Molecular cloning and expression of a novel Staphylococcus aureus antigen.

A novel gene encoding an antigen from Staphylococcus aureus was isolated from an expression library by screening with antisera from patients with deep Staphylococcus aureus infections. In one positive clone an open reading frame, named ORF-2, was identified. Recombinant ORF-2 protein reacted with human immune serum. ORF-2 was shown to be present in other Staphylococcus aureus strains, but not in related species.

The effect of antistaphylococcal agents used alone and in combinations on the survival of Staphylococcus aureus ingested by human polymorphonuclear leukocytes.

The intracellular activity of a number of drugs used alone and in combinations against Staphylococcus aureus was investigated using an experimental design which imitates the clinical situation and differs from other published methods. Staphylococci were phagocytosed by human polymorphonuclear leukocytes and, after differential centrifugation and washing, the granulocytes were incubated in 90% pooled human serum with clinically relevant drug concentrations. When exposed to antibiotics, more than 40-50% of the bacteria were located intracellularly. Fusidic acid (100 mg/l), erythromycin (20 mg/l), and clindamycin (20 mg/l) all had a bacteriostatic effect during the first 6 h of incubation, whereas rifampicin (1 and 5 mg/l), vancomycin (5 and 20 mg/l), and ciprofloxacin (2 mg/l) all acted bactericidally with decreases in viable counts between 1.3-1.9 log10. The greatest bactericidal effect was achieved with tobramycin (10 mg/l), which produced more than a 4 log10 decrease in viable counts at 6 h. Combinations of fusidic acid with other antibiotics all resulted in killing kinetics different from those achieved with the drugs used individually. The bactericidal effect of ciprofloxacin and dicloxacillin during the first 6 h was abolished when these drugs were combined with fusidic acid. However, at 24 h no significant difference was found between the effect of dicloxacillin alone versus the combination dicloxacillin and fusidic acid. The combination of fusidic acid and rifampicin resulted in a killing identical to that achieved with rifampicin used alone during the first 6 h, but at 24 h the killing by the combination was significantly greater. The bactericidal effect of the combination dicloxacillin (20 mg/l) and tobramycin (10 mg/l) equalled that obtained with tobramycin (10 mg/l) used alone. Rifampicin (5 mg/l) antagonized the bactericidal effect of ciprofloxacin (2 mg/l) during the first 6 h of incubation but at 24 h the combination acted synergistically. The results obtained are partly in agreement and partly in conflict with previous results.

Killing curve activity of ciprofloxacin is comparable to synergistic effect of beta-lactam-tobramycin combinations against Haemophilus species endocarditis strains.

Nine Haemophilus species strains, all beta-lactamase negative, isolated from patients with endocarditis were tested in killing curve experiments. Antibiotics used were penicillin, amoxicillin, aztreonam alone and in combination with tobramycin, as well as ciprofloxacin alone. Synergism between beta-lactams and tobramycin with reduction of colony counts to zero was seen after 24 h for H. influenzae, H. parainfluenzae and H. segnis strains. Ciprofloxacin was as effective as beta-lactam-tobramycin combinations. The H. aphrophilus strain was not killed as effectively as other strains by any of the antibiotics.

Microbiologically verified diagnosis of infectious spondylitis using CT-guided fine needle biopsy.

Using a computed tomography (CT)-guided technique we have been able to obtain fine needle spine biopsies directly from an affected vertebra or disk plate in 14 patients suspected of infectious spondylitis. The bioptic material was cultivated immediately and incubated for 14 days. Cultures from eight patients were positive. No single microbiological agent was predominant though coagulase-negative staphylococci were frequent. In no case were mycobacteria found. Bioptic material from six patients did not give rise to growth of microorganisms. We were able to successfully treat the eight patients with a culture-positive biopsy. We think that biopsies are crucial for establishing a microbiological diagnosis. The whole procedure takes less than one hour; it is performed under local anaesthesia and is thus not very stressful for the patient: The success rate for obtaining a positive spine biopsy was 57%.

Serological diagnosis of experimental Enterococcus faecalis endocarditis.

A modified rat model of endocarditis with catheterization for 2 days was established in female Lewis rats using different inocula of Enterococcus faecalis (strain no. EF 19) in order to measure IgG antibodies in serum during the course of infection. Increasing the inocula intravenously resulted in an increase in the CFU/g vegetation and the CFU/g spleen, the ID50 being about 10 CFU/ml and the ID90 about 1x10(2) CFU/ml. The lowest bacterial inoculum infecting 100% of the rats was 3x10(3) CFU/ml, and for further investigations we used this inoculum size. Rats were sacrificed on day 2, 5, 7, 9, 11 and 28 after infection. The CFU/g vegetation and the CFU/g spleen increased until day 7 and then decreased. Serum samples were collected from 129 rats at different times after challenge. Three different ELISA systems were established to measure the IgG antibody responses: E. faecalis sonicate ELISA (a pool of four sonicates of strain no. EF 10, EF 11, EF 19 and EF 48), E. faecalis whole cell ELISA (strain no. EF 19) and E. faecalis purified cell wall ELISA (strain no. EF 19). An IgG antibody response was detected already on day 2, and except for a minor decrease on day 6/7 the antibody response continued to increase until day 14 (whole cell ELISA and sonicate ELISA) and day 21 (purified cell wall ELISA) when a plateau was reached. Significant increases in IgG antibody responses (p<0.05) were found between groups of rats from days 0-2, 2-8/9 and 8/9-14 in the E. faecalis whole cell and sonicate ELISAs and from days 0-2, 2-10/11 and 10/11-21 in the E. faecalis purified cell wall ELISA. In conclusion, we established a model of endocarditis in rats with catheterization for 2 days and were able to demonstrate an increase in IgG antibodies during the course of infection.

