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Amino acid sequence and molecular modelling of a thermostable two (4Fe-4S) ferredoxin from the archaebacterium Methanococcus thermolithotrophicus.

The amino acid sequence of a two (4Fe-4S) ferredoxin from the methanogenic bacterium Methanococcus thermolithotrophicus (FdMt) has been determined. This thermostable protein comprises 60 amino acid residues (Mr 6541) and two (4Fe-4S) clusters chelated to the protein through the eight cysteines. FdMt contains a relatively high number of lysines [5], threonines [4] and valines [10]. The three-dimensional molecular model generated from the Peptococcus aerogenes X-ray structure keeps the characteristic overall ferredoxin folding thanks to complementary substitutions of residues of the hydrophobic core. The major structural features of the model are the different environments of both clusters, and the patch of three lysines at one end of the molecule. The possible role of several structural factors in the thermostability of the protein is discussed.

Amino Acid Sequence↗

Non-sporeforming anaerobic bacteria in clean surgical wounds--air and skin contamination.

The contamination of clean surgical wounds with anaerobic and aerobic bacteria was studied in 52 hip operations. In addition to wound samples, samples from air and the patients' skin were taken. The median of the total number of bacteria isolated from the wounds was 26 colony-forming units (cfu). The median percentage of anaerobic bacteria in the wound counts was 30. Propionibacterium spp. were found in 71 per cent of the wounds and anaerobic or microaerophilic cocci, most often Peptococcus spp., were found in 23 per cent. In six of 43 patients the same bacterial flora was seen in the skin samples and wounds. The geometric mean of the total number of bacteria in the air was 70.3 cfu/m3. Of these the median percentage of anaerobic bacteria was 30.3. When operating clothing of a disposable fabric (Barrier 450, Johnson & Johnson) was used, the counts of airborne bacteria were a little less than half those found when conventional cotton clothing was worn. Probably because of the overall low air counts in the operating theatre and the great variability in the individual bacterial counts from the operation wounds, a significant decrease in wound contamination was not observed. A positive correlation was found between the duration of operation and wound contamination.

Adult↗

Phylogenetic and chemotaxonomic characterization of Acidaminococcus fermentans.

The phylogenetic position of Acidaminococcus fermentans was determined by comparative sequence analysis of the 16S rRNA. This Gram-negative bacterium is a member of the Sporomusa cluster that is defined by other Gram-negative bacteria, i.e. Sporomusa, Megasphaera, Selenomonas, Butyrivibrio, Pectinatus, and Zymophilus. The branching point of this group within the radiation of Gram-positive bacteria of the Clostridium/Bacillus subphylum and adjacent to Peptococcus niger could be confirmed. Chemotaxonomic data were provided for a more detailed characterization of A. fermentans.

Amino Acids, Diamino↗

The detection of obligate anaerobic bacteria in udder secretions of dry cattle with mastitis during summer: a comparison between gas-liquid chromatography and bacteriological culturing methods.

Udder secretions sampled during the summer in 1984 and 1985 from mastitic quarters of 51 non-lactating cattle, mainly heifers less than 2 years of age, were examined bacteriologically for the presence of (facultative) aerobic and obligate anaerobic bacteria (OAB) and by gas-liquid chromatography (GLC) in order to detect volatile fatty acids (VFA), metabolic end-products of OAB. Forty-nine samples yielded positive cultures and in 20 cases these were mixtures of (facultative) aerobes and OAB. Only two specimens appeared to be sterile and from one specimen only were OAB cultured. Corynebacterium pyogenes was isolated from 35% of the cases and Peptococcus indolicus and Fusobacterium necrophorum from 31 and 22%, respectively. In most specimens (19/21) which yielded OAB after culturing, VFA (C3-C6) could be detected by GLC. Detection of VFA in summer mastitis secretions appeared to be a useful technique to evaluate the importance and association of OAB with summer mastitis. Because samples can be easily collected and stored at -20 degrees C, this is especially advantageous in situations where adequate facilities for the isolation of OAB are not readily available.

Acute Disease↗

Cutaneous abscesses: natural history and management in an outpatient facility.

