Biomedical subjects
Publications and source records attributed to P de Micco.
"Chelex 100" chelating resin has been previously proposed for the rapid extraction of human DNA for polymerase chain reaction. Protocols are given for the rapid extraction of bacterial and viral DNA from cultures or clinical samples. The DNA samples obtained were suitable for use in polymerase chain reaction.
We evaluated an alkaline phosphatase-labeled oligonucleotide probe for the rapid identification of Mycobacterium tuberculosis and mycobacteria belonging to the M avium and M intracellulare complex (MAIS). Sixty-two strains of mycobacteria and eight strains belonging to related genera were studied. All M tuberculosis strains hybridized with the tuberculosis probe. All M avium and M intracellulare gave a strong signal with their probes. However the 3 M xenopi strains tested hybridized with all probes for MAIS complex.
Activity of the new fluoroquinolone sparfloxacin against 30 strains of M. avium complex and 25 strains of M. xenopi was tested in vitro. Sparfloxacin was used alone (determination of MICs and MBCs) and in combination with ethambutol and rifabutin. Synergy studies with determination of the FIC and FBC indices showed that the sparfloxacin-ethambutol combination was synergistic against 10 M. avium complex strains and 12 M. xenopi strains. With the three-drug combination (sparfloxacin-ethambutol-rifabutin), synergy was found against 12 M. avium and 14 M. xenopi strains. Studies of intracellular bacteria showed that the decrease in viable bacteria with the three-drug combination was 1 log for M. avium and 2 log for M. xenopi.
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The isolation of Enterobacter amnigenus from a human source is reported. This microorganism was grown from an intravenous catheter and, along with Pseudomonas aeruginosa, from the blood of a heart transplant patient. The biochemical characteristics and antimicrobial susceptibilities of the isolate are described.
Efficacy of various antibiotics: amikacin, rifampicin, rifabutin, ciprofloxacin, temafloxacin, erythromycin and clarithromycin was evaluated against pigmented and unpigmented variants of Mycobacterium avium complex isolated from patients with acquired immunodeficiency syndrome. The minimal inhibitory concentrations of antibiotics against unpigmented variants were multiplied by 2 to 16 compared to those of pigmented variants. Observations with intracellularly growing bacteria showed that unpigmented variants were more resistant to antibiotics than variants with a difference of approximately 0.25 a 0.3 log 10 in decrease of viable or bacterial counts. The combinations of temafloxacin or ciprofloxacin with rifabutin and amikacin were most effective against unpigmented variants.
Fourteen strains of Mycobacterium marinum were isolated from patients with granulomatous skin lesion. All the strains were resistant to isoniazid and pyrazinamid and five resistant to rifampicin. The following antibiotics are used alone or in combination: ethambutol, rifabutin, ciprofloxacin, temafloxacin, erythromycin, clarithromycin, minocyclin. Minimal inhibitory concentrations are evaluated using agar dilution method. After what, activity of antibiotics against Mycobacterium marinum strains within human macrophages in investigated. After treating the monolayers at 24 and 96 h, a bactericidal effect is observed with rifabutin, ciprofloxacin, temafloxacin, clarithromycin and minocyclin. Combinations of ciprofloxacin, temafloxacin or clarithromycin with ethambutol and rifabutin produced synergistic effect.
This investigation was undertaken to assess the effectiveness of an enzyme-linked immunosorbent assay (ELISA) using A60 antigen in ascertaining diagnosis in hospitalized patients suspected to have pulmonary tuberculosis (TB) but with negative sputum stains. Cultures were performed to confirm active or inactive disease. IgG and IgM antibody activity was determined by adding a 1:100 dilution of serum to plates coated with A60 antigen. After addition of peroxidase-conjugated antihuman IgG or IgM and color development, optical density (OD) was determined. A total of 83 patients was studied, taking into account their current disease status and prior history. Using as a cutoff value the mean value +/- 2 SD measured in the negative culture, no TB history group, that is, OD = 0.50 for IgG measurements and 0.43 for IgM measurements, the sensitivity, specificity, and positive predictive value of IgG measurements were equal to 48, 71, and 50%, respectively. Using IgM measurements, these parameters were equal to 76, 98, and 95%, respectively. Combining the results of IgG and IgM measurements, sensitivity, specificity, and positive predictive value were equal to 68, 100, and 100%, respectively. Thus, the ELISA described here can greatly facilitate the diagnosis of TB in patients with negative smears.
