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Methods for testing antibiotic sensitivity of anaerobic bacteria.

Problems are still encountered in the performance and interpretation of tests of anaerobe sensitivity to antibiotics. A review of the methods currently used was carried out in order to determine factors modifying the activity of antibiotics. The sensitivity of Escherichia coli, Staphylococcus aureus and Clostridium perfringens to various drugs was tested under different conditions (including different culture media and incubation atmospheres). Gentamicin and kanamycin showed no activity in tests with brain heart infusion agar incubated anaerobically or in 10% CO2. The activity of lincomycin was much more readily influenced by test conditions than that of clindamycin. All the drugs examined except gentamicin and kanamycin showed more activity when tested in brain heart infusion agar than in Mueller Hinton blood agar.

Anaerobiosis↗

[Antibiotic resistance of anaerobic bacteria (author's transl)].

Reference data on the overall resistance patterns of anaerobic bacteria are needed since in individual clinical cases susceptibility testing is complicated by the long generation time and difficulties of test conditions. Benzylpenicillin and tetracyclines are the drugs of choice against Clostridium perfringens. Newer cephalosporins are also active, whereas other Clostridium species have been found resistant. Beta-lactam antibiotics show high activity against Peptococcaceae; resistance to clindamycin should be noted, however. The good in vitro activity of tetracyclines, clindamycin and erythromycin against Propionibacterium acnes could be confirmed in the systemic and local therapy of acne vulgaris. In contrast to gram-positive anaerobic bacteria, only some penicillin and cephalosporin derivatives are sufficiently active against gram-negative rods of the Bacteroides group. Sporadic resistance is seen to the newer tetrayclines, nitroimidazole compounds and to lincomycins. Nevertheless, these substances must be considered as standard therapeutics for infections caused by anaerobic gram-negative bacteria. It should be mentioned that cefoxitin may offer new therapeutic possibilities.

Actinomyces↗

A comparative study of osteomyelitis and purulent arthritis with special reference to aetiology and recovery.

We analysed the records of 44 paediatric cases of acute haematogenous osteomyelitis (age 0-14 years) and 25 cases of purulent arthritis (age 0-13 years). The annual incidences were 4.5 and less than two per 100,000 children, respectively. Bacteriologic diagnosis was achieved in 82% of the acute haematogenous osteomyelitis cases and in 40% of the acute purulent arthritis cases. Staphylococcus aureus was responsible for 70% of the proven acute haematogenous osteomyelitis and acute purulent arthritis cases combined, followed by streptococci (20%) and Haemophilus influenzae (7%), which caused only acute purulent arthritis. Acute haematogenous osteomyelitis was localized in the femur in 41% of the cases and acute purulent arthritis in the knee joint in 76%. Surgery (in most cases drilling, fenestration or arthrotomy) was performed on 82% of the acute haematogenous osteomyelitis and on 32% of the acute purulent arthritis patients. Although six of the acute haematogenous osteomyelitis patients (but none of the acute purulent arthritis patients) underwent surgery for a second time, permanent damage, which was functionally non-significant, developed in only 14%. No sequelae were found in the acute purulent arthritis group. The average duration of antimicrobial therapy was 44 days in the acute haematogenous osteomyelitis group and 29 days in the acute purulent arthritis group. The prognosis for the children was similar, irrespective of whether the drugs used were staphylococcal penicillins, ampicillin, lincomycin or clindamycin.

Adolescent↗

Antibacterial drugs and their interference with the biogenesis of mitochondria in animal and human cells.

Mitochondrial RNA and protein synthesis in mammalian cells is sensitive to inhibition by a variety of antibiotics which are used in medical practice. In spite of the intrinsic sensitivity of the synthetic processes to these drugs it appears that inhibition in intact cells and living organisms is not observed in all cases because the cellular membranes may act as barriers which prevent the antibiotics from reaching their intramitochondrial targets. This holds for the rifamycins, the lincomycins and a number of macrolides, but not for chloramphenicol and its analogues. Some of the toxic side-effects of the latter antibiotics can be related to their antimitochondrial action. For the tetracyclines selectivity in permeability exists in the sense that some cell types are permeable and other are not. The hypothesis is developed that the deliberate inhibition of mitochondrial protein synthesis in vivo may lead to cell proliferation arrest and offer a device in combined modality treatments of malignant growths. This hypothesis is supported by the results of two studies with experimental tumour models in rats and by retrospective and prospective clinical data.

Animals↗

In vitro susceptibility of Corynebacterium group D2 and Corynebacterium jeikeium to twelve antibiotics.