This study investigated the natural history and treatment of cutaneous abscesses in an outpatient setting. Incision, drainage, aerobic and anaerobic cultures were done on all 78 patients entered in the study. Tenderness and fluctuance were noted in more than 80% of the patients; erythema and induration in more than 60%. Forty-one percent of all abscesses were in the anogenital region. Forty-two percent of cultured abscesses grew aerobes exclusively, 28% grew anaerobes exclusively, and 27% grew a mixture of aerobes and anaerobes. The predominant aerobic organisms were Staphylococcus and Streptococcus, which were mostly isolated from the head/neck, extremities, and axillary regions. The predominant anaerobic organisms were Peptococcus and Bacteroides, which were primarily isolated from the anogenital regions. Nearly 60% of the patients returned for reevaluation. They were equally divided between those patients taking antibiotics and those not on antibiotics. However, all patients were clinically improved.

Abscess↗

New drugs--reports of new drugs recently approved by the FDA. Dirithromycin.

Dirithromycin is a semisynthetic derivative of erythromycin, a 14-membered ring macrolide antibiotic. The drug is converted during absorption and distribution, to an active metabolite 9-(S)-erythromycylamine, which is the predominant compound found in plasma and extravascular tissues. High tissue concentration of erythromycylamine is achieved after oral doses of dirithromycin, with slow release back into the circulation. The mechanism of action of dirithromycin is like that of erythromycin and other macrolides. These compounds inhibit RNA-dependent protein synthesis. It has recently been suggested that all macrolides stimulate dissociation of peptidyl-tRNA from ribosomes during the elongation phase, leading to inhibited protein synthesis. The antimicrobial spectrum of dirithromycin is similar to that of erythromycin, although the drug offers no significant advantage with regard to MIC values. In vitro against Gram-positive isolates, dirithromycin exhibits similar potency to that of clarithromycin, erythromycin, roxithromycin, and clindamycin. In vivo, dirithromycin is active against penicillin-susceptible Staphylococcus aureus, beta-hemolytic streptococci, and Streptococcus pneumoniae. Dirithromycin is as effective as penicillin VK against streptococcal pharyngitis and tonsilitis, and as effective as erythromycin against acute superimposed chronic bronchitis and skin and soft-tissue infections. In comparison with other newer macrolides, dirithromycin has shown similar or lesser in vitro activity. In particular, Haemophilus influenzae, Bacteroides spp., Peptococcus-Peptostreprococcus spp., Clostridium perfringens, Legionella spp., Neisseria gonorrhoeae, and Chlamydia trachomatis were all less sensitive to dirithromycin than azithromycin or clarithromycin. Once-daily oral administration of dirithromycin (500 mg) has been demonstrated to be similar in efficacy to erythromycin (250 mg, 4 times daily), each for approximately 7 days, in the treatment of acute bronchitis or acute-exacerbations of chronic bronchitis in controlled studies. Proven or presumed pathogen eradication rates were 83 and 86% for acute bronchitis patients treated with dirithromycin and erythromycin, respectively. Corresponding bacteriological response rates in acute exacerbations of chronic bronchitis were 75 to 84% with dirithromycin and 75 to 82% with erythromycin. Both agents achieved clinical cure or improvement in over 85% of the patients with either condition. The main advantage of dirithromycin over erythromycin appears to be once-daily administration. Lilly launched dirithromycin in September 1993, in Spain, received approval from FDA in August 1995, and launched it during October 1995.

Anti-Bacterial Agents↗

In vitro activities of tigecycline against clinical isolates from Shanghai, China.