The effect of two new macrolides, azithromycin and roxithromycin used either alone or in combination with amikacin rifabutine and 1.25 (HO)2 vitamin D3 was examined in vitro. Macrophage monolayers infected with M. avium complex or M. xenopi were treated with antibiotics or 1.25 (OH)2 vitamin D3 by using different protocols: antibiotics or 1.25 (OH)2 vitamin D3 was added to the macrophage monolayers immediately after infection and released by washing out after 24 h; antibiotics or 1.25 (OH)2 vitamin D3 was replenished daily for 4 days; or infected macrophage monolayers were treated with antibiotics plus vitamin D3 for 4 consecutive days. Treatment for 24 h resulted in an inhibition of growth. After treating the macrophage monolayers with the four antibiotics alone or in combination for 4 consecutive days intracellular killing of mycobacteria was observed and the combinations were significantly more lethal than antibiotics alone. The mycobactericidal effect was enhanced when vitamin D3 was added to the culture.
The authors compared the post-antibiotic effect (PAE) of imipenem, amikacin, ciprofloxacin, and latamoxef against Serratia marcescens ATCC 13880 (type strain) and against 12 clinical strains belonging to Grimont's most frequent biotypes: A2a, A3a, A3b, A4a, A4b, A5, A6a, A8a, A8b, A8c, TT, TCT. PAE was determined by measuring bacterial growth kinetics after one hour exposure to concentration of 2 x MIC of 10(6) CFUs in Mueller-Hinton broth. Drug removal was by 10-3 dilution of the exposed culture. A PAE was consistently present with imipenem (range 0.8-2.9 hrs), amikacin (range 1.0-4.9 hrs), ciprofloxacin (range 1.4-2.8 hrs). The duration of PAE did not correlate with MIC or Grimont's biotypes.
A DNA biotinylated probe for the rapid identification of Campylobacter jejuni in culture (Enzo Biochem, New York) has been evaluated. The hybridized biotinylated DNA probe is detected by its interaction with streptavidin linked to horseradish peroxidase. Sixteen strains of C. jejuni, including type strain. 24 strains of other Campylobacter and Helicobacter species, and 59 strains of other general have been studied. The probe was highly sensitive (100%) and specific (100% inside the genera Campylobacter and Helicobacter). All Campylobacter strains gave strong signals, and only three weak signals have been observed with non-Campylobacter strains. Our results indicate that specific recombinant DNA probe should offer a reliable and rapid method for routine diagnosis of Campylobacter jejuni enteritis.
We evaluated a DNA biotinylated probe for the rapid identification of Neisseria gonorrhoeae in culture (Ortho Diagnostics System, Raritan, NJ). Twenty-one strains of N. gonorrhoeae, including type strain, 57 strains of other Neisseria species, and 104 strains of other genera were studied. The probe was highly sensitive (100%) and specific (96%). All N. gonorrhoeae strains gave strong signals, and only two cross-reaction were observed with N. lactamica, which has a close genetic relationship to N. gonorrhoeae. Our results indicate that specific recombinant DNA probe should offer a reliable and rapid method for routine diagnosis of Neisseria gonorrhoeae.
The authors have tested the sensibility to rifabutine and clofazimine of seventeen strains of M. avium and eleven strains of M. xenopi. They compared the ability of these and other antimicrobial agents ciprofloxacin, amikacin and erythromycin alone or in combinations to inhibit the growth of strains of the M. avium complex and of M. xenopi. The purpose of this investigation was also to examine the bactericidal action of these antimicrobial agents and to determine the ability of combinations to kill strains of M. avium and M. xenopi.
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Exogen cell fusion induced in vitro by a wild strain of Visna virus of Sheep is compared with two mutant strains isolated from the precedent. One of them produces large plaques in vitro (strain LPF), and the other produces small plaques (strain SPF). These strains behave in different ways according to infection multiplicity, type of cells, temperature and timing of fusing activity. Taking the wild strain K 796 as a base of reference, the strain SPF seems to have a higher rate fusing activity and the strain LPF lower rate of fusing activity.
A method is described to demonstrate and measure the cell-fusion in vitro induced by viruses. This technique has been established using Sendai virus. It has been used to study the fusing ability of the Scrapie agent which is responsible of a slowly progressing spongiform encephalopathy in Mice. In Vero cells, the Scrapie agent induces a fusion which appears slowly and remains moderate. This test can help to detect the human spongiform encephalopathies.
A studying method of viruses induced cell-fusion is described, using monolayers of Vero monkey cells. Nature and density of cells and infection multiplicity are studied. The sensibility is equal to 4 hemagglutining units of Sendaï and accuracy about +/- 3 % of polykaryocytosis. Quantification. comparison and kinetics study of cell-fusion are allowed by this method which may lead to systematic and routine exploration of cell-fusion ability of biological extracts from slow degenerative diseases of the human central nervous system.
The authors study cell fusion in vitro induced by two kinds of slow viruses : visna virus of sheep and scrapie of mouse and Creutzfeldt-Jakob disease of man agents. Cell-fusion induced by visna virus is either slow and late occuring (fusion from within) or early and rapid (fusion from without) according to experimental conditions. Whereas, cell-fusion induced by spongiform encephalopathies agents is only of slow appearance, looking an endogenous phenomenon.