Citrobacter

Clin Infect Dis, 1996 Sep, 23(3), 543 - 9
Bacteremia due to Citrobacter species: significance of primary intraabdominal infection; Shih CC et al.; From 1982 to 1994, 45 patients (1.22 episodes per 10,000 discharged patients) were treated for citrobacter bacteremia at National Taiwan University Hospital (Taipei) . All patients had at least one underlying disease . Citrobacter bacteremia most commonly occurred in patients with malignancies (48.9%) or hepatobiliary stones (22.2%) . Intraabdominal tumors comprised the majority (59.1%) of malignancies . Bacteremia commonly originated from sites such as the abdominal cavity (51.1%), urinary tract (20%), and lung (11.1%) . Polymicrobial bacteremia was diagnosed in 15 patients (33.3%); for nine (60%) of these patients, the source of the infection was intraabdominal . Prior treatment with a third-generation cephalosporin was significantly associated (P < .01) with the development of multidrug resistance among the isolates . The mortality associated with citrobacter bacteremia was 17.8% . Poor prognostic factors included pneumonia, altered mental status on presentation, hypothermia, oliguria, septic shock, deterioration in mental status, hyperbilirubinemia, azotemia, and thrombocytopenia . Combination therapy, as compared with other regimens, improved the outcome of citrobacter bacteremia.

Appl Environ Microbiol, 1996 Sep, 62(9), 3350 - 4
Confirmational identification of Escherichia coli, a comparison of genotypic and phenotypic assays for glutamate decarboxylase and beta-D-glucuronidase; McDaniels AE et al.; Genotypic and phenotypic assays for glutamate decarboxylase (GAD) and beta-D-glucuronidase (GUD) were compared for their abilities to detect various strains of Escherichia coli and to discriminate among other bacterial species . Test strains included nonpathogenic E . coli, three major groups of diarrheagenic E . coli, three other non-coli Escherichia species, and various other gram-negative and -positive bacteria found in water . The genotypic assays were performed with hybridization probes generated by PCR amplification of 670- and 623-bp segments of the gadA/B (GAD) and uidA (GUD) genes, respectively . The GAD enzymes catalyze the alpha-decarboxylation of L-glutamic acid to yield gamma-aminobutyric acid and carbon dioxide, which are detected in the phenotypic assay by a pH-sensitive indicator dye . The phenotypic assay for GUD involves the transformation of 4-methylumbelliferyl-beta-D-glucuronide to the fluorogenic compound 4-methylumbelliferone . The GAD phenotypic assay detected the majority of the E . coli strains tested, whereas a number of these strains, including all representatives of the O157:H7 serotype and several nonpathogenic E . coli strains, gave negative results in the GUD assay . Both phenotypic assays detected some but not all strains from each of the four Shigella species . A strain of Citrobacter freundii was also detected by the GUD assay but not by the GAD assay . All E . coli and Shigella strains were detected with both the gadA/B and uidA probes . A few Escherichia fergusonii strains gave weak hybridization signals in response to both probes at 65 degrees C but not at 68 degrees C . None of the other bacterial species tested were detected by either probe . These results were consistent with previous reports which have indicated that the GAD phenotypic assay detects a wider range of E . coli strains than does the GUD assay and is also somewhat more specific for this species . The genotypic assays for the two enzymes were found to be equivalent in both of these respects and superior to both of the phenotypic assays in terms of the range of E . coli strains and isolates detected.

Antimicrob Agents Chemother, 1996 Aug, 40(8), 1926 - 30
Comparative characterization of the cephamycinase blaCMY-1 gene and its relationship with other beta-lactamase genes; Bauernfeind A et al.; A plasmidic beta-lactamase which hydrolyzed cephamycins was first detected and reported in 1989 . At that time its description was restricted to phenotypic characteristics . We analyzed nucleotide sequence of its gene and explored it genetic relationship with other bla genes . The deduced amino acid sequence of the blaCMY-1 product was compared with those of other known plasmidic cephamycinases and of chromosomal AmpC beta-lactamases . The results indicate that the relationship of CMY-1 is closest to MOX-1 among the plasmidic cephamycinases and to AmpC of Pseudomonas aeruginosa among the chromosomal cephalosporinases . We conclude that the plasmidic cephamycinases described up to now may be classified into three families, as follows: CMY-1, MOX-1, and FOX-1 with AmpC of P . aeruginosa; CMY-2, BIL-1 and LAT-1 with AmpC of Citrobacter freundii; and MIR-1 with AmpC of Enterobacter cloacae . Plasmidic cephamycinases are now recognized as clinically relevant class C beta-lactamases.

APMIS, 1996 Jul-Aug, 104(7-8), 583 - 90
The influence of incubation conditions on the adherence of oral Enterobacteriaceae to HeLa cells; Sedgley CM et al.; The adherence of 12 oral isolates and 4 type strains of Enterobacteriaceae (equally representing Enterobacter cloacae, Klebsiella pneumoniae, Escherichia coli and Citrobacter freundii) to HeLa cell monolayers following five different incubation conditions (sucrose, D-mannose, serum, MEM and Candida albicans GDH 1957) was investigated . Incubation with sucrose and D-mannose resulted in the greatest and least adherence, respectively . The presence of preadherent C . albicans GDH 1957 on the HeLa cells tended to enhance the adherence of certain strains of E . cloacae and C . freundii, but had no overall impact on Enterobacteriaceae adherence . While heterogeneity of behaviour existed between strains within species, E . cloacae was the most, and K . pneumoniae the least, adherent species irrespective of incubation conditions . Haemagglutination assays indicated the presence of mannose-resistant type 1 fimbriae associated with all Enterobacteriaceae . In clinical terms, the variations in adherence properties observed in vitro may contribute to an understanding of the different prevalence rates of oral Enterobacteriaceae reported in the literature.

Trans R Soc Trop Med Hyg, 1996 Jul-Aug, 90(4), 347 - 52
Royal Society of Tropical Medicine and Hygiene meeting at Manson House, London, 14 December 1995 . Enteropathogenic Escherichia coli--mucosal infection models; Frankel G et al.; The formation of attaching and effacing (A/E) lesions is central to the pathogenesis of enteropathogenic Escherichia coli (EPEC)-mediated disease in humans and Citrobacter rodentium-mediated transmissible colonic hyperplasia in mice . Closely related outer membrane proteins, known as intimins, are required for formation of the A/E lesion by both EPEC and C . rodentium . In this study we found similar ultrastructural damage in small intestinal biopsies from an EPEC-infected child and large bowel specimens from C . rodentium-infected mice . The C . rodentium-infected large bowel biopsies revealed massive hyperplastic reactions and the infected human small intestinal biopsies showed an increase in total crypt cell number and mitotic index . EPEC-infected small intestinal organ cultures revealed bacteria adhering in a localized pattern and evidence of A/E lesions . Covaspheres coated with a biologically active cell-binding domain of intimin also adhered to cells in a localized fashion but did not induce the characteristic A/E lesions.

Antimicrob Agents Chemother, 1996 Jul, 40(7), 1736 - 40
Transferable cefoxitin resistance in enterobacteria from Greek hospitals and characterization of a plasmid-mediated group 1 beta-lactamase (LAT-2); Gazouli M et al.; Cefoxitin resistance in Klebsiella pneumoniae from Escherichia coli strains isolated in Greek hospitals was found to be due to the acquisition of similar plasmids coding for group 1 beta-lactamases . The plasmids were not self-transferable but were mobilized by conjugative plasmids . These elements have also been spread to Enterobacter aerogenes . The most common enzyme was a Citrobacter freundii-derived cephalosporinase (LAT-2) which differed from LAT-1 by three amino acids.

J Clin Microbiol, 1996 Jul, 34(7), 1788 - 93
Evaluation of CHROMagar Orientation for differentiation and presumptive identification of gram-negative bacilli and Enterococcus species; Merlino J et al.; A new chromogenic plate medium, CHROMagar Orientation, was evaluated for use in the differentiation and presumptive identification of gram-negative bacilli and Enterococcus species by a multipoint inoculation (replicator) technique . In this study, 1,404 gram-negative bacilli and 74 enterococcal isolates were tested on CHROMagar Orientation . Six control American Type Culture Collection strains were also included with the testing to ensure quality control of the media . Of the Escherichia coli isolates (n = 588) tested, 99.3% produced a pink-to-red color . Only in four isolates that were O-nitrophenyl-beta-D-galactopyranoside (ONPG) negative did this result differ . Proteus mirabilis and P . vulgaris were well differentiated on this medium . P . mirabilis (n = 184) produced a clear colony with diffusible brown pigment around the periphery . By contrast, 15 of 16 P . vulgaris isolates produced bluish-green colonies with a slight brown background . All Aeromonas hydrophila isolates (n = 26) tested produced clear to pink colonies at 35 to 37 degrees C . This colony color changed to blue after 2 to 3 h of incubation at room temperature . A . hydrophila exhibited stronger color and better growth at 30 degrees C . Serratia marcescens (n = 29) demonstrated an aqua blue color that deepened to a darker blue when exposed to room temperature . All enterococcal isolates (n = 74) resulted in a blue color and gave pinpoint colonies on purity subcultures at 35 to 37 degrees C after 18 h of incubation . Similarity in color resulted in failure to discriminate accurately between Klebsiella, Enterobacter, and Citrobacter species . However, these species could be readily differentiated from other members of the family Enterobacteriaceae . Pseudomonas aeruginosa (n = 151) was easily differentiated from members of the Enterobacteriaceae but was less easily distinguishable from other gram-negative nonmembers of the Enterobacteriaceae . The medium was found to facilitate easy visual detection of mixed bacterial isolates in culture . When used in a replicator system, it easily detected mixed growths of organisms which may have otherwise led to false antibiotic susceptibility results . These mixed growths were not obvious on the routine susceptibility testing medium (Isosensitest).