To evaluate the in vitro activity of tigecycline, the minimum inhibitory concentrations (MICs) of tigecycline against 1,201 strains of recent clinical isolates from 10 hospitals in Shanghai, China were determined and compared with selected comparators. Results showed that tigecycline had broad-spectrum antimicrobial activity. It was highly active against Gram-positive cocci, including methicillin-resistant Staphylococcus spp., penicillin-intermediate Streptococcus pneumoniae, Enterococcus faecalis and E. faecium. Tigecycline also had good activity against most strains of Enterobacteriaceae, Haemophilus influenzae, Neisseria gonorrhoeae, and Moraxella catarrhalis. However, it was poorly active against Acinetobacter baumannii and Pseudomonas aeruginosa. Tigecycline was highly active against anaerobic Gram-positive cocci such as Peptococcus spp. The in vitro activity of tigecycline was significantly better than that of minocycline and tetracycline. It was as active as or slightly more active than vancomycin and teicoplanin in the activity against resistant aerobic Gram-positive cocci. Tigecycline was bactericidal against all Gram-positive cocci tested except Enterococcus spp. Inoculum size but not pH of medium or concentration of human serum in broth had significant effect on the in vitro activity of tigecycline. Aged media (48-72 hours after preparation) used in the test and specific resistance problem in China may have some effects on MIC values of tigecycline.

Anti-Bacterial Agents↗

Cefmenoxime: clinical evaluation.

Cefmenoxime was evaluated in an open trial consisting of 41 patients. Forty infections in 36 patients could be evaluated. Thirteen patients had pyelonephritis due to Escherichia coli (two bacteremic), Pseudomonas aeruginosa, Klebsiella pneumoniae, or Streptococcus faecalis; all improved and 12 of 13 were clinically cured, but one relapse (S. faecalis) occurred at two weeks. Six patients with cystitis due to E. coli, Citrobacter freundii, Serratia marcescens, P. aeruginosa, or S. faecalis all improved, but relapse or reinfection, or both, occurred in five due to P. aeruginosa, S. faecalis, C. fruendii, or E. coli. Neurogenic bladder or other complications were present in five of 13 patients with pyelonephritis and five of six with cystitis. Ten patients with pneumonia and one with tracheobronchitis due to Hemophilus influenzae, S. pneumoniae, S. agalactiae, or Neisseria meningitidis all improved and seven had resolution without relapse, but P. aeruginosa emerged in two patients, one of whom died. Eight soft tissue infections due to Staphylococcus aureus, Peptococcus prevotti, Streptococcus species, or infections of mixed origin resolved in six. Sterility of blood cultures was obtained in one patient with endocarditis due to S. anginosus, but other therapy was substituted. Clinical resolution of the toxic shock syndrome and subsequent negative endocervical cultures for S. aureus occurred in one. Granulocytopenia of unverified cause in four (with less than 1,500 mm3) and two (with less than 2,000 mm3) was reversible. Headache during treatment occurred in six patients and a possible disulfiram-like effect in three. Elevations of serum glutamic oxalacetic transaminase and alkaline phosphatase occurred in five, Coombs' positivity in two, and diarrhea in three. Clinical efficacy of cefmenoxime was significant. Possible side effects require further study.

Abscess↗

Results of noncomparative studies of cefotetan in the treatment of obstetric and gynecologic infections.

In a multicenter trial involving 11 centers, 160 women were enrolled to evaluate the safety and effectiveness of 1 or 2 gm of cefotetan administered every 12 hours in the treatment of obstetric and gynecologic infections. The 133 evaluable patients generally were under 25 years of age, were nonwhite, and had hospital-acquired endometritis or pelvic inflammatory disease caused by both aerobic and anaerobic bacteria. Escherichia coli, Neisseria gonorrhoeae, group D streptococci, Bacteroides sp., and Peptococcus sp. were among the most frequently isolated pathogens. The patients were treated for a mean of 5.6 +/- 1.6 days and received a total dose of 19.27 gm. The signs and symptoms of infection were cleared or improved in 93% of the 133 patients evaluable for clinical response. Of the 116 evaluated bacteriologically, 95% had a satisfactory or presumed satisfactory response; only six patients (5%) were considered to be bacteriologic failures. Differences in the results of several clinical laboratory tests performed before and after treatment were statistically, but not clinically, significant (p less than 0.05). Safety was evaluated in the 158 patients who received cefotetan, and only four (3%) had adverse reactions considered related to the drug. Cefotetan was clearly effective and produced no untoward reactions in these women with obstetric and gynecologic infections caused by both aerobic and anaerobic organisms when administered at 1 or 2 gm every 12 hours.

Adult↗

Moxalactam versus clindamycin plus tobramycin in the treatment of obstetric and gynecologic infections.