The in vitro susceptibility of 83 Corynebacterium group D2 strains and 44 Corynebacterium jeikeium strains to 12 antimicrobial agents was determined by an agar dilution technique using Mueller-Hinton agar supplemented with Tween 80 (0.025%). All strains of Corynebacterium group D2 were highly sensitive to fusidic acid, pristinamycin, teicoplanin and vancomycin (geometric mean MICs 0.047, 0.048, 0.338 and 0.396 mg/l respectively). Most of the strains were resistant to other antibiotics tested (ciprofloxacin, erythromycin, gentamicin, lincomycin, rifampin and tetracycline). However, a few strains were highly sensitive (MICs less than or equal to 0.2 mg/l). The overall pattern of susceptibility of 44 strains of Corynebacterium jeikeium was similar; the geometric mean MICs of fusidic acid, pristinamycin, vancomycin and teicoplanin were 0.234, 0.235, 0.557 and 0.652 mg/l respectively.

Anti-Bacterial Agents↗

World-wide development of antibiotic resistance in pneumococci.

Antibiotic-resistant pneumococci, especially penicillin-resistant strains, are being increasingly isolated. Pneumococci with intermediate penicillin-resistance (MIC 0.1-1.0 micrograms/ml) have been reported from many parts of the world over the past two decades, and highly resistant strains (penicillin MICs greater than or equal to 2 micrograms/ml) have also appeared. Infection may be acquired in the hospital or community, and nosocomial outbreaks may occur which require control measures to limit organism spread. Most infections occur in children with diminished host responses. Disease caused by pneumococci with intermediate penicillin-resistance may be treated with high doses of penicillin, but disease caused by highly resistant strains, especially meningitis, may require alternative therapy. Pneumococci resistant to sulfonamides, tetracyclines, erythromycin, lincomycin, clindamycin, chloramphenicol, aminoglycosides and rifampin have also appeared. Strains resistant to all the above-mentioned agents, including all beta-lactam antibiotics tested, have been reported from South Africa and Spain. Alternative therapy for resistant strains may include vancomycin, cefotaxime, cefoperazone, ceftriaxone and imipenem. Pneumococci isolated from sites suggestive of infection, especially blood and cerebrospinal fluid, should be routinely tested for penicillin-susceptibility.

Anti-Bacterial Agents↗

In vitro and in vivo susceptibility of Borrelia burgdorferi.

The antispirochetal activity in vitro and in vivo of several antibiotics against ten isolates of Borrelia burgdorferi from human spinal fluids and skin biopsies was determined. Borrelia burgdorferi was most susceptible in vitro to erythromycin, ceftriaxone and cefotaxime (MIC90: 0.06, 0.06, 0.12 mcg/ml respectively). Less activity was observed with tetracycline, amoxycillin and lincomycin (MIC90: 0.50 mcg/ml), imipenem and augmentin (MIC90: 0.25 mcg/ml), oxacillin (MIC90: 1 mcg/ml), ciprofloxacin (MIC90: 2 mcg/ml) and ofloxacin (MIC90: 4 mcg/ml). Penicillin G, normally regarded as appropriate treatment for Lyme disease, had an MIC90 of only 4 mcg/ml. With the exception of erythromycin, activity in vitro corresponded to the activity in vivo. Erythromycin, however, was less active in vivo, and penicillin G showed poor activity both in vitro and in vivo.

Animals↗

Corneal ulcers in Nepal.

In Kathmandu, Nepal, 133 cases of corneal ulcers were examined and scraped by the investigators during 1981. Corneal ulcers were seen to be more common in females during the active years of life, occurring more frequently during monsoon and autumn seasons. Trauma by vegetative matter was the common antecedent. Organisms were grown in about 50.0% of the cases. Bacteria were seen to be responsible in about 75.0%, and fungi in about 25.0% of growth-positive cases. Sensitivity tests in vitro showed that carbenicillin, chloramphenicol, cephaloridine, methicillin, and lincomycin were most effective against bacteria and clotrimazole was most effective against fungi.

Adolescent↗

Cryopreserved aortic allograft replacement of infected prosthetic grafts in man: processing and clinical results.

Aortic allografts preserved at 4 degrees C have been used successfully for the replacement of infected prosthetic grafts, but have a limited storage duration and this does not allow for rigourous security of the allograft. Original cryopreservation protocol has been developed, characterized by the use of polyethylene glycol 20,000 30 g/l associated with 12.5% DMSO, high concentration of antibiotics (lincomycin 300 mg/l, vancomycin 125 mg/l), controlled freezing rate, and storage in the vapour phase of liquid nitrogen (-150 degrees C). Cryopreserved arterial allografts were used for the replacement of infected prostheses in 22 patients. In 2 patients, allograft-related dysfunction was observed, 3 patients died in the early postoperative period from non-allograft-related causes, and, 17 patients were cured of infection without any dysfunction of the allograft. These results are comparable to replacement by fresh allograft, but cryopreservation allows for better microbiological security, long-term storage, and, thus, better management of the available allografts.