Am J Med, 1996 Jun 24, 100(6A), 45S - 51S
Bacteriologic and clinical applications of a new extended-spectrum parenteral cephalosporin; Segreti J et al.; Although third-generation cephalosporins have been considered the backbone of antibiotic therapy for the treatment of many kinds of serious infections, including those in hospitalized patients, lack of activity against some important pathogens still exists among currently available drugs . In addition, increasing accounts of antibiotic resistance, particularly in the hospital environment, are of deep concern and have thus led to the need for the development of newer antimicrobial agents . Cefepime is a now parenteral cephalosporin with an extended spectrum of antibacterial activity that includes both aerobic gram-negative and gram-positive bacteria . It is also active against many gram-negative organisms resistant to ceftriaxone and cefotaxime, as well as many strains of Enterobacter and Citrobacter resistant to ceftazidime . Cefepime appears to be less likely to select out resistant organisms, and it may be less likely to change hospital flora than currently available antimicrobials . Cefepime has been shown to be very well tolerated and effective in the treatment of a variety of infections including moderate-to-severe pneumonia (including cases associated with concurrent bacteremia), complicated and uncomplicated urinary tract infections (also including cases associated with concurrent bacteremia), and skin and skin-structure infections . Clinical response rates are > or = 75% for most infections and have been comparable to ceftazidime in comparative trials . In addition, pretreatment susceptibility testing indicates that >94% of organisms isolated in patients enrolled in clinical trials were susceptible to cefepime.

Am J Med, 1996 Jun 24, 100(6A), 26S - 38S
Comparative activity of eight antimicrobial agents against clinical bacterial isolates from the United States, measured by two methods; Thornsberry C et al.; In a surveillance study conducted during 1992-1993 at 83 medical institutions of different types and sizes (e.g., laboratories, community hospitals, teaching hospitals) and from different geographical areas of the United States, clinical bacterial isolates were tested for their susceptibility to eight comparative antimicrobial agents (cefepime, ceftazidime, cefotaxime, ceftriaxone, ciprofloxacin, gentamicin, imipenem, and piperacillin) . A total of 12,574 isolates were tested by either the Etest method (AB Biodisk) or a microdilution method (MicroScan) in the participating laboratories; 11.8% of these isolates were subsequently retested for quality assurance purposes by both methods in a central laboratory . The results obtained in the central laboratory were essentially the same as the results obtained in the participating laboratories . This article presents data for gram-negative and gram-positive isolates other than Streptococcus pneumoniae, the results of which have been previously published . Antimicrobial susceptibility results obtained with the two different minimum inhibitory concentration (MIC) methods--MicroScan and Etest--showed that most isolates of Enterobacteriaceae were susceptible to cefepime, exceeding the activity of ceftazidime, ceftriaxone, and cefotaxime, principally because of the greater activity of cefepime against the species that produce Bush group 1 beta-lactamases (predominantly Enterobacter cloacae, Enterobacter aerogenes, and Citrobacter freundii) . In addition, the activity of cefepime against Pseudomonas aeruginosa isolates was essentially equivalent to that of ceftazidime and greater than that of third-generation cephalosporins . Most methicillin-susceptible Staphylococcus aureus were susceptible to all the cephalosporins, whereas methicillin-resistant S . aureus and enterococci were resistant . Overall, the most active antimicrobials in this study were imipenem, ciprofloxacin, and cefepime, but the activity of all the antimicrobials varied with different species . Categorically, the results from the microdilution and Etest methods were equivalent.

Am J Med, 1996 Jun 24, 100(6A), 13S - 19S
Susceptibility of bacterial isolates to beta-lactam antibiotics from U.S . clinical trials over a 5-year period; Kessler RE et al.; Results are reported for agar dilution susceptibility testing of 3,075 isolates of aerobic bacteria collected from >200 U.S . institutions, located in 30 different states . These isolates were collected from 1987 through 1991 from patients who participated in cefepime clinical trials . Cefepime susceptibility was compared with ceftazidime, cefotaxime, ceftriaxone, cefoperazone, and imipenem . To avoid duplication of strains, only initial isolates were included . Cefepime minimum inhibitory concentration (MIC90) values for Enterobacteriaceae were < or = 0.5 microg/mL, except for two species, Citrobacter freundii and Providencia stuartii, with MIC90 values of 2 and 1, respectively . The MIC90 values of the other cephalosporins were higher, especially for Enterobacter aerogenes and C . freundii . The MIC90 values of cefepime for methicillin-susceptible Staphylococcus aureus (4 microg/mL) and Pseudomonas aeruginosa (8 microg/mL) were similar to those of cefotaxime for S . aureus (4 microg/mL), and to ceftazidime for P . aeruginosa (8 microg/mL) . Streptococcus pneumoniae was similar in susceptibility to cefotaxime at 0.06 microg/mL . The activity of cefepime against a diverse group of gram-positive and gram-negative (1987-1991) bacteria isolates demonstrates the excellent activity of cefepime compared to third-generation cephalosporins and imipenem, particularly among C . freundii and E . aerogenes isolates, which were often resistant to other cephalosporins.

J Appl Bacteriol, 1996 Jun, 80(6), 659 - 66
Use of two 16S DNA targeted oligonucleotides as PCR primers for the specific detection of Salmonella in foods; Lin CK et al.; A 16S DNA targeted polymerase chain reaction (PCR) method specific for the detection of Salmonella isolates with various serotypes was developed . The primers used for such a PCR method were 16SF1 and 16SIII . 16SF1 is the reverse and complementary strand of 16SI which has been shown to be able to hybridize with Salmonella and Citrobacter spp . 16III on the other hand, is able to hybridize with Klebsiella and Serratia spp . in addition to Salmonella . Although 16SF1 and 16SIII were not specific to Salmonella only, when they were used as PCR primers, only the Salmonella isolates could be specifically detected . The interference from Citrobacter, Klebsiella and Serratia spp . could be prevented . None of the other non-Salmonella isolates including strains of the family of Enterobacteriaceae closely related to Salmonella would generate the false-positive reaction . When this PCR system was used for the detection of Salmonella cells artificially contaminated in food samples, results obtained were satisfactory . A detection limit of N x 10(0) cells per assay could be obtained.

J Immunol, 1996 Jun 1, 156(11), 4466 - 73
Inhibition of the hemolytic activity of the first component of complement C1 by an Escherichia coli C1q binding protein; van den Berg RH et al.; Molecular mimicry is a well established mechanism via which bacteria protect themselves from complement-mediated killing . We have previously demonstrated that a number of human cells express receptors for C1q (C1qR) and that the soluble form of this receptor inhibits activation of the classical pathway of complement . We now investigated whether Escherichia coli possesses a C1qR-like protein that protects these bacteria from complement-mediated injury . By FACS analysis it was shown that approximately 60% of the bacteria bound C1q directly in the absence of Abs . With ELISA we confirmed that the bacterial cell envelope was able to bind C1q in a dose-dependent fashion . We isolated a cell envelope associated C1q binding protein (C1qBP) by C1q affinity chromatography, then by anion exchange chromatography and gel filtration chromatography . On SDS-PAGE, the m.w . of C1qBP appeared to be 57 kDa and 51 kDa under reducing and nonreducing conditions, respectively . It was demonstrated that C1qBP specifically binds C1q and inhibits the hemolytic activity of C1q in both a dose- and time-dependent fashion . The binding of C1qBP to C1q is inhibited by C1q itself and also by the collagen-like stalks and the globular heads of C1q . In this respect, bacterial C1qBP is different from human C1qR because the binding of C1q to C1qR is only inhibited by the collagen-like stalks of C1q and not by the globular heads of C1q . C1qBP, when bound to C1q, prevents the assembly with C1r and C1s to form a functional C1 complex . The occurrence of C1qBP is not limited to certain E . coli strains, but is also found on Staphylococcus aureus, Citrobacter freundii, and Pseudomonas aeruginosa . Also, the binding of 125(I)-labeled C1q to these bacteria is specific because the binding of C1q to these bacteria is inhibitable with isolated soluble C1qBP . These findings provide evidence for the existence of a C1qR-like protein on bacteria that might protect them from complement-mediated damage.