The clinical efficacy of moxalactam versus clindamycin/tobramycin was evaluated in a comparative, randomized, prospective study. Sixty patients were treated: 30 with moxalactam and 30 with clindamycin/tobramycin. There were 15 cases of tuboovarian abscess, 36 cases of severe pelvic inflammatory disease with peritonitis, eight cases of endomyometritis, and one wound abscess. Aerobic and anaerobic cultures from the sites of infection yielded 441 microorganisms from 53 patients; an average of 8.3 bacteria per infection (4.5 anaerobes and 3.8 aerobes). The infections tended to be mixed aerobic-anaerobic with anaerobes isolated in 90% of cases. The most frequently isolated possible pathogens were Bacteroides sp. (37), Bacteroides bivius (23), Bacteroides asaccharolyticus (12), Peptococcus asaccharolyticus (29), Peptostreptococcus anaerobius (19), unidentified anaerobic gram-positive cocci (18), Escherichia coli (17), nonhemolytic streptococci (16), Neisseria gonorrhoeae (13), and Gardnerella vaginalis (38). Clinical cure was noted in 29 of 30 moxalactam-treated and 29 of 30 clindamycin/tobramycin-treated patients. Moxalactam was effective in five of six cases of tuboovarian abscess, all 22 cases of pelvic inflammatory disease with peritonitis, the one case of endomyometritis and the one wound abscess. Clindamycin/tobramycin was effective in eight of nine cases of tuboovarian abscess, all 14 cases of pelvic inflammatory disease with peritonitis, and all seven cases of endomyometritis. No adverse hematologic, renal, or hepatic effects were noted with either regimen.

Adolescent↗

A multicenter comparative study of cefotetan once daily and cefoxitin thrice daily for the treatment of infections of the skin and superficial soft tissue.

To compare the effectiveness of cefotetan administered at 2 g once a day with cefoxitin at 1 or 2 g three times a day in the treatment of hospitalized patients with skin and superficial soft tissue infections, 194 patients from eight centers were enrolled in an open, randomized trial. Most of the 104 evaluable patients in the cefotetan group and 50 in the cefoxitin group were young men with community-acquired, moderate or severe cellulitis, or abscesses of the upper and lower extremities caused by Staphylococcus aureus, Streptococcus species, Escherichia coli, Proteus mirabilis, Bacteroides fragilis and other species of bacteroides, peptococcus species, and peptostreptococcus species. The mean duration of treatment was 7.5 days for cefotetan and 7.1 days for cefoxitin. A successful clinical response was achieved in 97 percent of the cefotetan patients and in 94 percent of the cefoxitin patients. Of the 88 and 39 bacteriologically evaluable patients in the cefotetan and cefoxitin groups, respectively, a satisfactory bacteriologic response occurred in 96 percent and 87 percent of the patients. No clinically significant changes in clinical laboratory determinations were noted. The incidence of adverse reactions in the cefotetan group (17 percent) was significantly different from that for the cefoxitin group (6 percent) (p less than 0.05); however, the incidence of treatment-related reactions was not significant and the events were mild. Discontinuation of therapy was necessary only in two patients in whom allergic-type reactions developed. A once-daily regimen of cefotetan was as effective as thrice-daily cefoxitin in this study in the treatment of primarily polymicrobial, moderate, or severe infections of the skin and superficial soft tissue.

Abscess↗

Bacteriology and therapy of lung abscess in children.

The bacteriology and clinical findings of ten pediatric patients with lung abscess are presented. Bacteriologic data were based on percutaneous transtracheal aspiration obtained before initiation of antimicrobial therapy. Anaerobic bacteria were present in all ten patients; in nine they were mixed with aerobic bacteria. The most frequent isolates were Peptostreptococcus (8), Peptococcus (5), Bacteroides melaninogenicus (6), and Bacteroides fragilis (3). The aerobic isolates most frequently recovered were alpha-hemolytic streptococci (5), group A beta-hemolytic streptococci (4), and Escherichia coli and Klebsiella pneumoniae (4 each). Virtually all abscesses were located in dependent pulmonary segments. Antimicrobial therapy was guided by the bacteriologic findings and was successful in all instances.

Adolescent↗

Identification of anaerobic bacteria using high-performance liquid chromatography.

