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PubMed1992

The E-Test applied to susceptibility tests of gonococci, multiply-resistant enterococci, and Enterobacteriaceae producing potent beta-lactamases.

The E-Test (AB Biodisk, Solna, Sweden), a new and novel approach developed to test antimicrobial susceptibility, was compared with the agar dilution method. A collection of 57 organisms, including those with known resistances to various antimicrobial agents was tested against 15 drugs using the National Committee for Clinical Laboratory Standard (NCCLS) methods. The E-Test quantitative accuracy (+/- 2 log2 dilutions) compared with agar dilution results was 99% for Neisseria gonorrhoeae, 95% for Escherichia coli strains with extended-spectrum beta-lactamases, 100% for Enterobacter-Citrobacter spp. with type-1 stably derepressed enzymes, and 95% for Enterococcus spp. with various drug resistances (penicillins, ciprofloxacin, gentamicin, and vancomycin). The E-Test procedures with these species were easy to perform, reproducible, and quantitatively and qualitatively accurate (89%-100%). The E-Test appears to be an acceptable option for susceptibility testing of these difficult-to-treat, drug-resistant bacterial strains.

Anti-Bacterial Agents↗

PubMed1994

Comparative in vitro activity of FK-037, a new cephalosporin antibiotic.

The in vitro activity of FK-037, a novel parenteral oxime-type cephalosporin, was compared with that of cefepime, cefpirome, ceftazidime, imipenem, and gentamicin against a total of 668 recent clinical isolates. Minimum inhibitory concentrations were determined by a standard agar dilution procedure, and all isolates were tested at two inocula (10(4) and 10(6) colony forming units). FK-037 inhibited 90% of isolates of Escherichia coli, Klebsiella species, Proteus mirabilis, P. vulgaris, Morganella morganii, Serratia marcescens, Providencia stuartii, Citrobacter freundii, Salmonella typhi, Shigella sonnei, Yersinia enterocolitica, Aeromonas species, and Haemophilus influenzae at < or = 1 microgram/ml. FK-037 was less active against Enterobacter species, Acinetobacter species, and Pseudomonas species, requiring 16 micrograms/ml to inhibit 90% of isolates, and was inactive against Xanthomonas maltophilia. FK-037 inhibited 90% of methicillin-susceptible Staphylococcus aureus at < or = 1 microgram/ml and 90% of methicillin-resistant S. aureus at < or = 8 micrograms/ml.

Bacteria↗

PubMed1994

Cefquinome (HR 111V). In vitro evaluation of a broad-spectrum cephalosporin indicated for infections in animals.

Cefquinome (formerly HR 111V), an aminothiazolyl cephalosporin, was compared with cefepime, cefpirome, cefotaxime, and ceftazidime against 681 clinical cultures and a challenge set of bacteria with well-characterized resistance mechanisms. Cefquinome minimum inhibitory concentrations (MIC90) for the enterobacteriaceae ranged from < or = 0.12-2 micrograms/ml with the highest MIC (4 micrograms/ml) obtained among Citrobacter freundii, Enterobacter cloacae, and Providencia stuartii strains. A total of 90% of the Pseudomonas aeruginosa were inhibited by cefquinome at < or = 8 micrograms/ml. Cefquinome activity of particular note for Gram-positive isolates included Corynebacterium jeikeium (MIC90, 8 micrograms/ml) and enterococci (MIC50, 4-8 micrograms/ml). Oxacillin-resistant Staphylococcus aureus was 32-fold less susceptible (MIC90, 16 micrograms/ml) to cefquinome than oxacillin-susceptible (MIC90, 0.5 micrograms/ml) strains. Cefquinome was very potent against fastidious isolates such as Moraxella catarrhalis (MIC90, 0.25-2 micrograms/ml); Haemophilus influenzae (MIC90, 0.06-1 micrograms/ml), Neisseria gonorrhoeae (MIC90, 0.06-0.5 micrograms/ml), and Streptococcus species (MIC90, < or = 0.03-006 micrograms/ml). When tested against organisms possessing Bush group 2 enzymes (including extended spectrum beta-lactamases), cefquinome remained active (MIC, < or = 8 micrograms/ml) against the majority of strains. This compound should be very active against pathogens generally found in animal infections and possesses a potency and spectrum comparable to the "fourth-generation" cephalosporins (cefepime and cefpirome) being investigated for human infectious diseases.