J Bacteriol, 1996 Jun, 178(11), 3072 - 6
Growth suppression in early-stationary-phase nutrient broth cultures of Salmonella typhimurium and Escherichia coli is genus specific and not regulated by sigma S; Barrow PA et al.; We have studied the growth suppression seen in early-stationary-phase LB broth cultures of Salmonella typhimurium . Multiplication of small numbers of an antibiotic-resistant S . typhimurium mutant was prevented when the mutant was added to 24-h cultures of the antibiotic-sensitive parent strain, whereas an antibiotic-resistant mutant of an Escherichia coli strain added to the same culture grew well . A 24-h E . coli culture produced a similar specific bacteriostatic inhibition against E . coli . In older cultures, a specific bactericidal effect similar to that observed by M . M . Zambrano and R . Kolter (J . Bacteriol . 175:5642-5647, 1993) was also observed . Whether incubated statically or shaken, sufficient nutrients were present in the filtered supernatants of 24-h cultures for small inocula of the same strain to multiply to ca . 10(9) CFU/ml after reincubation . Introduction of the rpoS mutation had no effect on the specific bacteriostatic inhibition . Similar specific inhibition was also observed in strains of Citrobacter freundii, Klebsiella pneumoniae, Enterobacter agglomerans, and Shigella spp . Experiments in which the 24-h culture was physically separated from the antibiotic-resistant mutant by using a dialysis membrane were carried out . These results indicated that the inhibition might be mediated by a diffusible but labile chemical mediator.

Gig Sanit, 1996 May-Jun, (3), 22 - 3
{Enterobacteria isolated from commercial fishes from the Volga-Caspian basin}; Lartseva LV et al.; Enteric bacteria, such as Proteus, Citrobacter {correction of Cytobacter}, and Providencia, were found to be prevalent particularly in the gills and intestines of marketable fishes of the Volga and Caspian Seas . Half of all the enteric bacteria was detected in the fish kidneys, spleen, and liver . This shows it necessary to standardize different types of fish raw materials.

Lett Appl Microbiol, 1996 May, 22(5), 378 - 80
The use of a rapid Salmonella latex serogrouping test (Spectate) to assist in the confirmation of ELISA-based rapid Salmonella screening tests; Cheesbrough S et al.; Spectate, a 10 min, simple, latex-based agglutination test for serogrouping of salmonellae, has been investigated as a tool to assist in the confirmation of ELISA presumptive positive broth samples when screening for salmonellae in foods . When obtaining a combined ELISA and Spectate positive result, there was a 90% (27/30) confidence limit of a genuine positive result, some one or two working days quicker than the traditional methods . Of the 10% (3/30) that were not confirmed as salmonellas, two gave a Spectate result which is advised as being a possible Citrobacter spp . and one sample gave a positive confirmation by Spectate only, which suggested a failure of the traditional confirmation process, a finding confirmed at other sites . Extensive studies performed at several food company microbiology laboratories showed Spectate to be a useful additional tool in the confirmation process of ELISA screening techniques for salmonellae in food . Additionally the concept of a "false positive' may need to be refined to that of a "not culturally' positive, given the apparent possible failure of the traditional confirmation methods.

Z Naturforsch {C}, 1996 May-Jun, 51(5-6), 363 - 70
The catalytic mechanism of tyrosine phenol-lyase from Erwinia herbicola: the effect of substrate structure on pH-dependence of kinetic parameters in the reactions with ring-substituted tyrosines; Faleev NG et al.; Apparently homogeneous tyrosine phenol-lyase (TPL) from Erwinia herbicola has been prepared by a new method . The pH-dependencies of the main kinetic parameters for the reactions of Erwinia TPL with tyrosine, 2-fluorotyrosine, 3-fluorotyrosine, 2-chlorotyrosine, and 3,4-dihydroxyphenylalanine (DOPA) have been studied . The pattern of pH-dependence of V(max) depends on the nature of the substituent in the aromatic ring . For the substrates bearing small substituents (H, 2-F, 3-F) V(max) values were found to be pH-independent . For 2-chlorotyrosine and DOPA V(max) decreased at lower pH, the effect being described by equation with one pKa . Generally two bases are reflected in the pH dependence of V(max)/Km . The first base, probably is responsible for the abstraction of alpha-proton, while the second one, interacts with the phenolic hydroxyl at the stage of binding . The reaction of TPL with DOPA differs from the reactions with other tyrosines by the requirement of an additional base which is reflected in the pH-profiles of both V(max) and V(max)/Km . For the reaction of TPL from Citrobacter intermedius with DOPA only V(max)/Km values could be determined . The activity of Citrobacter enzyme towards DOPA is considerably less than that of E . herbicola enzyme, and its maximal value is attained at higher pH.

Enferm Infecc Microbiol Clin, 1996 Apr, 14(4), 211 - 4
{Fluoroquinolone and aminoglycoside resistance in chromosomal cephalosporinase-overproducing gram-negative bacilli strains with inducible beta-lactamase}; Lopez-Yeste M et al.; INTRODUCTION . The increasing prevalence of stable derepressed mutants over-producers of type I chromosomal cephalosporinase in inducible Enterobacteriaceae and Pseudomonas aeruginosa challenges the adequacy of third generation cephalosporins in the empirical treatment of certain nosocomial infections . We sought to determine the frequency of stable over-producers of type I enzyme and their associated resistance to fluoroquinolones and aminoglycosides . METHODS . Disc-diffusion and MIC determinations to extended-spectrum beta-lactams, imipenem, ciprofloxacin and gentamicin were performed in all cell isolates of inducible enteric bacteria (Enterobacter spp., Citrobacter spp., Serratia spp., Morganella morganii, Providencia spp.) and P . aeruginosa collected during the period of study (1992-1993) . RESULTS . A total of 1,426 isolates of inducible enteric bacteria and P . aeruginosa were studied . Each one represented a single patient . Among the 511 isolates of enteric bacteria 15.1% of strains were found to be stable derepressed mutants (Serratia 2.2%; Morganella spp., 3%; Providencia and Proteus 3%; Citrobacter spp., 10%; Enterobacter spp., 23.6%); among the 916 P . aeruginosa isolates studied, 9.2% were stable over-producers . Among Citrobacter, Providencia and Proteus spp., 53.1% of stable over-producers were resistant to ciprofloxacin versus 20.2% of non-over-producers (p < 0.01); in P . aeruginosa, 35.3% of over-producers were resistant to gentamicin versus 25.0% in non-over-producers (p < 0.01) . CONCLUSION . The prevalence of stable derepressed mutants is high among enteric bacteria and P . aeruginosa with type I inducible beta-lactamase . These strains frequently exhibit resistance to fluoroquinolones and aminoglycosides, reducing considerably the available therapeutic options.

FEMS Microbiol Lett, 1996 Apr 1, 137(2-3), 141 - 6
Iron utilization studies in Citrobacter species; Khashe S et al.; Seventy-one strains of Citrobacter were screened for iron scavenging mechanisms by biologic and chemical assays . Essentially all citrobacteria (70/71) were found to elaborate enterobactin-like siderophores by both biologic and chemical assays, however only c . koseri (C . diversus) was found to produce aerobactin . The concentration of ethylenediamine di(o-hydroxyphenylacetic acid) (EDDA) required to inhibit the growth of individual Citrobacter strains by depleting free iron ranged from 250 micrograms/ml to 100 micrograms/ml . Iron utilization studies of selected citrobacter isolates indicated that hemin and hematin could reverse the effects of iron limitation on growth under iron-stressed conditions (1000 micrograms/ml of EDDA) . Two C . koseri strains grown under iron-restricted conditions showed similar changes in their whole cell protein profiles including induction of high molecular mass proteins (72-83 kDa) which may play a role in iron acquisition under iron-stressed conditions . The collective results support an additional virulence-associated mechanism for C . koseri strains which may help explain the greater pathogenic potential this group has for causing serious extraintestinal disease in humans.

Appl Environ Microbiol, 1996 Apr, 62(4), 1448 - 51
3-Hydroxypropionaldehyde, an inhibitory metabolite of glycerol fermentation to 1,3-propanediol by enterobacterial species; Barbirato F et al.; Glycerol fermentation by Enterobacter agglomerans revealed that both growth and 1,3-propanediol production ceased after consumption of about 430 mM glycerol, irrespective of the initial glycerol content . This phenomenon was assigned to the production of 3-hydroxypropionaldehyde, which was identified by proton nuclear magnetic resonance and which showed a bacteriostatic effect . The accumulation during glycerol fermentation was also observed with two other enterobacterial species, i.e., Klebsiella pneumoniae and Citrobacter freundii.