Adult↗

Specific inhibition of 50S ribosomal subunit formation in Staphylococcus aureus cells by 16-membered macrolide, lincosamide, and streptogramin B antibiotics.

The translational functions of the bacterial ribosome are the target for a large number of antimicrobial agents. The 14- and 16-membered macrolides, the lincosamides, and the streptogramin B type antibiotics are thought to share certain inhibitory properties, based on both biochemical and genetic studies. We have shown previously that the 14-membered macrolides, like erythromycin, have an equivalent inhibitory effect on translation and the formation of the 50S ribosomal subunit in growing bacterial cells. To extend this work, we have now tested the 16-membered macrolides spiramycin and tylosin, the lincosamides lincomycin and clindamycin, and 3 streptogramin B compounds pristinamycin I(A), virginiamycin S, and CP37277. Each of these was a specific inhibitor of 50S subunit formation, in addition to having an inhibitory effect on translation. By contrast, two streptogramin A compounds, virginiamycin M1 and CP36926, as well as chloramphenicol, were effective inhibitors of translation without showing a specific effect on the assembly of the large ribosomal subunit. A combination of an A and B type streptogramin (virginiamycin M1 and pristinamycin I(A)) demonstrated a synergistic inhibition of protein synthesis without exhibiting a specific inhibition of 50S subunit formation. These results extend our observations on 50S assembly inhibition to the entire class of MLS(B) antibiotics and reinforce other suggestions concerning their common ribosome-binding site and inhibitory functions.

Anti-Bacterial Agents↗

Protein synthesis by isolated Acetabularia chloroplasts. In vitro synthesis of the apoprotein of the P-700-chlorophyll alpha-protein complex (CP i).

Acetabularia chloroplasts can incorporate radioactive amino acids for up to several hours in vitro. The incorporation is sensitive to chloramphenicol and lincomycin, insensitive to cycloheximide, and completely light-dependent. At least 35 discrete labelled bands can be separated by SDS-polyacrylamide gel electrophoresis: 20--24 in the soluble fraction and 13--15 in the membrane fraction. Most of the label (80--85%) is in the membrane fraction, and 90% of that is in a polypeptide of 32 000 daltons. Chlorophyll-protein complexes were purified from in vitro labelled chloroplasts by SDS-polyacrylamide gel electrophoresis. CP I (P-700-chlorophyll alpha-protein complex) and its apoprotein were both labelled. This shows that the apoprotein is synthesized on chloroplast ribosomes, and can be integrated correctly into the thylakoid membrane in the absence of any cytoplasmic contribution. In contrast, no label was incorporated into the two polypeptides of CP II, the light-harvesting chlorophyll a/b complex.

Acetabularia↗

Differential regulation by trophic conditions of phosphorylating and non-phosphorylating NADP(+)-dependent glyceraldehyde-3-phosphate dehydrogenases in Chlorella fusca.

The two NADP(+)-dependent glyceraldehyde-3-phosphate dehydrogenases present in the green alga Chlorella fusca, namely, the phosphorylating (chloroplastic) enzyme and the non-phosphorylating (cytosolic) enzyme, are differently affected by the trophic conditions prevailing in the cell cultures. The addition of metabolizable sugars to cell cultures growing in the light promotes a marked decrease of the phosphorylating enzyme activity down to a barely detectable cellular level. In contrast, the cellular level of the non-phosphorylating enzyme is even enhanced in the presence of such sugars. These effects are not observed, however, with a number of non-assimilable sugar analogs. After sugar removal, a recovery of the phosphorylating activity--in a process which is inhibited by cycloheximide but not by lincomycin--is observed in illuminated cells but not in darkness, thus indicating a light-dependent nuclear synthesis of the chloroplastic enzyme. It seems therefore that the two dehydrogenases are adaptative enzymes subject to differential regulation by nutritional conditions.

Carbohydrates↗

Dual effects of macrolide antibiotics on rat liver cytochrome P-450. Induction and formation of metabolite-complexes: a structure-activity relationship.

Previous studies have shown that the macrolide antibiotics, troleandomycin and erythromycin, are able to induce their own transformation into a metabolite forming an inactivated complex with rat liver cytochrome P-450. This paper reports the results of a study on the effects of several macrolide antibiotics including oleandomycin, erythromycin derivatives, josamycin, methymycin, tylosin, spiramycin and rifampicin, as well as antibiotics of other series, such as tetracycline and lincomycin, on rat liver cytochromes P-450 in vivo and in vitro. Only the antibiotics containing the desosamine and mycaminose amino sugars were able to give the dual effects already found with troleandomycin: induction of cytochrome p-450 and formation of an inhibitory cytochrome P-450--iron--nitrosoalkane metabolite complex in vivo or in vitro. From these studies, it appears that two structural factors are important for a macrolide antibiotic to lead to such effects: the presence of a non-hindered readily accessible N-dimethylamino group and the hydrophobic character of the molecule. These data are discussed in relation to the adverse effects observed during drug associations involving some of these macrolide antibiotics.