Animals↗

PubMed1994

In vitro activity of CP-99,219, a novel 7-(3-azabicyclo[3.1.0]hexyl) naphthyridone antimicrobial.

The in vitro activity of CP-99,219 was compared with that of ciprofloxacin and sparfloxacin against 814 clinical bacterial isolates using a microdilution method with brain-heart infusion broth. CP-99,219 was the most potent agent tested against methicillin-resistant, ciprofloxacin-susceptible staphylocci (minimum inhibitory concentration [MIC]90 < or = 0.25 microgram/ml). CP-99,219 was 32-fold and fourfold more potent than ciprofloxacin and sparfloxacin, respectively, against Streptococcus pneumoniae, including strains resistant to penicillin G and erythromycin (MIC90 < or = 0.25 microgram/ml). CP-99,219 was also the most potent agent tested against S. pyogenes and Enterococcus faecalis (MIC90 < or = 0.5 microgram/ml). The activity of CP-99,219 against Enterobacteriaceae was comparable to that of sparfloxacin, with 90% of Escherichia coli, Enterobacter cloacae, Enterobacter aerogenes, Klebsiella pneumoniae, Citrobacter freundii, C. diversus, Helicobacter pylori, and K. oxytoca being inhibited by < or = 0.5 microgram/ml. Serratia marcescens, Morganella morganii, and Pseudomonas aeruginosa were less susceptible, with MIC90 values to CP-99,219 of 4, 2, and 2 micrograms/ml, respectively. The MIC90 for Bacteroides fragilis was 0.39 microgram/ml for CP-99,219 compared with 12.5 micrograms/ml for ciprofloxacin. CP-99,219 was highly bactericidal at 1 x to 4 x MIC against both Gram-positive and Gram-negative organisms; its activity was similar in nutrient, trypticase soy, and cation-supplemented Mueller-Hinton broths. The spectrum and potency observed with CP-99,219 warrant further testing with this novel quinolone.

Anti-Infective Agents↗

PubMed1994

Update of lomefloxacin in vitro activity and spectrum. A multicenter trial testing contemporary pathogens following Food and Drug Administration validation guidelines. Lomefloxacin Activity Study Group.

The United States Food and Drug Administration recently recommended that the antimicrobial product package insert (PPI) subsection on microbiology be annually validated with regard to the compound's spectrum and potency. To address this request, a nine-laboratory trial was organized to test (two methods) lomefloxacin, a newer fluoroquinolone, and nine comparison drugs against PPI-listed pathogens (1934 strains). A broad geographic sampling (nine medical centers) was achieved, and lomefloxacin was determined to be active [minimum inhibitory concentration (MICs), < or = 2 micrograms/ml for > or = 90% of strains] for all PPI-listed species except Pseudomonas aeruginosa, Citrobacter freundii, and Providencia rettgeri (42%-87% inhibited). Comparison fluoroquinolones also had a similarly compromised spectrum of activity against these species. Additional organism species, including Neisseria gonorrhoeae, N. meningitidis, Salmonella enteriditis, and Shigella species, should be added to the lomefloxacin PPI (MIC90s, 0.03-0.25 microgram/ml) following data generated in this study. These in vitro results indicate that lomefloxacin remains active against the vast majority of clinically "indicated" species, and that it has a spectrum compatible with other marketed fluoroquinolones for these tested pathogens, monitored in 1994.

Anti-Bacterial Agents↗

PubMed1994

Antimicrobial susceptibility patterns of bacterial isolates at the American University Medical Center in Lebanon.