Jpn J Antibiot, 1996 Apr, 49(4), 338 - 51
{Antibacterial activity of sulopenem, a new parenteral penem antibiotic}; Inoue E et al.; Sulopenem, a new penem antibiotic, was compared with other antibiotics with regard to in vitro antibacterial and bactericidal activities, stabilization against beta-lactamases, and effect on the release of lipopolysaccharide from Gram-negative bacteria . The results are summarized as follows . 1 . Sulopenem showed more potent activities than other antibiotics against both Gram-positive and Gram-negative bacteria except Pseudomonas aeruginosa . 2 . Sulopenem showed potent bactericidal activities (MIC/MBC) against both Gram-positive and Gram-negative bacteria . Time kill studies against Staphylococcus aureus, Escherichia coli, Enterobacter cloacae and Citrobacter freundii showed potent bactericidal activities of sulopenem . 3 . Sulopenem was found to possess a stronger activity than other antibiotics against beta-lactamase-producing strains except P . aeruginosa and Stenotrophomonas maltophilia . 4 . In particular, sulopenem was found to be more stable to the hydrolysis by various beta-lactamases produced by Gram-negative bacteria than any other antibiotics tested . Vmax/Km values of sulopenem were smaller than those of cefotiam for all tested beta-lactamases, which reflected a broad antibacterial spectrum of sulopenem . 5 . E . coli ML4707 exposed to sulopenem and imipenem released less endotoxin than did controls at all concentration ranges tested . In contrast, the strain exposed to ceftazidime at bacteriostatic concentrations released a large amount of endotoxin.

FEMS Immunol Med Microbiol, 1996 Apr, 13(4), 261 - 8
The occurrence of glycine in bacterial lipopolysaccharides; Gamian A et al.; The aminoacyl analysis of endotoxic lipopolysaccharides (LPS) isolated from several bacteria revealed essential amounts of glycine, among the inherent LPS components . Significant amounts of the glycine was detected in lipopolysaccharides isolated from over 30 strains of Escherichia, Salmonella, Hafnia, Citrobacter and Shigella species . Glycine as a single amino acid was found only in a core part of LPS . Molar ratio of glycine in core oligosaccharide fraction ranged from 0.2 to 0.6 per 3 heptoses . The oligosaccharide enriched in glycine was isolated using the HPLC . The amino acid appeared to be terminally located in a core oligosaccharide . The labelling of the lipopolysaccharide cores was achieved when the bacteria were cultivated in the presence of radioactive {14C}glycine . The labelled core oligosaccharide released the radioactivity during treatment with mild alkali or acid (0.1 M NaOH or HCl, 100 degrees C, 4 h) . The radioactivity in SDS-polyacrylamide gel electrophoresis migrated exclusively with LPS . The results indicate that amino acid is an integral constituent of core oligosaccharide in lipopolysaccharide.

Biotechnol Appl Biochem, 1996 Apr, 23 ( Pt 2), 149 - 54
Affinity purification and binding characteristics of Citrobacter freundii AmpR, the transcriptional regulator of the ampC beta-lactamase gene; Roh IK et al.; The transcriptional regulator of the Citrobacter freundii ampC beta-lactamase gene, AmpR, was purified as a single SDS/PAGE-gel band by using various techniques, including DNA-Sepharose 4B affinity chromatography . The purified AmpR consisted of a 32.5 kDa monomer that interacted with three operator sequences: two binding sequences, at positions -75 to -70 and -67 to -51 with respect to the transcriptional start site, were located in the LysR motif (-72 to -60), and the third sequence was at position -43 to -38 . Equilibrium binding studies raise the possibility that the adjacent operator sequence could exert a positive influence on the ability of AmpR to bind to these sites.

J Bacteriol, 1996 Apr, 178(7), 2094 - 101
Isolation, identification, and transcriptional specificity of the heat shock sigma factor sigma32 from Caulobacter crescentus; Wu J et al.; We report the identification of the Caulobacter crescentus heat shock factor sigma32 as a 34-kDa protein that copurifies with the RNA polymerase holoenzyme . The N-terminal amino acid sequence of this protein was determined and used to design a degenerate oligonucleotide as a probe to identify the corresponding gene, rpoH, which encodes a predicted protein with a molecular mass of 33,659 Da . The amino acid sequence of this protein is similar to those of known bacterial heat shock sigma factors of Escherichia coli (41% identity), Pseudomonas aeruginosa (40% identity), and Citrobacter freundii (38% identity) . The isolated C . crescentus gene complements the growth defect of an E . coli rpoH deletion strain at 37 degrees C, and Western blot (immunoblot) analysis confirmed that the gene product is related to the E . coli sigma32 protein . The purified RpoH protein in the presence of RNA polymerase core enzyme specifically recognizes the heat shock-regulated promoter P1 of the C . crescentus dnaK gene, and base pair substitutions in either the -10 or -35 region of this promoter abolish transcription . S1 nuclease mapping indicates that rpoH transcripts originate from two promoters, P1 and P2, under the normal growth conditions . The P2 promoter is similar to the sigma32 promoter consensus, and the P2-specific transcript increases dramatically during heat shock, while the P1-specific transcript remains relatively constant . These results suggest that although the structure and function of C . crescentus sigma32 appear to be very similar to those of its E . coli counterpart, the C . crescentus rpoH gene contains a novel promoter structure and may be positively autoregulated in response to environmental stress.

Mikrobiol Z, 1996 Mar-Apr, 58(2), 3 - 7
{Enterobacteria in areas of water along the Crimean coast}; Puchenkova SG; Characteristic of coliform bacteria in the coastal sea ecosystems is given . A decrease of Escherichia coli indicator value was ascertained and presence of Klebsiella sp . and Citrobacter freundii was revealed, in the sea areas with high level of anthropogenic load . Wide distribution of bacteria of Enterobacter agglomerans kind was determined in the sea substrates and their stability to unfavourable influences was shown.

J Bacteriol, 1996 Mar, 178(5), 1363 - 73
Identification of TonB homologs in the family Enterobacteriaceae and evidence for conservation of TonB-dependent energy transduction complexes; Larsen RA et al.; The transport of Fe(III)-siderophore complexes and vitamin B12 across the outer membrane of Escherichia coli requires the TonB-dependent energy transduction system . A set of murine monoclonal antibodies (MAbs) was generated against an E . coli TrpC-TonB fusion protein to facilitate structure and function studies . In the present study, the epitopes recognized by these MAbs were mapped, and their distribution in gram-negative organisms was examined . Cross-species reactivity patterns obtained against TonB homologs of known sequence were used to refine epitope mapping, with some epitopes ultimately confirmed by inhibition experiments using synthetic polypeptides . Epitopes recognized by this set of MAbs were conserved in TonB homologs for 9 of 12 species in the family Enterobacteriaceae (including E . coli), including previously unidentified TonB homologs in Shigella, Citrobacter, Proteus, and Kluyvera species . These homologs were also detected by a polyclonal alpha-TrpC-TonB serum that additionally recognized the known Yersinia enterocolitica TonB homolog and a putative TonB homolog in Edwardsiella tarda . These antibody preparations failed to detect the known TonB homologs of either Pseudomonas putida or Haemophilus influenzae but did identify potential TonB homologs in several other nonenteric gram-negative species . In vivo chemical cross-linking experiments demonstrated that in addition to TonB, auxiliary components of the TonB-dependent energy transduction system are broadly conserved in members of the family Enterobacteriaceae, suggesting that the TonB system represents a common system for high-affinity active transport across the gram-negative outer membrane.

Antimicrob Agents Chemother, 1996 Feb, 40(2), 454 - 9
Chromosomal beta-lactamase genes of Klebsiella oxytoca are divided into two main groups, blaOXY-1 and blaOXY-2; Fournier B et al.; The chromosomally encoded beta-lactamase gene (blaOXY-2) of the wild-type Klebsiella oxytoca SL911 was cloned and sequenced . Its nucleotide sequence similarity with the previously sequenced K . oxytoca beta-lactamase gene (blaOXY-1) (Y . Arakawa, M . Ohta, N . Kido, M . Mori, H . Ito, T . Komatsu, Y . Fujii, and N . Kato, Antimicrob . Agents Chemother . 33:63-70, 1989) is 87.3%, and its amino acid similarity is 89.7% . This group of K . oxytoca beta-lactamases is related to chromosomal beta-lactamases of Citrobacter diversus, Proteus vulgaris, and Yersinia enterocolitica and to the plasmid-mediated extended-spectrum beta-lactamases MEN-1 and Toho-1 . By colony hybridization with 86 strains susceptible and resistant to aztreonam, isolated in six countries, K . oxytoca beta-lactamase genes hybridized with either a specific blaOXY-1 DNA probe (668 bp) or a blaOXY-2 DNA probe (723 bp) . Thus, beta-lactamase genes could be divided into two groups: blaOXY-1 (47% of the strains) and blaOXY-2 (53% of the strains) . A study of isoelectric points confirmed the great variability reported in the literature . However, the two beta-lactamase groups were each represented by four different pIs: for OXY-2, 5.2, 5.7, 6.4, and 6.8, with the 5.2 form representing 59% of all OXY-2 enzymes, and for OXY-1, 7.1, 7.5, 8.2, and 8.8, with the 7.5 form representing 88% of all OXY-1 enzymes.

