-determinants in human pathogens was due to their horizontal spread by transformation, transduction and conjugation. Furthermore, transposition has enabled bacteria to efficiently distribute R-determinants between independent DNA-molecules. Since the genetic processes involved in the development of resistance are rare events, the selective pressure exerted by antibiotics has significantly contributed to the overall evolutionary picture. With few exceptions, experimental data about the role of antibiotic usage outside human medicine with respect to the resistance problem in human pathogens are missing. Epidemiological data about the occurrence of resistance in human pathogens seem to indicate that the major contributing factor to the problem we face today was the extensive use of antibiotics in medicine itself.

Purification and characterization of an aminoglycoside inactivating enzyme from Staphylococcus epidermidis FK109 that nucleotidylates the 4'- and 4''-hydroxyl groups of the aminoglycoside antibiotics.

The resistance to aminoglycoside antibiotics in Staphylococcus epidermidis FK109, is mediated by an enzyme that catalyzes transfer of the nucleotide monophosphate moiety from the nucleotide triphosphates, either to the 4'-hydroxyl group, that is in the equatorial plane of the aminoglycoside molecule. The enzyme, modifying the two sites, appears as a single and homogeneous entity in affinity chromatography, in chromatography on DEAE-Sepharose CL-6B, in isoelectric focusing and in gel-filtration. It requires divalent cations, notably Mg++, and dithiothreitol for optimal adenylylation. It has a molecular weight of 46,770 and an isoelectric point of 5.0. The ability of the enzyme ANT (4', 4'') to modify the two hydroxyl groups of aminoglycoside molecules, enables it to have a spectrum of substrates that surpasses, in range, the substrate spectrum of all the aminoglycoside-modifying enzymes which have been characterized hitherto.

Apparatus for extruding wires of soft metals under vacuum or inert atmospheres.

A bakeable apparatus is described that was constructed for extruding wires from small ingots of soft metals like the alkalis, the alkaline earths, and the divalent rare earths, while at the same time safeguarding the purity of the sample material. The unit is completely enclosed yet small enough to be hand transferred into and out of an inert atmosphere glove box where load/unload operations are performed. Extrusions are effected with the apparatus outside the glove box but with the sample material under the captured atmosphere of the box or under a vacuum. Given an external heating source it is possible to carry out extrusions at temperatures above room temperature as well as in situ recrystallizations of extruded materials.

In vitro activity of oral cephalosporin BAY v 3522 compared with other oral cephalosporins.

Minimal inhibitory concentrations (MICs) of the oral cephalosporin BAY v 3522, and of cephprozyl, cefaclor, cefixime, cefuroxime, cefetamet, cefpodoxime and cefotaxime were determined against Gram-positive and Gram-negative clinical isolates with the NCCLS agar dilution procedures. BAY was the most active drug against Gram-positive organisms. MICs ranged from 0.01 mg/l against group A streptococci to 16 mg/l against S. faecium. Although mean MICs of BAY against methicillin-resistant S. aureus and S. epidermidis were between 0.9-1.8 mg/l, respectively, such strains showed typical heteroresistance in population studies. In addition, the biochemical correlate of methicillin-resistance, the PBP-2', showed similar low affinity to BAY as methicillin. beta-lactamase-producing H. influenzae and B. catarrhalis were inhibited by 2-8 and 0.25-2 mg/l, respectively, whereas non-producers were inhibited by 0.25-2 and 0.12-1 mg/l of the drug. The activity of BAY against enterobacteriaceae was rather low. Ampicillin-susceptible E. coli strains were inhibited by 2-8 and resistant strains by 8-32 mg/l. The mean MIC against cephalothin-susceptible K. pneumoniae strains was 2.8, and that against resistant strains 27.4 mg/l. MICs against beta-lactamase-producing enterobacteriaceae determined in broth dilution were 4-8 times higher than those determined in agar dilution. Bactericidal activity was measured in killing-curve experiments at 4 times the MIC. BAY killed equally well as standard control drugs.

Infection in the upper body: hand and burn-wound microbiology and considerations for antimicrobial therapy.