In Lebanon, knowledge of the prevailing pattern of bacterial resistance to antimicrobial agents has been limited, particularly because of 15 years of civil strife. Thus, the current study was conducted to determine the antimicrobial susceptibility patterns of nonselected bacterial isolates recovered from recent clinical specimens, using the standardized disk agar diffusion technique. A total of 5216 isolates (1443 Gram positive and 3773 Gram negative) were examined. Over 92% of Staphylococcus aureus and coagulase-negative staphylococci (CNS) were resistant to penicillins. Methicillin resistance was more frequently noted among CNS (28%) compared with S. aureus (18%). For the pneumococci, 27% of the isolates were resistant to penicillin G. High but variable rates of multidrug resistance were encountered among Acinetobacter spp., Pseudomonas spp., Serratia spp., Citrobacter spp., and Enterobacter spp. Ampicillin resistance was detected in 65% of Escherichia coli and in 20% of Haemophilus influenzae isolates. Although one resistant Salmonella typhi strain was observed, 17% of other Salmonella spp. and 60% of Shigella spp. proved to be resistant to ampicillin, chloramphenicol, and cotrimoxazole. Among Vibrio cholerae isolates, high resistance to tetracycline (71%) and trimethoprim-sulfamethoxazole (94%) was observed. The overall antimicrobial resistance rates in Lebanon seem to fall between figures reported from the Arabian Gulf countries (higher) and those from medical centers in the United States (lower).

Drug Resistance, Microbial↗

PubMed1995

National survey of the in vitro spectrum of piperacillin-tazobactam tested against more than 40,000 aerobic clinical isolates from 236 medical centers.

Hospital microbiology laboratories from 41 states participated in a bacterial antimicrobial susceptibility study comparing in vitro results generated by the standardized disk diffusion method. Over 41,000 freshly isolated aerobic and facultative strains, representing all specimen types (except stools and urines), were tested for their susceptibility to piperacillin-tazobactam and 21 other antimicrobial agents. Enterococcus spp. was the second or third most common isolate from intraabdominal, gynecologic, and cutaneous infections, confirming its growing importance as a nosocomial pathogen. Escherichia coli was the most frequent isolate overall, despite the exclusion of urinary tract specimens from the study. Pseudomonas aeruginosa was the second most prevalent species, ranking first in frequency of recovery from lower-respiratory-tract specimens. Piperacillin-tazobactam was the most active beta-lactamase inhibitor combination tested against Gram-negative bacteria. Its activity against Gram-positive bacteria and Haemophilus influenzae was similar to that of ampicillin-sulbactam (95-97% susceptible). Imipenem and piperacillin-tazobactam displayed similar spectrums of activity against Gram-positive organisms and Haemophilus influenzae. Against Enterobacteriaceae, piperacillin-tazobactam and ceftazidime exhibited similarly wide spectrums of activity, but with some gaps, particularly among Enterobacter spp. and Citrobacter freundii. In this large-scale in vitro study, piperacillin-tazobactam and imipenem displayed the widest antimicrobial spectrums, inhibiting > 90% of all isolates tested.

Bacteria↗

PubMed1995

Beta-D-Glucuronidase activity among prototrophic and auxotrophic variants of Escherichia coli and other Enterobacteriaceae commonly implicated in urinary tract infections.

Glucuronidase (GUD) activity of 102 prototrophic, 91 cysteine-requiring, and 19 thymidine-requiring strains of Escherichia coli was examined using growth from MacConkey, CLED, and enriched brain heart infusion (BHI) agars. After 24 h incubation, GUD activity was detected in 92%-96% of prototrophic strains and a similar proportion of thymidine-requiring strains with most reactions detectable in shorter incubation periods. GUD activity among strains requiring cysteine was significantly less than that found amongst prototrophic strains. The effects of different sources of inocula were evident in the shorter incubation periods. Other strains of the Enterobacteriaceae and oxidative strains frequently implicated in urinary tract infection were also tested. Here, positive reactions were detected among Citrobacter and Enterobacter spp. and a strain of Klebsiella oxytoca, but only after 24 h incubation. GUD activity was not detected among the oxidative strains tested under the same conditions. Although an incubation time of 24 h is necessary to detect activity in a small number of "slow hydrolyzing" E. coli, the increased sensitivity thus attained compromises the specificity of the test for this organism by simultaneously enhancing detection of the enzyme in other enterobacteria.

Bacteriological Techniques↗

PubMed1993

Tempe fermentation: some aspects of formation of gamma-linolenic acid, proteases and vitamins.