Can J Microbiol, 1996 Feb, 42(2), 107 - 14
Distribution of diaminopropane and acetylspermidine in Enterobacteriaceae; Hamana K; Polyamines of 97 strains (60 species) belonging to 18 genera of the family Enterobacteriaceae were determined by high performance liquid chromatographic analysis . In addition to putrescine and cadavarine, diaminopropane was widely distributed in Enterobacteriaceae and almost ubiquitously within Enterobacter, Pantoea, Erwinia, Leminorella, Proteus, Leclercia, Morganella, Klebsiella, Hafnia, Rahnella, Serratia, and Tatumella species and sporadically within Citrobacter, Escherichia, Moellerella, Providencia, Yokenella, and Yersinia species . Histamine was detected in some cultures of Proteus and Morganella . Agmatine was sporadically spread . Heterogeneity in the occurrence of spermidine was observed within the spermidine-containing cultures . Distribution profiles of 18 genera . Acetylated spermidine was found concomitantly in the spermidine-containing cultures . Distribution profiles of diaminopropane, spermidine, and acetylspermidine in Enterobacteriaceae can serve as a chemotaxonomic marker to distinguish this family from other taxa of the gamma subclass of the class Proteobacteria.

J Mol Biol, 1996 Jan 12, 255(1), 176 - 86
Crystal structure of Citrobacter freundii restriction endonuclease Cfr10I at 2.15 A resolution; Bozic D et al.; The X-ray crystal structure of Citrobacter freundii restriction endonuclease Cfr10I has been determined at a resolution of 2.15 A by multiple isomorphous replacement methods and refined to an R-factor of 19.64% . The structure of Cfr10I represents the first structure of a restriction endonuclease recognizing a degenerated nucleotide sequence . Structural comparison of Cfr10I with previously solved structures of other restriction enzymes suggests that recognition of specific sequence occurs through contacts in the major and the minor grooves of DNA . The arrangement of the putative active site residues shows some striking differences from previously described restriction endonucleases and supports a two-metal-ion mechanism of catalysis.

Antibiot Khimioter, 1996, 41(12), 7 - 13
{Characterization of antimicrobial properties of cefpirome}; Sidorenko SV et al.; The results of a multicentre investigation of antibiotic susceptibility in 800 clinical isolates were analyzed . The levels of susceptibility to cefpirome and other antibiotics in gram-negative organisms with inducable production of chromosomal beta-lactamases (Enterobacter, Serratia, Citrobacter and others) were compared and the advantages of cefpirome over other cephalosporins were shown: 89 per cent of the susceptible strains against 58 to 78 per cent . With respect to other microorganisms the advantages of cefpirome were less pronounced.

Postepy Hig Med Dosw, 1996, 50(5), 459 - 72
{Lipopolysaccharides containing sialic acid}; Gamian A; Sialic acids which are important constituents of animal tissue glycoconjugates are also present in antigens of some bacterial strains . Capsular polysaccharides with sialic acid have been extensively studied whereas little is known on lipopolysaccharides which contain sialic acid . The paper presents review of the data concerning structure of bacterial endotoxic lipopolysaccharides . Methodological peculiarities were described . Specially emphasized were endotoxins of Escherichia coli O24, O56, O104, Salmonella toucra O48, Citrobacter freundii O37 and Hafnia alvei strain PCM2386.

Microbiol Immunol, 1996, 40(12), 915 - 21
Re-speciation of the original reference strains of serovars in the Citrobacter freundii (Bethesda-Ballerup group) antigenic scheme of West and edwards; Miki K et al.; The antigenic scheme for the Bethesda-Ballerup group of bacteria established by West and Edwards in 1954 has continued to be applied as a serotyping scheme for Citrobacter freundii . In 1993, however, the classification of the Citrobacter was drastically revised and the species C . freundii redefined by Brenner et al . Accordingly, to judge the propriety to continuously use a single antigenic scheme for the C . freundii complex, the 90 reference strains listed in the antigenic scheme for C . freundii by West and Edwards were characterized phenotypically and specified based on the revised classification . Of these 90 strains, two strains of Hafnia alvei and one of Escherichia coli were found . Among the remaining 87 reference strains, Citrobacter youngae was the predominant species (40 strains), followed by Citrobacter braakii (25 strains), Citrobacter werkmanii (13 strains), and the unnamed Citrobacter genospecies 10 of Brenner et al (six strains) . Citrobacter freundii, as redefined, accounted for only three strains and ranked behind the other four species . No overlapping with most of the 42 O-groups and 82 H-antigens was recognized between species with few exceptions . O-groups 1-9 inclusive, which were estimated to represent more than 90% of the former C.freundii strains, occurred in strains of C . youngae and C . braakii; and all nine strains of O-group 29, formerly known as the Ballerup group, were identified as C . braakii . These findings suggest that further study of the serotyping system is needed for all H2S-producing Citrobacter species.

Ann Pharm Fr, 1996, 54(6), 276 - 9
Screening of in vitro antibacterial activity from Syzygium Guineense (Willd) hydrosoluble dry extract; Tsakala TM et al.; Diarrhoea is one of the most important causes of infant mortality in the world . As modern drugs are expensive or unavailable in developing countries, many people use traditional medicines in Africa for the treatment of several diseases . In our study, we investigated the antibacterial activity of Syzygium Guineense extract in order to assess its activity on some bacterial strains involved in diarrhoeal diseases and to justify its use . The aqueous dry extract was prepared by decoction followed by evaporating to dryness and tested according to dilution method . This extract showed an antibacterial activity against some bacterial strains: Salmonella E., Shigella D., Shigella F., E . Coli., Enterobacter A . It did not show any activity against Citrobacter F., Proteus M., Klebsiella P . Storage conditions (27 degrees C, glass flask) did not affect antibacterial properties of the extract.

Microbios, 1996, 86(349), 205 - 12
A simple method to detect bacteriolytic enzymes produced by enterobacteriaceae; Branca G et al.; The production of bacteriolytic enzymes by Enterobacteriaceae in various growth conditions was investigated . Peptone-based media containing killed Gram-negative cells facilitated detection of bacteriolytic enzyme production in the highest number of species . These belonged to the genera Serratia, Proteus, Morganella and Providencia . In contrast, Escherichia coli, Shigella, Salmonella, Klebsiella, Enterobacter and Citrobacter species did not produce bacteriolytic activities in any of the conditions tested.

FEMS Immunol Med Microbiol, 1996 Jan, 13(1), 1 - 8
Structural and serological studies of lipopolysaccharides of Citrobacter O35 and O38 antigenically related to Salmonella; Kocharova NA et al.; Structural analysis using 13C NMR spectroscopy and methylation showed that lipopolysaccharides (LPSs) of Citrobacter freundii O35 and Salmonella arizonae O59 have structurally identical O-specific polysaccharide chains, and those of C . freundii O38 and Salmonella kentucky differ only in the presence of O-acetyl groups in the former . Serological relationships between the structurally similar LPSs were demonstrated using inhibition of ELISA, rocket immunoelectrophoresis, double gel diffusion, and immunoblotting . The O-acetyl groups present in C . freundii O38 LPS are of little importance for its serological specificity . A cross-reaction was observed in immunoblotting between O-antisera to C . freundii O35 and S . arizonae O59 and a structurally related LPS of Pseudomonas aeruginosa O11a, 11b (Lanyi-Bergan classification).

Antimicrob Agents Chemother, 1996 Jan, 40(1), 221 - 4
Characterization of the plasmidic beta-lactamase CMY-2, which is responsible for cephamycin resistance; Bauernfeind A et al.; The phenotype of Klebsiella pneumoniae HEL-1 indicates a plasmidic cephamycinase gene (blaCMY-2) . Its sequence shows one open reading frame coding for a protein of 381 amino acids . CMY-2 is classified as class C beta-lactamase that is closely related to the plasmidic enzymes BIL-1 and LAT-1 and the chromosomal AmpC of Citrobacter freundii . The blaCMY-2 gene possibly was translocated onto a plasmid of C . freundii which spread to K . pneumoniae.

Ann Urol (Paris), 1996, 30(3), 108 - 11
Citrobacter emphysematous pyelonephritis in a tuberculous kidney caused by citrobacter . A case report in a diabetic patient; Fischer C et al.; Emphysematous pyelonephritis caused by gas-producing bacteria like Escherichia coli or Klebsiella pneumonia is generally observed in female diabetic patients . We report a case in which Citrobacter was the microbiologically documented pathogen . High-dose antibiotic regimen was administered, but nephrectomy was necessary to overcome the life-threatening situation.