Animals↗

Control of Campylobacter fetus in artificially contaminated bovine semen by incubation with antibiotics before freezing.

Fresh, diluted semen containing 1.55 X 10(6) cfu/ml of Campylobacter fetus subsp. venerealis was incubated with 500 iu of penicillin, 500 micrograms of streptomycin, 160 micrograms of lincomycin and 300 micrograms of spectinomycin per ml at 35 degrees C for 0, 1, 2, 5, 10, 20 or 40 minutes. The semen was cooled to 5 degrees C, packaged in 0.25 ml French straws and then frozen in liquid nitrogen for 2 weeks. Immediately after thawing and removal of the antibiotics by centrifugation semen samples from each of the seven treatment groups were cultured as for C. fetus. Semen samples were also examined by in-vitro tests for sperm motility prior to and post-freezing. Incubation with the antibiotics for 5, 10, 20 or 40 min prior to freezing reduced the numbers of C. fetus in the semen to non-detectable levels in 38%, 69%, 88% and 100% of samples respectively. The incubated semen showed no significant reduction of sperm motility although fertility trials have not been done.

Animals↗

A comparison of josamycin with macrolides and related antibiotics on isolated rat atria.

The effects of macrolide (Josamycin, erythromycin, spiramycin and oleandomycin) and related antibiotics (clindamycin and lincomycin) were examined and compared in spontaneously beating right atrial preparations and in electrically driven left atrial preparations of rats. Josamycin and erythromycin (10(-7)-10(-4) M) produced a dose-dependent decrease in rate. contractile force and maximum following frequency and prolonged the sinus node recovery time and the refractory period. The negative inotropic effect of josamycin was not modified by pretreating the atria with atropine or with a mixture of antagonists containing phentolamine, practolol, diphenhydramine, cimetidine, methysergide and indomethacin. In isolated right atria, josamycin did not block the positive inotropic and chronotropic responses to isoprenaline but shifted the dose-response curve to Ca to the right. Josamycin and erythromycin reduced in a dose-dependent manner the slow responses induced in K-depolarized right atria by isoprenaline but this effect was reversed by increasing the Ca concentration in the bathing media. These findings demonstrate a direct negative inotropic effect of josamycin and suggest that this effect could in some way be explained by inhibiting transmembrane Ca influx into atrial cells. The clinical significance of these results is discussed.

Animals↗

Peptidyl puromycin synthesis; effect of several antibiotics which act on 50 S ribosomal subunits.

The effects of several antibiotics which are know to bind with 50 S ribosomal subunits, on the formation of several di- and tri-peptidyl puromycins have been examined. Tylosin and spiramycin inhibited the formation of phenylalanyl-(14)C-phenylalanyl-puromycin, glycyl-(14)C-phenyllalanyl-puromycin, leucyl-(14)C-phenylalanyl-puromycin, N(epsilon)-carbobenzoxylysyl-(14)C-phenylalanyl-puromycin, and valyl-glycyl-(14)C-phenylalanyl-puromycin as well as N-acetyl-(14)C-phenylalanyl-puromycin. Of these compounds, erythromycin and oleandomycin selectively inhibited the formation of phenylalanyl-(14)C-phenylalanyl-puromycin. Although chloramphenicol and lincomycin inhibited the formation of most of these peptidyl puromycins, the formation of phenylalanyl-(14)C-phenylalanyl-puromycin and leucyl-(14)C-phenylalanyl-puromycin was found to be resistant to these antibiotics. So far, no significant effect of siomycin has been observed on pepetidyl puromycin formation in the absence of G factor.

Journal Article↗

A method for in vitro clearance of mycoplasma from human cell lines.

Rapid and reproducible clearance of mycoplasma contaminating human cell lines was achieved using macrophages and antibiotics. Human peripheral blood monocytes were purified by Percoll density gradient centrifugation and allowed to mature into macrophages by 7 days culture in vitro. To the adherent monolayers of macrophages were added the cells to be cleared. Optimal results were obtained with a macrophage to cell concentration of 100:1, together with 200 micrograms/ml of the antibiotics tylosin and lincomycin. The cleared cells were recovered after 7 days of treatment. Monitoring with the Hoechst 33258 stain demonstrated that cells cleared by this method have remained mycoplasma-free for over 6 months. The method is unlikely to cause cell mutation or to introduce mouse viruses and is effective on both adherent and non-adherent cell lines.

Cell Line↗