During a tempe fermentation the concentrations of linoleic, and alpha-linolenic acids (ALA) decreased while the concentration of oleic acid increased. During fatty acid synthesis Rhizopus sp. produced only gamma-linolenic acid (GLA) instead of ALA. The amount of GLA in tempe were influenced by varying external parameters. The proteolytic capacity of 36 strains of the genus Rhizopus isolated from Indonesian tempe or tempe inocula was examined. There was a distinct increase in the amount of free amino acids during tempe fermentation. Fermentations with mixed populations of bacteria and Rhizopus yielded a lower level of free amino acids, but an increase in total amount of amino acids. In comparison to intracellular, and extracellular proteases the proteases of the cell wall fraction are most responsible for proteolytic capacity of the different Rhizopus strains. Two isolated strains of Citrobacter freundii were found to be the best vitamin B(12) producers during the soaking of soybeans. In the solid substrate fermentation the Rhizopus molds formed vitamin B(6), riboflavin, and nicotinic acid. The addition of bacteria to the solid substrate fermentation resulted in a strong increase of active vitamin B(12) in tempe. In the presence of the Rhizopus mold, the vitamin B(12) formation by C. freundii was three times higher than that of a fermentation without the mold.

Journal Article↗

PubMedDate not supplied

Palindromic unit highly repetitive DNA sequences exhibit species specificity within Enterobacteriaceae.

Palindromic units (PU, or REP for repetitive extragenic palindrome) constitute a family of DNA sequences of 40 nucleotides which is highly repeated in the genome of Escherichia coli. We analysed the presence of PU sequences in 99 different bacterial genomes by cross-hybridization. When PU sequences were used as a probe, only DNA from Enterobacteriaceae closely related to E. coli exhibited an appreciable hybridization signal: Shigella sonnei, Shigella boydii, Salmonella enteritica serotype Typhimurium, Citrobacter freundii and Levinea malonatica. Furthermore, these bacteria could be divided into two groups which corresponded to a slight difference in their PU sequence: the E. coli group includes S. sonnei and S. boydii; the S. enteritica serotype Typhimurium group includes C. freundii and L. malonatica.

Blotting, Southern↗

PubMed1992

Novobiocin, brilliant green, glycerol, lactose agar: a new medium for the isolation of Salmonella strains.

We describe a medium, novobiocin, brilliant green, glycerol, lactose (NBGL) agar, for the routine isolation of Salmonella strains from stool samples. The NBGL agar principle involved the use of the antisaprophytic effect of brilliant green and novobiocin. Glycerol and lactose were added in order to distinguish between Citrobacter and Salmonella. NBGL was used in parallel with salmonella-shigella (SS) and Hektoen (H) agar for culturing 2,853 stool samples, of which 184 were confirmed to be salmonellae. NBGL showed a high sensitivity: 94% in direct plating compared to 74% (p < 10(-3)) and 65% (p < 10(-5)) for H and SS, respectively, and 96% in enrichment broth plating vs. 83% (p < 10(-4)) and 86% (p < 10(-3)), respectively, for H and SS. In direct plating using NBGL, 95% of black-centred colonies were confirmed to be salmonellae (vs. 31% and 36%, H and SS). In enrichment plating using NBGL, this figure was 82% (vs. 26% and 28%). The results suggest that NBGL agar is advantageous for the isolation of non-Typhi H2S+ salmonellae.

Anti-Infective Agents, Local↗

PubMed2005

Vacuum-assisted closure for the treatment of complex chest wounds.

BACKGROUND: Destruction of chest wall musculature from trauma, empyema, or local infection limits closure options, especially with muscle flaps. While the vacuum-assisted closure system (VAC; KCI International, San Antonio, TX) has been used for wounds in other anatomic locations, we have found no series for chest wounds. METHODS: This is a retrospective review of trauma registry data from the R Adams Cowley Shock Trauma Center, University of Maryland School of Medicine from 2000 to 2003. RESULTS: Seventeen patients were identified and divided into two groups. Group I consisted of seven patients with primary chest wall processes: four necrotizing soft-tissue infections and three with thoracic trauma resulting in significant loss of chest wall musculature. Group II consisted of ten patients with empyema and varying levels of chest wall extension. Six were postpneumonic and four postoperative. Wound size averaged 16 x 7 cm (range, 7 x 3 cm to 21 x 11 cm). The VAC duration averaged nine days (range, 3 to 21 days) and changed every two to three days. Fourteen wounds were culture positive; nine staphylococcus aureus, two alpha hemolytic streptococcus, and one each with enterococcus, Citrobacter, and anaerobes. Eight were polymicrobial. There were no deaths. All wounds healed without rotational muscle flaps. Ten underwent delayed primary closure, four split-thickness skin graft, and three healed by secondary intention. There was one significant complication: a wound infection after delayed primary closure which required reoperation. CONCLUSIONS: Closure of complex chest wall wounds can present significant technical challenges. The VAC system is a simple, useful, and novel alternative to conventional wound care even with large, infected wounds.