Chemotherapy, 1996 Jan-Feb, 42(1), 11 - 20
Comparative evaluation of orally active antibiotics against community-acquired pathogens: results of eight European countries; Cullmann W; In this multicenter study conducted in eight European countries, 13,173 pathogens--all isolated from community-acquired infections in 1992 and 1993--were evaluated for their susceptibility to the following orally active antibiotics: penicillin G, ampicillin, amoxycillin plus clavulanic acid, cefaclor, cefuroxime, cefetamet, doxycycline and erythromycin . Ten centers in Italy, five in Germany, in the Netherlands and Switzerland, four in Greece and Spain, three in Hungary and one in Finland contributed to this study; ready-to-use standardized microtiter panels (Sceptor system, BBL, Heidelberg, Germany) were used throughout all assays . The most frequently encountered species were: Escherichia coli, Proteus mirabilis, Klebsiella pneumoniae and non-typhoid Salmonella spp., Enterobacter cloacae, Streptococcus agalactiae, Haemophilus influenzae, Citrobacter freundii, Staphylococcus pyogenes, Streptococcus pneumoniae, Proteus vulgaris, Moraxella catarrhalis and Shigella spp . The percentage of susceptible isolates was assessed for each of the above-mentioned countries and European-wide with all the data available . For many species, the percentage of resistant isolates did not differ markedly between the countries considered . However, one of the most striking exceptions was the high prevalence of high-level penicillin-G-resistant S . pneumoniae isolates in Hungary and Spain; some of the low-level penicillin-G-resistant strains remained susceptible to cefuroxime, whereas complete cross-resistance occurred for all other beta-lactams studied . The high frequency of ampicillin-resistant H . influenzae isolates in Spain deserves mentioning; this could be attributed mainly to the prevalence of a beta-lactamase, as the addition of clavulanic acid rendered these strains susceptible to ampicillin . The penicillin compounds exhibited the greatest activity against Gram-positive pathogens, whereas cefetamet was the most active agent against Gram-negative pathogens with a well-balanced spectrum of activity.

J Clin Microbiol, 1996 Jan, 34(1), 76 - 9
Outbreak of TEM-24-producing Enterobacter aerogenes in an intensive care unit and dissemination of the extended-spectrum beta-lactamase to other members of the family enterobacteriaceae; Neuwirth C et al.; We report an outbreak of Enterobacter aerogenes in an intensive care unit (ICU) and two medicine departments that produced the extended-spectrum beta-lactamase TEM-24, which was difficult to detect by disk agar diffusion . The strains were compared by DNA restriction fragment length polymorphism after pulsed-field gel electrophoresis following cleavage with XbaI . This typing method indicated that a single strain, first isolated in the ICU, spread throughout the other medical departments as a result of patient transfer . We also observed the transfer in vivo of the plasmid encoding TEM-24 from the strain of Enterobacter aerogenes to different strains of Escherichia coli and Citrobacter freundii in the ICU . It therefore appears that the epidemic involved results from two events: dissemination of one strain of Enterobacter aerogenes and dissemination of the plasmid encoding TEM-24 among various members of the family Enterobacteriaceae.

Acta Clin Belg, 1996, 51(1), 28 - 35
Belgian multicentre study on the in vitro activity of cefepime against gram-negative bacilli; Verbist L et al.; The in vitro activity of cefepime has been compared with that of cefotaxime, ceftazidime, aztreonam, and piperacillin against 1826 Enterobacteriaceae including 537 inducible Enterobacteriaceae (Enterobacter spp., Serratia spp., Citrobacter spp, . Morganella morganii) and 572 non-fermenters, including 401 Pseudomonas aeruginosa and 111 Acinebacter spp . isolated from hospitalized patients in 28 Belgian hospitals . Overall, cefepime was found substantially more active than the third-generation cephalosporins, aztreonam and piperacillin against Enterobacteriaceae species producing inducible type I cephalosporinases . Notably, cefepime remained active against 96% of Enterobacteriaceae resistant to cefotaxime, ceftazidime or aztreonam while it displayed a similar activity against E . coli and the other Enterobacteriaceae . Against non-fermenters, cefepime was found less active than ceftazidime but more than cefotaxime or aztreonam.

Arch Surg, 1996 Jan, 131(1), 95 - 7
Aerobic and anaerobic microbiology of superficial suppurative thrombophlebitis; Brook I et al.; OBJECTIVE: To study the aerobic and anaerobic microbiologic characteristics of superficial suppurative thrombophlebitis . DESIGN: Retrospective review of microbiologic and clinical data . SETTING: Navy Hospital in Bethesda, Md . RESULTS: Sixty-one isolates, 36 aerobic and 25 anaerobic, were isolated from samples obtained from 42 patients . Aerobic bacteria only were found in 26 (62%) patients; anaerobic only, in 11 (26%); and mixed aerobic and anaerobic bacteria, in five (12%) . The predominant aerobic bacteria were Staphylococcus aureus (n = 9), Escherichia coli (n = 7), Pseudomonas aeruginosa (n = 4), and Klebsiella sp (n = 3) . The most frequently recovered anaerobic bacteria were Peptostreptococcus sp (n = 8), Propionibacterium acnes (n = 6), Bacteroides fragilis group (n = 5), Prevotella intermedia (n = 3), and Fusobacterium nucleatum (n = 3) . Propionibacterium acnes and Peptostreptococcus sp were associated with cannula-related superficial suppurative thrombophlebitis; B fragilis and Enterobacteriaceae, with abdominal surgery or pathology; and S aureus and P aeruginosa and Citrobacter sp, with burns . CONCLUSION: These data illustrate the importance of anaerobic bacteria in superficial suppurative thrombophlebitis.

Mol Gen Genet, 1995 Dec 15, 249(5), 474 - 86
SlyA, a regulatory protein from Salmonella typhimurium, induces a haemolytic and pore-forming protein in Escherichia coli; Ludwig A et al.; A chromosomal fragment from Salmonella typhimurium, when cloned in Escherichia coli, generates a haemolytic phenotype . This fragment carries two genes, termed slyA and slyB . The expression of slyA is sufficient for the haemolytic phenotype . The haemolytic activity of E . coli carrying multiple copies of slyA is found mainly in the cytoplasm, with some in the periplasm of cells grown to stationary phase, but overexpression of SlyB, a 15 kDa lipoprotein probably located in the outer membrane, may lead to enhanced, albeit unspecific, release of the haemolytic activity into the medium . Polyclonal antibodies raised against a purified SlyA-HlyA fusion protein identified the overexpressed monomeric 17 kDa SlyA protein mainly in the cytoplasm of E . coli grown to stationary phase, although smaller amounts were also found in the periplasm and even in the culture supernatant . However, the anti-SlyA antibodies reacted with the SlyA protein in a periplasmic fraction that did not contain the haemolytic activity . Conversely, the periplasmic fraction exhibiting haemolytic activity did not contain the 17 kDa SlyA protein . Furthermore, S . typhimurium transformed with multiple copies of the slyA gene did not show a haemolytic phenotype when grown in rich culture media, although the SlyA protein was expressed in amounts similar to those in the recombinant E . coli strain . These results indicate that SlyA is not itself a cytolysin but rather induces in E . coli (but not in S . typhimurium) the synthesis of an uncharacterised, haemolytically active protein which forms pores with a diameter of about 2.6 nm in an artificial lipid bilayer . The SlyA protein thus seems to represent a regulation factor in Salmonella, as is also suggested by the similarity of the SlyA protein to some other bacterial regulatory proteins . slyA- and slyB-related genes were also obtained by PCR from E . coli, Shigella sp . and Citrobacter diversus but not from several other gram-negative bacteria tested.

Am J Infect Control, 1995 Dec, 23(6), 357 - 63
Polymicrobial gram-negative bacteremia associated with saline solution flush used with a needleless intravenous system; Chodoff A et al.; BACKGROUND: During a 2-week period, seven cases of nosocomial polymicrobial gram-negative rod bacteremia occurred on a 39-bed medical and cardiac step-down unit . Combinations of Enterobacter cloacae (seven isolates), Klebsiella pneumoniae (five isolates), and Citrobacter freundii (two isolates) were isolated from blood cultures . METHODS: Concurrent and retrospective chart reviews were used to look for further cases and common exposures . Epidemiologic methods were used to refine determination of common exposure . Restriction enzyme DNA analysis was performed on the isolates . RESULTS: Concurrent and retrospective chart reviews revealed four additional possible cases during the same period . All case patients were exposed, through peripheral saline solution locks or central venous catheters, to saline solution "flush" from a central 0.9% saline solution bag and a needleless dispensing pin . Epidemiologic methods implicated probable extrinsic contamination of a single bag and pin used during a 24-48-hour period (Fisher's Exact Test, p < 0.002) . There were no other common exposures . Restriction enzyme DNA analysis of the isolates further supported a common source for the outbreak . CONCLUSIONS: The introduction of needleless intravascular systems has been embraced for employee protection . Our report is the first to raise the question of patient safety with such systems . This outbreak highlights the inherent risks in rapid introduction of new technologies and points out the delicate balance among patient health, employee safety, and cost containment.