Opportunistic screening for Chlamydia trachomatis cervicitis: the value of cytobrush specimens for detection by PCR compared with cell culture.

UNLABELLED: Endocervical sampling for microbiological and pathological screening is laborious and expensive due to different sampling devices and techniques. The purpose of this study was to examine if the routine procedure could be simplified by using a cytobrush for concurrent cytology and sampling for Chlamydia trachomatis detection using the PCR method or cell culture. As a sampling device control we used a conventional rayon swab. RESULTS: Culture: Out of 873 paired endocervical specimens, C. trachomatis was isolated in 68 swab specimens and in 65 cytobrush specimens (overall detection rate 8.4%). The cytobrush proved less suitable than the swab for the isolation of C. trachomatis as 31.5% of the cytobrush samples showed cytotoxicity to the cultured cells vs 0.9% of the swab samples. PCR: In a random sample of 427 paired endocervical specimens, C. trachomatis was detected in 45 pairs without any difference between the two sampling devices. The sensitivity of PCR was 93.8% vs 89.6% and 87.5% in cultured swab and cultured cytobrush specimens, respectively. The cytobrush can therefore be recommended as a cervical sampling device if a PCR assay is used for the detection of C. trachomatis, but not if the cell culture method is used, due to high cytotoxicity. Furthermore, the same cytobrush may be used for cervical cytological sampling and thereafter placed in transport medium for subsequent C. trachomatis detection if the PCR technique is used.

New developments in the treatment of infective endocarditis infective cardiovasculitis.

The natural history of infective endocarditis has undergone remarkable changes over the past 100 years as regards both the demographic characteristics of the disease and changes in the incidence of the so-called diagnostic signs. Alongside these changes and the development of new and better diagnostic tools and criteria, we are also facing new problems with the precise definition of cardiovascular infections and calculation of the incidence of the disease. Nosocomial endocarditis presents an emerging problem of diagnosis and treatment after heart valve surgery, with pace-maker catheters, defibrillators and a very large variety of foreign materials used in connection with heart valve surgery. New technological progress including new types of prosthetic valves and use of homografts or the Ross operation will give a greater possibility of choosing the best solution in a particular case. Antimicrobial chemotherapy is mainly based on our understanding of the pathophysiology of the disease and efficacy of the antibiotics achieved in an experimental animal model of endocarditis. Important recommendations of single or combined drug therapy or the dosing regimens of antibiotics are still an expression of expert opinion not always supported by experimental or clinical proof. A typical example is the recommendation of two divided doses of gentamicin for treatment of streptococcal endocarditis. Nevertheless, it is the author's opinion that the development of uncomplicated, easy to handle diagnostic and treatment regimens are justified in order to achieve better compliance with these recommendations.

[Acute injection-induced injuries in drug addicts. A review of 146 consecutive admissions]. / Akutte injektionsskader hos narkomaner. En gennemgang af 146 konsekutive indloeggelser.

Admissions of drug abusers with acute soft tissue lesions at the injection site increased in number over the period 1985-1989. There were 146 admissions in 90 patients. The diagnoses were: 58 superficial abscesses, 27 deep abscesses, 57 cellulitis, one tenosynovitis, one purulent arthritis, one not further categorised abscess and one case of arterial spasm. The commonest location was the groin (25%). Serious complications occurred in 17 cases, including four femoral amputations due to arterial lesions. Ultrasonic examination is recommended before surgery if central vessels can be involved, especially in the groin. Sufficient microbiological examination was performed in 78 cases. There was a predominance of polybacterial infections (53% polybacterial, 38% monobacterial, 9% sterile). The most common bacteria were Streptococcus spp. with a preponderance of oropharyngeal bacteria, Staphylococcus aureus, and anaerobes, especially Bacteroides spp. Typical intestinal bacteria were rare. We recommend that aerobic and anaerobic culturing with susceptibility testing always be carried out, that primary antibacterial therapy should be with an antistaphylococcal agent like dicloxacillin plus metronidazole and that free injection paraphernalia with disinfection swabs should be made easily available.

[CT-guided spinal biopsy in spondylodiscitis]. / CT-vejledt borebiopsi ved spondylodiscitis.

Since February 1987 percutaneous CT-guided spine biopsy was performed in 18 patients with spondylodiscitis at the X-ray Department of Bispebjerg Hospital. Eleven cases were spontaneous and seven followed spinal surgery. The infection was located in five cases in the thoracic spine and in 13 cases in the lumbar spine. Only one biopsy was performed during general anaesthesia, the rest under local anaesthesia. No complications were observed. The bioptic material was cultivated immediately beside the patient and incubated for 14 days. The infective organism was isolated in 12 cases (67%). Thus, material obtained through a fine needle was satisfactory for microbiological investigation. A biopsy is crucial for establishing a microbiological diagnosis and thereby enabling prompt adequate treatment.

[Spontaneous and postoperative spondylodiscitis. A material concerning 23 patients]. / Spontan og postoperativ spondylodiscitis. Et materiale på 23 patienter.

A retrospective study of 23 patients with spondylodiscitis is reported. Sixteen cases were spontaneous. Five of these were seen in the acute phase with S. aureus grown from the blood. Eleven patients were investigated with CT-guided biopsy of the spine with identification of different microorganisms in eight cases. In four of seven cases of spondylodiscitis after operation for disc herniation coagulase-negative staphyloccoci were grown after CT-guided biopsy. In spontaneous cases pain disappeared and CRP was normalized within a few weeks after treatment with antibiotics, but radiological changes might progress for several months. Antibiotics were given for two to six months, with a mean of 5.1 months in purulent bacterial infections, and all patients were considered cured after this.