Gram-positive bacteria are the predominant organisms in hand infection and in burn wounds of the upper extremities. In a recent study of isolates from patients who were treated at our institution, Staphylococcus aureus and beta-hemolytic Streptococcus group A organisms were the most common organisms in infection of the hand; they were found in 36.3% and 14.4% of cases, respectively. The most common organisms in burn wounds were Enterococcus species, S. aureus, and Escherichia coli, which were found in 21.2%, 20.5%, and 16.7% of patients, respectively. Between 1969 and 1989, the prevalence of Pseudomonas species in burns decreased markedly, whereas that of S. aureus remained relatively stable and that of Enterococcus increased substantially. Over this period, both enterococci and coagulase-negative staphylococci emerged as troublesome pathogens in patients with burns. Methicillin-resistant S. aureus, which was first seen in our institution in 1981, continues to be found in a small proportion of patients. We have achieved successful results in certain surgical settings with the use of gentamicin-dispersing polymethyl-methacrylate beads to provide sufficient antimicrobial concentrations in poorly vascularized or avascular tissue. Additional topical antimicrobials that are potent against gram-positive bacteria are needed.

[Ganglion cyst of the cervical spine causing radiculopathy]. / Le kyste ganglionnaire de la colonne cervicale responsable d'une radiculopathie.

A case of ganglion cyst of the cervical spine causing radiculopathy is presented. This epidural mass is rare at the cervical level. Computed tomography suggests the diagnosis by the postero-lateral position of the mass close to facet joint. The trilobed configuration and the tissue characterisation of the cyst are well documented on MRI.

[Bacterial sensitivity to chemotherapeutic agents (Zurich, 1990)]. / Die Empfindlichkeit von Bakterien gegen Chemotherapeutika (Zürich, 1990).

This paper describes the incidence of susceptibility of gram-negative and positive bacteria towards antibacterial agents. The data are based on all susceptibility tests performed at the Department of medical microbiology of the University of Zurich. The evaluation of the results from 1975 to 1990 shows that susceptibilities against the antimicrobial agents tested have not changed markedly in this period. These tables may be a help for the physician in his decision for a "calculated chemotherapy" of bacterial infections.

[In-vitro activity of cefuroxime-axetil against pathogens of bacterial infections of the respiratory tract]. / In-vitro-Aktivität von Cefuroxim-Axetil gegen Erreger bakterieller Infektionen des Respirationstrakts.

Cefuroxime-Axetil is an ester of cefuroxime, which can be used as an oral antimicrobial agent. The prodrug is hydrolysed by esterases of the gut mucosa, setting free the active cefuroxime. This second generation cephalosporin is well known since more than a decade as a cephalosporin possessing high stability against gram positive and gram negative beta-lactamases. Due to its pharmacokinetic properties and its wide spectrum of activity, cefuroxime-axetil was recommended to be used as an oral agent in bacterial infections of the respiratory tract. This paper describes the in vitro activity of cefuroxime against respiratory tract pathogens and compares it with that of ampicillin and amoxicillin, amoxicillin/clavulanic acid, cefaclor, chloramphenicol, tetracycline, erythromycin and trimethoprim/sulphamethoxazole. Cefuroxime had good activity against beta-lactamase-producing and beta-lactamase-negative H. influenzae and M. catarrhalis. As expected, the aminopenicillins showed reduced activity against beta-lactamase-producing strains of these organisms. Amoxicillin/clavulanic acid and, to a lesser degree, cefaclor also showed good activity against these bacteria. Cefuroxime was highly active against penicillin-susceptible pneumococci, but showed impaired activity against penicillin-resistant strains. Therefore, this drug should not be used in infections caused by penicillin-resistant pneumococci. Resistance to penicillin in pneumococci isolated in Switzerland is not a problem at the present time. For many years, such strains were isolated only sporadically. Cefuroxime showed also a high in vitro activity against streptococci of serogroups A, B, C and G, which are encountered as respiratory tract pathogens. Against beta-lactamase-positive and -negative staphylococci, cefuroxime showed good activity. The drug was inactive, however, against methicillin-resistant strains.

[Bacterial susceptibility to chemotherapeutic agents (Zurich, 1987)]. / Die Empfindlichkeit von Bakterien gegen Chemotherapeutika (Zürich, 1987).

This paper describes the frequency of susceptibility of Gram-positive and Gram-negative bacteria against antibacterial agents. The data are based on all susceptibility tests performed at the Department of Medical Microbiology of the University of Zurich. The evaluation of the results from 1975 to 1987 shows that susceptibility against standard antimicrobial agents has not decreased in this period. These tables may be a valuable help for the physician in his decision for a "calculated chemotherapy".