J Chemother, 1995 Dec, 7(6), 509 - 14
Antimicrobial resistance and prevalence of extended spectrum beta-lactamase among clinical isolates of gram-negative bacteria in Riyadh; El-Karsh T et al.; The activity of ciprofloxacin, imipenem and 12 other commonly used antibiotics was evaluated against 106 documented clinical isolates from a medical Intensive Care Unit (ICU) . The resistance rates to ceftriaxone, cefotaxime, aztreonam and ceftazidime were 42, 25, 24 and 21%, respectively . Apart from Pseudomonas aeruginosa, all isolates were sensitive to ciprofloxacin and imipenem . Complete cross resistance among tested beta-lactam groups was uniformly evident in Enterobacter cloacae, Citrobacter freundii and P . aeruginosa . On the other hand, penicillins and second generation cephalosporins showed cross resistance among Escherichia coli and Klebseilla pneumoniae isolates . Induction experiments indicate that 70 and 62% of P . aeruginosa and E . cloacae or C . freundii produce class I cephalosporinase, respectively . Among all tested isolates, plasmid mediated extended spectrum beta-lactamase (ESBL) was detected in one isolate of K . pneumoniae . The plasmid mediated beta-lactamase is transferable and inhibited by beta-lactamase inhibitors . The transconjugates not only expressed resistance to extended spectrum beta-lactams and aztreonam but also toward tested aminoglycoside antibiotics, with the exception of gentamicin . The obtained transconjugates conferred high level resistance to ceftazidime and aztreonam but considerably low resistance to ceftriaxone and cefotaxime . The isoelectric point for the extended-spectrum beta-lactamase is 8.2.

Antimicrob Agents Chemother, 1995 Dec, 39(12), 2787 - 91
In vitro antibacterial properties of T-5575 and T-5578 novel parenteral 2-carboxypenams; Watanabe Y et al.; T-5575 and T-5578, novel 2-carboxypenams in which a carboxyl group has been introduced into the C-2 beta position of the nucleus, were evaluated for their in vitro antibacterial properties . The spectrum of activity of T-5575 was similar to that of aztreonam . However, it showed stronger activities than those of aztreonam against most gram-negative bacteria . T-5575 also showed potent activities against isolates of Enterobacter cloacae, Citrobacter freundii, and Pseudomonas aeruginosa resistant to ceftazidime, with MICs at which 90% of the isolates were inhibited of 0.39, 0.39, and 3.13 microgram/ml, respectively . T-5578 showed moderate levels of activity against gram-negative bacteria, compared with those of T-5575 . Its activity against P . aeruginosa, however, was superior to those of T-5575 and the reference drugs tested . The most characteristic feature of T-5578 was its potent activities against ceftazidime-, imipenem-, and gentamicin-resistant P . aeruginosa isolates, with MICs at which 90% of the isolates were inhibited at 0.39, 3.13, and 3.13 microgram/ml, respectively . These two compounds were unfortunately poorly active against gram-positive bacteria, such as Staphylococcus aureus and streptococci . Both compounds were found to be stable for hydrolysis by various kinds of beta-lactamases and to have low affinities for these enzymes, with Ki values of > 100 microM . These novel penams bound most tightly to penicillin-binding protein 3 of Escherichia coli and P . aeruginosa . These results indicate that T-5575 and T-5578 can be regarded as promising 2-carboxypenams specially targeted against gram-negative pathogens.

Jpn J Antibiot, 1995 Dec, 48(12), 1906 - 19
{Antimicrobial activities of cefepime against clinically isolated strains}; Suzuki Y et al.; In order to evaluate antimicrobial activity of cefepime (CFPM), minimum inhibitory concentrations (MICs) of CFPM and other drugs were determined against clinical isolates that were obtained in 1994 . 1 . CFPM showed a wide antibacterial spectrum against Staphylococcus spp . and glucose non-fermentative Gram-negative rods ((G)NF-GNR) . Antimicrobial activities of CFPM against Staphylococcus spp . were stronger than those of ceftazidime (CAZ) and somewhat stronger than those of cefotaxime (CTX), and antimicrobial activity of CFPM against Pseudomonas aeruginosa was same as that of CAZ . 2 . Antimicrobial activities of CFPM against almost all of Enterobacteriaceae were stronger than those of CAZ and CTX . And CFPM showed strong antimicrobial activities against CAZ-resistant Escherichia coli, Citrobacter freundii and Enterobacter spp . 3 . Antimicrobial activities of CFPM were weaker than those of CAZ against some of strains of Klebsiella oxytoca, beta-lactamase high producing strains of Moraxella subgenus Branhamella catarrhalis and than those of CTX against beta-lactamase high producing strains of Prevotella spp . 4 . The feature of new cephems was demonstrated in that CFPM had wider antibacterial spectrum than cephems of previous genenations against Staphylococcus spp . and (G)NF-GNR and CFPM showed strong antimicrobial activities against almost all of oxacephem-resistant Enterobacteriaceae.

J Appl Bacteriol, 1995 Dec, 79(6), 635 - 9
Detection of Escherichia coli in potable water using direct impedance technology; Colquhoun KO et al.; Direct impedance measurement utilizing a medium previously described as being specific for Escherichia coli and which contains trimethylamine N-oxide (TMAO) and glucuronic acid was used to detect E . coli in water samples . The system was compared with the Colilert presence/absence test and the United Kingdom standard membrane filtration technique using membrane lauryl sulphate broth . The impedance method correlated well with both the traditional membrane method (93%) and the Colilert method (93.95%) for a number of different water types . No interference from Citrobacter spp . (as reported in previous studies) was detected in this study although some Salmonella spp . did give false-positive results . The data presented here suggest that the use of direct impedance may offer an alternative to conventional methods for the detection of E . coli in water.

Clin Infect Dis, 1995 Nov, 21(5), 1107 - 13
The relationship between antecedent antibiotic use and resistance to extended-spectrum cephalosporins in group I beta-lactamase-producing organisms; Jacobson KL et al.; Gram-negative pathogens are increasingly resistant to extended-spectrum cephalosporins (ESCs) . Using a prospective, case-controlled observational study, we examined the prevalence and the risk factors for development of resistance to ESCs in group I beta-lactamase-producing organisms . Of the 386 isolates of Enterobacter species, Pseudomonas aeruginosa, Citrobacter species, and Serratia marsescens from 340 consecutive patients, 70 (18.1%) were resistant to ESCs; the highest rates of resistance were found among Citrobacter freundii (40.9%), Enterobacter cloacae (31.1%), and Enterobacter aerogenes isolates (18.9%) . Patients' prior antibiotic use and the mean number of antibiotics used were significantly greater in association with resistant vs . susceptible isolates . Resistance was associated with prior use of ceftizoxime or cefotaxime (P = .008), ceftazidime (P = .004), and piperacillin (P = .001) . Other antibiotics were not associated with resistance . Resistance was less frequent in patients receiving ESCs and an aminoglycoside . We conclude that prior use of ESCs is associated with the isolation of resistant group I beta-lactamase-producing organisms . Concomitant use of an aminoglycoside may decrease this risk.

Antimicrob Agents Chemother, 1995 Nov, 39(11), 2494 - 8
DNA sequence differences of ampD mutants of Citrobacter freundii; Stapleton P et al.; Three groups of mutants with increased levels of beta-lactamase synthesis were selected from Citrobacter freundii 382010 by beta-lactam antibiotics at concentrations just above the MIC . Uninduced cultures of the hyperinducible group had 3- to 5-fold more beta-lactamase activity than the parent strain, with one mutant (termed type b) expressing 19 times the activity of the parent strain; the partially derepressed group had a relative 55-fold increase, while fully derepressed strains exhibited a 460-fold increase . Upon induction by growth in the presence of cefoxitin (32 micrograms/ml) for 2 h, the hyperinducible and derepressed groups had similar relative beta-lactamase activities of 650 and 725, respectively . Induction of beta-lactamase activity from partially derepressed mutants resulted in a relative activity of only 240 . The ampD gene including its promoter region was amplified from the parent strain and the mutant strains by PCR . The sequence of ampD from the parent strain showed only three nucleotide changes from a previously published sequence, none of which resulted in a change to the deduced amino acid sequence . Hyperinducible mutant strains of type a had an amino acid change of either a tryptophan in codon 95 to an arginine (Trp-95-->Arg) (three mutants) or Ala-158-->Asp (one mutant) . The hyperinducible type b strain had the change Tyr-102-->Asp . The derepressed strains had the following changes: Val-33-->Gly (one mutant), Asp-164-->Glu (one mutant), and Trp-95-->termination codon (two mutants) . We infer that the amino acid changes in the hyperinducible mutants result in altered AmpD activity, whereas, in contrast, they lead to an inactive protein in derepressed mutants . No nucleotide differences were found in the ampD gene from partially derepressed strains.