Adult↗

PubMed2003

Mutagenesis of SugE, a small multidrug resistance protein.

The small multidrug resistance protein family has two subclasses. In this study we used a mutation approach to see what is necessary to convert a SUG subgroup member into a quaternary ammonium compound (QAC) transporter. We chose four key residues (H24, M39, I43, and A44) conserved within SUGs but conserved differently within the QAC transporters. Altogether, seven mutants were generated in Citrobacter freundii SugE. Surprisingly, the mutated SugE demonstrated an increased sensitivity to representative QACs. Additionally, ethidium uptake is found to be more prominent in the hypersensitive mutants. We conducted orientation studies using topology reporter gene fusions which indicated that SugE and the QAC transporter EmrE both have their N- and C-termini in the cytoplasm as predicted. The results imply that SugE can be converted to a QAC transporter with only a single mutation. However, because hypersensitivity was observed, the SugE mutant proteins are behaving as importers rather than as exporters.

ATP Binding Cassette Transporter, Subfamily B↗

PubMed2005

Identification of human and rat FAD-AMP lyase (cyclic FMN forming) as ATP-dependent dihydroxyacetone kinases.

Rat liver FAD-AMP lyase or FMN cyclase is the only known enzymatic source of the unusual flavin nucleotide riboflavin 4',5'-cyclic phosphate. To determine its molecular identity, a peptide-mass fingerprint of the purified rat enzyme was obtained. It pointed to highly related, mammalian hypothetical proteins putatively classified as dihydroxyacetone (Dha) kinases due to weaker homologies to biochemically proven Dha kinases of plants, yeasts, and bacteria. The human protein LOC26007 cDNA was used to design PCR primers. The product amplified from human brain cDNA was cloned, sequenced (GenBank Accession No. ), and found to differ from protein LOC26007 cDNA by three SNPs. Its heterologous expression yielded a protein active both as FMN cyclase and ATP-dependent Dha kinase, each activity being inhibited by the substrate(s) of the other. Cyclase and kinase activities copurified from rat liver extracts. Evidence supports that a single protein sustains both activities, probably in a single active center. Putative Dha kinases from other mammals are likely to be FMN cyclases too. Future work will profit from the availability of the structure of Citrobacter freundii Dha kinase, which contains substrate-interacting residues conserved in human Dha kinase/FMN cyclase.

Adenosine Triphosphate↗

PubMed2006

A novel phytase with preferable characteristics from Yersinia intermedia.

A Yersinia intermedia strain producing phytase was isolated from glacier soil. The phytase gene, appA, was isolated by degenerate PCR and TAIL-PCR. The full-length fragment contained 2354bp with a 1326-bp open reading frame encoding 441 amino acids. APPA contained the active site RHGXRXP and HD sequence motifs that are typical of histidine acid phosphatases. To our knowledge, this is the first report of the detection of phytase activity and cloning of the relevant gene from Y. intermedia. The gene was overexpressed in Pichia pastoris, and the purified recombinant APPA had a specific activity for sodium phytate of 3960U/mg, which is higher than that of the Citrobacter braakii phytase (previously the highest specific activity known). Recombinant APPA had high activity from pH 2 to 6 (optimum 4.5) and optimal temperature of 55 degrees C; the enzyme was resistant to pepsin and trypsin. These characteristics suggest that APPA may be highly suitable for use in the feed industry.

6-Phytase↗

PubMed2005

Isolation and characterization of enteric bacteria from the hindgut of Formosan termite.