20 species of the genera Clostridium, Bacteroides, Fusobacterium and the type strains of Peptostreptococcus anaerobius, Peptococcus asaccharolyticus, Veillonella parvula and Propionibacterium acnes were examined for the production of volatile (VFA) and nonvolatile (NVFA) short-chain fatty acids using high-performance liquid chromatography (HPLC) with a column for organic acids (Aminex HPX-87H). 10 min are needed for sample preparation and the VFA and NVFA were detected simultaneously in a single chromatographic run. The total time required to run each chromatogram was approximately 60 min. With regard to the production of short-chain fatty acids in culture media it was possible to identify the species of the genera rapidly and clearly. The results illustrate the role of HPLC in determinating short-chain fatty acid products as an additional means for rapid differentiation between closely related anaerobic bacterial species. The aim of the method developed is to establish a complete automatization for the identification of anaerobic bacteria using an automatic sampling system and a microprocessor-controlled chromatography unit.

Bacteria, Anaerobic↗

Deep fasciitis of the biceps region.

An infection of the biceps fascia, later myositis, and spreading to the pectoralis is described in six patients. The majority were synergistic infections with an anaerobic Peptococcus, Eikenella, or Bacteroides and an aerobic Streptococcus. Five of the patients were drug abusers. Excision of the biceps and portions of the pectoralis major was necessary in four patients.

Adult↗

A prospective, randomized, clinical study to compare the clinical safety, effectiveness, and cost of oral ofloxacin/clindamycin vs intravenous clindamycin/gentamicin for the treatment of postpartum endomyometritis.

Objective: The primary objective of this prospective, randomized, clinical study was to compare the safety, clinical and microbiologic efficacy, and cost of oral ofloxacin in combination with clindamycin vs intravenous (IV) clindamycin/gentamicin in the early empiric treatment for hospitalized patients with mild to moderate postpartum endomyometritis. The secondary objective is to reduce total hospital and patient treatment cost. Postpartum endomyometritis is a major cause of infectious morbidity in the obstetric patient. It is the most common complication associated with cesarean delivery. Careful timing and amniotomy, limited vaginal examinations, and prophylactic antibiotics for cesarean section delivery may help to reduce the incidence and severity of endomyometritis. Endomyometritis is caused by bacteria that compose the normal cervicovaginal flora. These are anaerobic gram-positive cocci (Peptostreptococcus and Peptococcus), aerobic streptococci (Group B Streptococci and enterococci), Enterobacteriaceae, Bacteroides (B. fragilis, B. bivius, and B. disiens), and clostridium species.Ofloxacin is a synthetic broad-spectrum antibacterial agent for intravenous and oral administration. Following oral administration, the bioavailability in tablet form is 98% with maximum serum concentrations in 1 to 2 hours. Steady state concentrations are achieved after 4 doses. Ofloxacin usually is bactericidal in action. A synthetic broad-spectrum antibacterial agent for intravenous and oral administration. Ofloxacin inhibits DNA topoisomerase (ATP-hydrolyzing), commonly referred to as DNA-gyrase. DNA-gyrase causes double-stranded DNA breakage; it inhibits duplication, transcription, and repair of bacterial DNA.Methods: This is a preliminary study that has enrolled 19 evaluable patients towards the overall enrollment of 60 patients for statistical significance. Patients clinically diagnosed as having postpartum endomyometritis who meet the inclusion/exclusion criteria were entered into the trial. Patients were examined for the presence of fever (102.2 degrees F), pelvic pain, and foul lochia. A medical history, physical examination, and laboratory analysis were obtained prior to the first dose of antibiotic treatment. A signed consent was obtained prior to the study enrollment and randomization. Appropriate endometrial, blood, and urine culture specimens were obtained prior to the initiation of antibiotic therapy.Patients in Group 1 were treated with oral therapy using ofloxacin 400 mg q12h plus clindamycin 900 mg q8h until 24 hrs of afebrility. In Group 2, patients were treated with clindamycin 900 mg IV q8h plus gentamicin IV 5mg/kg/d q 8h until afebrility. Antibiotic therapy was continued for at least 48 hours unless significant clinical deterioration occurred necessitating the withdrawal of the patient from the study.Results:Conclusions: We found in our preliminary study that oral ofloxacin in combination with oral clindamycin was equally as efficacious, well tolerated, and safe as the combination of intravenous therapy with clindamycin and gentamicin for the treatment of postpartum endomyometritis.