The Formosan subterranean termite, Coptotermes formosanus Shiraki, is an aggressive, invasive termite species that has caused billions of dollars of damage across the United States for the past 50 years. Termites depend on intestinal microorganisms for cellulose digestion. Symbiotic microorganisms in the termite gut play key physiological functions such as cellulose and hemicellulose digestion, acetogenesis, hydrogenesis, methanogenesis, sulfate reduction, and nitrogen fixation. Additionally, intestinal microbes create suitable conditions for symbiotic protozoans through the production of nutrients and the maintenance of the pH and the anaerobic conditions in the gut. Although extensive research has been done on the symbiotic relationship of these termites and the microbes found in its gut, there is little information available on the role of facultative anaerobes in the gut. We isolated four enteric bacteria from the hindgut of Formosan subterranean termite, C. formosanus. All isolates were facultative anaerobes and G-. The isolates were identified as Serratia marcescens, Enterobacter aerogens, Enterobacter cloacae, and Citrobacter farmeri by using BIOLOG assay and fatty acid methyl ester analysis (FAME). Each isolate was characterized using sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis and biochemical study. This is the first report on the presence of facultative microbes in termite gut. Results of this first study on facultative microbes in the termite gut indicate that the role of facultative organisms in the Formosan termite gut may be to scavenge oxygen and create anaerobic conditions for the anaerobic microorganisms, which are essential for digestion of cellulose consumed by the termite.

Acyltransferases↗

PubMed2026

Multi-omics insights into the physiological mechanisms of bile acid accumulation in the gallbladder in brumation-like snakes.

Hibernation/brumation represents an important physiological adaptation for animals to cope with seasonal environmental changes. Field observations suggested increased gallbladder weight in the Five-pacer viper (Deinagkistrodon acutus) during brumation, and our quantitative measurements confirmed this increase together with bile acid accumulation. By integrating a multi-omic approach, this study elucidates the regulatory mechanisms of bile acid accumulation in the gallbladder during brumation. Results showed that taurocholic acid (TCA) and taurodeoxycholic acid (TDCA) were the major components in the gallbladder of the brumation-like group, with significantly elevated concentrations of bile acids, whereas bile acid concentrations in serum and intestinal contents were markedly reduced, indicating suppression of the enterohepatic circulation and consequent accumulation of bile acids in the gallbladder. Hepatic transcriptomic analysis revealed significant downregulation of bile acid synthesis and regulatory genes in brumation-like snakes. In contrast, the alternative synthesis pathway gene sterol 27-hydroxylase (CYP27A1) and some transporter genes were slightly upregulated. Further, some modification genes and regulatory genes showed no significant differences between active and brumation-like states. Gut microbiota analysis demonstrated Akkermansia muciniphila, Bacteroides fragilis, and Citrobacter freundii were more enriched in the active group, which were common microbes related to bile acid metabolism, and the correlation analysis confirmed this relationship. Taken together, these findings indicate that the "physiological bile acid accumulation" observed in snakes during brumation-like state is jointly driven by suppressed hepatic synthesis, reduced enterohepatic circulation, and remodeled microbial community structure. The study provides novel comparative physiological insights into extreme metabolic homeostasis in animals.

Animals↗

PubMed2004

Bacterial infection and tissue-specific Hsp72, -73 and -90 expression in western painted turtles.

Heat shock proteins (Hsps) are molecular chaperones that assist intracellular folding, assembly and translocation of proteins in prokaryotic and eukaryotic cells. A variety of stresses including hyperthermia, radiation, heavy metals, ischemia, anoxia and reoxygenation have been shown to increase the expression of Hsps. Likewise, bacterial infection represents a stress for the host cell. In this study, expression of the constitutive (Hsp73) and inducible (Hsp72) isoforms of Hsp70 and Hsp90 was monitored in brain, heart, liver and skeletal muscle from the western painted turtle Chrysemys picta bellii diagnosed with Septicemic Cutaneous Ulcerative Dermatitis (SCUD). This disease is caused by a gram-negative bacterium probably belonging to the Citrobacter spp. The expression of Hsp73 increased 1.8-fold in brain and liver, 2.2-fold in heart but did not change in skeletal muscle; Hsp72 expression increased 5.5-fold in brain and 3-fold in liver but did not change in heart or skeletal muscle; Hsp90 expression increased 9-fold in brain, 2.7-fold in heart and 2.4-fold in skeletal muscle but did not change in liver. These results suggest a tissue-specific Hsp response during bacterial infection and a role for Hsps in immunopathological events in reptiles.

Animals↗