Journal Article↗

Effects of laparoscopic lavage on adhesion formation and peritoneum in an animal model of pelvic inflammatory disease.

STUDY OBJECTIVES: To develop an animal model of pelvic inflammatory disease (PID) that simulates intraluminal tubal damage and adhesion formation seen in human PID, and to evaluate peritoneal effects of laparoscopic lavage with 1% povidone-iodine, 0.5% povidone-iodine, and 0.05% chlorhexidine gluconate solutions. DESIGN: Prospective, randomized, controlled study (Canadian Task Force classification I). SETTING: Animal research operating room facility. SUBJECTS: Forty-five 1-year-old Pasteurella-free New Zealand white rabbits. INTERVENTION: Laparoscopy with instillation of one of five lavage solutions after bacterial inoculation with Neisseria gonorrhoeae, Bacteroides fragilis, Escherichia coli, and Peptococcus niger. MEASUREMENTS AND MAIN RESULTS: Histologic evaluation revealed significantly more peritoneal inflammation after lavage with all three solutions than after no lavage or lavage with normal saline. Gross adhesion formation was visible in only one rabbit after lavage with normal saline. One animal having lavage with 1% povidone-iodine died immediately after injection for sedation before laparoscopic lavage, and was not included in the final results. CONCLUSION: Bacterial inoculation using an intrauterine insemination catheter did not produce clinical adhesions in 43 of 44 rabbits. Histologic evaluation revealed significantly more peritoneal inflammation in the lavage with povidone-iodine and chlorhexidine gluconate compared with no lavage or lavage with normal saline.

Analysis of Variance↗

Dispersal of non-sporeforming anaerobic bacteria from the skin.

Dispersal of non-sporeforming anaerobic bacteria was studied. Skin samples were taken from the subjects, and dispersed from different parts of the body was examined. The number of anaerobic bacteria dispersed was not correlated to their density on the surface of skin area exposed. The highest density of anaerobic bacteria on the skin was found in the face and upper trunk, but the highest yield of anaerobic bacteria dispersed came from the lower trunk. The dominant anaerobic bacteria dispersed were Propionibacterium acnes, but Propionibacterium avidum, Propionibacterium granulosum and Gram-positive cocci were also isolated from the dispersal samples. Peptococcus magnus was the most common coccus isolated. For the less frequently isolated bacteria, the best correlation was found between the perineal flora and airborne bacteria. A comparison was also made of bacterial dispersal by naked and dressed subjects. The dispersal of both aerobic and anaerobic bacteria was higher when the subjects were dressed in conventional operating theatre cotton clothing than when they were naked. The increased dispersal of anaerobic bacteria when the subjects were dressed was mainly due to increased dispersal of Propionibacterium sp.

Actinomycetales↗

Comparison of native and mutant proteins provides a sequence-specific assignment of the cysteinyl ligand proton NMR resonances in the 2[Fe4S4] ferredoxin from Clostridium pasteurianum.

A sequence-specific assignment is presented for the eight low-field paramagnetically shifted cysteinyl ligand proton NMR resonances in the 2[Fe4S4] ferredoxin from Clostridium pasteurianum. The assignment is based upon comparison of chemical shifts in 1D and 2D NMR spectra of native oxidized protein and those of three mutants. The mutant proteins G12A and G41A were designed to produce minor local structural changes (hence small chemical shift perturbations) in either cluster I (glycine 12 to alanine) or in cluster II (glycine 41 to alanine). Observed chemical shift changes in spectra of the double mutant G12,41A support the interpretation. The comparison is aided by structural models derived from the crystal structure of the related ferredoxin from Peptococcus aerogenes. Each of the eight low-field resonances is assigned to a beta-proton from a different cysteinyl ligand, and so connectivities established from previous TOCSY and HMQC data allow assignment of all 24 cysteinyl ligand protons.

Amino Acid Sequence↗