FEMS Microbiol Lett, 1995 Nov 1, 133(1-2), 35 - 9
Cloning and nucleotide sequence of a gene upstream of the eaeA gene of enterohemorrhagic Escherichia coli O157:H7; Zhao S et al.; A DNA segment located immediately upstream of the eaeA gene of enterohemorrhagic Escherichia coli O157:H7 strain HA1 was cloned and sequenced . This segment contained an open reading frame encoding a predicted protein of 156 amino acids . A database search identified similar open reading frames upstream of the eaeA gene in two other bacterial pathogens, i.e . enteropathogenic E . coli and Citrobacter freundii . The predicted amino acid sequence of the enterohemorrhagic E . coli protein shared 96.8% and 94.2% identity with the enteropathogenic E . coli and C . freundii sequences, respectively . Because the open reading frame is located within the locus of enterocyte effacement region of the E . coli chromosome, a 'hot spot' for insertion of virulence factor genes, and shares high sequence homology with attaching and effacing EPEC and C . freundii, this protein may be associated with pathogenicity of E . coli O157:H7.

Infect Control Hosp Epidemiol, 1995 Oct, 16(10), 564 - 9
Vertical transmission of Citrobacter diversus documented by DNA fingerprinting; Harvey BS et al.; OBJECTIVE: To confirm the vertical transmission of Citrobacter diversus from a mother to her infant and to evaluate the epidemiologic usefulness of a new automated procedure for analysis of polymerase chain reaction (PCR)-generated DNA fingerprints . DESIGN: Repetitive element-based PCR (rep-PCR) analysis of C diversus isolates from the blood and amniotic fluid of a mother and the blood of her infant was performed . Unrelated C diversus isolates also were characterized and compared with the isolates from mother and infant . DNA fingerprints were generated by gel electrophoresis of PCR products derived from either unlabeled standard repetitive sequence-based oligonucleotide primers or fluorescent primers . The standard rep-PCR fingerprints were analyzed by visual inspection . The fluorescent primers were used in fluorophore-enhanced rep-PCR (FERP), and the FERP DNA fingerprints were analyzed by an Applied BioSystems (ABI) Model 373A laser scanning unit equipped with Genescan 672 software (Applied Biosystems, Inc, Foster City, CA) . SETTING AND PATIENTS: A mother and her newborn infant, both with invasive disease due to C diversus, in an urban tertiary-care hospital . RESULTS: The DNA fingerprints of the maternal blood, amniotic fluid, and infant blood isolates of C diversus were identical by both visual inspection of ethidium bromide-stained agarose gels and computer-aided analysis of FERP patterns . These strains appeared to differ from all but one control isolate, which had been collected 7 years earlier in the same city in which the infant was born . CONCLUSIONS: Vertical transmission of C diversus from mother to infant can occur in utero . Automated analysis of rep-PCR-generated DNA fingerprints derived using fluorescent primers is an objective means for comparing isolates of C diversus and in all likelihood would be useful for other species of bacteria that possess repetitive elements.

Microbiology, 1995 Oct, 141 ( Pt 10), 2433 - 41
Phosphatase production and activity in Citrobacter freundii and a naturally occurring, heavy-metal-accumulating Citrobacter sp; Montgomery DM et al.; The ability of a naturally occurring Citrobacter sp . to accumulate cadmium has been attributed to cellular precipitation of CdHPO4, utilizing HPO4(2-) liberated via the activity of an overproduced, Cd-resistant acid-type phosphatase . Phosphatase production and heavy metal accumulation by batch cultures of this strain (N14) and a phosphatase-deficient mutant were compared with two reference strains of Citrobacter freundii . Only strain N14 expressed a high level of acid phosphatase and accumulated lanthanum and uranyl ion enzymically . Acid phosphatase is regulated via carbon-starvation; although the C . freundii strains overexpressed phosphatase activity in carbon-limiting continuous culture, this was approximately 20-fold less than the activity of strain N14 grown similarly . Citrobacter strain N14 was originally isolated from a metal-contaminated soil environment; phosphatase overproduction and metal accumulation were postulated as a detoxification mechanism . However, application of Cd-stress, and enrichment for Cd-resistant C . freundii ('training'), reduced the phosphatase activity of this organism by about 50% as compared to Cd-unstressed cultures . The acid phosphatase of C . freundii and Citrobacter N14 had a similar pattern of resistance to some diagnostic reagents . The enzyme of the latter is similar to the PhoN acid phosphatase of Salmonella typhimurium described by other workers; the results are discussed with respect to the known phosphatases of the enterobacteria.

Lett Appl Microbiol, 1995 Oct, 21(4), 249 - 51
A comparison of immunomagnetic separation plus enrichment with conventional Salmonella culture in the examination of raw sausages; Coleman DJ et al.; Immunomagnetic separation with additional enrichment was used in conjunction with improved selective media to improve the isolation of salmonellae from raw sausages . The isolation rate achieved was almost double that of conventional culture with no increase in processing time . The selective media gave an overall specificity of approximately 74%; all false-positive pickoffs being identified as Citrobacter freundii . It is believed that this method represents a significant advance in the isolation of salmonellae from foods, although the ideal media both for enrichment and selection have yet to be found.

Biochemistry, 1995 Sep 26, 34(38), 12276 - 83
Site-directed mutagenesis of tyrosine-71 to phenylalanine in Citrobacter freundii tyrosine phenol-lyase: evidence for dual roles of tyrosine-71 as a general acid catalyst in the reaction mechanism and in cofactor binding; Chen HY et al.; Tyr71 is an invariant residue in all known sequences of tyrosine phenol-lyase (TPL) . The substitution of Tyr71 in TPL by phenylalanine results in a mutant Y71F TPL with no detectable activity (greater than 3 x 10(5)-fold reduction) for beta-elimination of L-tyrosine . Y71F TPL can react with S-alkylcysteines, but these substrates exhibit kcat values reduced by 10(3)-10(4)-fold, while the kcat/Km values are reduced by 10(2)-10(3)-fold, compared to wild-type TPL . However, for substrates with good leaving groups (S-(o-nitrophenyl)-L-cysteine,beta-chloro-L-alanine, and O-benzoyl-L-serine), Y71F TPL exhibits kcat values 1.85-7% those of wild-type TPL . Y71F TPL forms very stable quinonoid complexes with strong absorbance at 502 nm from L-phenylalanine, tyrosines (L-tyrosine, 3-fluoro-L-tyrosine, and {alpha-2H}-3-fluoro-L-tyrosine), and S-alkylcysteines (S-methyl-L-cysteine, S-ethyl-L-cysteine, and S-benzyl-L-cysteine) . The time courses of the formation of quinonoid intermediates in these reactions are biphasic . The slow phase shows a dependence on concentration of PLP and is due to the cofactor binding steps, while the fast phase is due to the amino acid alpha-deprotonation and reprotonation steps . The rate constants for the fast phase of the reactions of Y71F TPL with L-phenylalanine and S-methylcysteine are similar to those for alpha-deprotonation or reprotonation steps in the reactions of wild-type TPL . The PLP binding constant of Y71F TPL is estimated to be 1 mM by spectrophotometric titration, compared to 0.6 microM for wild-type TPL.(ABSTRACT TRUNCATED AT 250 WORDS)

J Antimicrob Chemother, 1995 Sep, 36(3), 537 - 43
Escherichia hermannii: susceptibility pattern to beta-lactams and production of beta-lactamase; Fitoussi F et al.; The susceptibility pattern of Escherichia hermannii, although closely related to Escherichia coli according to its biochemical patterns, was clearly distinguishable by its susceptibility to beta-lactams by both diffusion and dilution methods from E . coli penicillinase producing or non-producing strains, Citrobacter diversus, Klebsiella pneumoniae, and Klebsiella oxytoca . Beta-lactamase clavulanate-sensitive activity was localized with various isoelectric points from 7.0 to 8.5 . No cross hybridization with DNA intragenic probes (blaTEM, blaSHV, blaCARB and blaOXY) was observed by dot blot procedure.

J Antimicrob Chemother, 1995 Sep, 36(3), 483 - 96
The ability of beta-lactam antibiotics to select mutants with derepressed beta-lactamase synthesis from Citrobacter freundii; Stapleton P et al.; The ability of beta-lactam compounds to induce synthesis of the class C beta-lactamase of Citrobacter freundii was assessed both directly and indirectly, the latter by measurement of the different susceptibilities of strains with inducible and depressed beta-lactamase synthesis and those of a beta-lactamase-negative strain . Most beta-lactam compounds were poor inducers but ampicillin, amoxycillin and cephalexin were moderately good inducers and cefoxitin, imipenem and meropenem were strong inducers . The ability of the compounds to select mutants in which the synthesis of the beta-lactamase was derepressed was also assessed . Imipenem and temocillin, which had a high degree of stability to the beta-lactamase failed to select such mutants or were very poor selectors . In contrast, most other compounds showed considerable lability to the beta-lactamase and readily selected derepressed mutants, whether they were strong inducers of beta-lactamase synthesis (for example cefoxitin) or poor inducers (cefotaxime, ceftazidime and many other compounds) . Thus it appears that lability to the beta-lactamase is a more important factor in determining whether or not a compound will select derepressed mutants than the power of the compound as an inducer of beta-lactamase synthesis, since induction results in the production of less beta-lactamase than derepression as a result of mutation.

Lijec Vjesn, 1995 Sep-Oct, 117(9-10), 249 - 53
{Clinical and laboratory significance of inducible beta-lactamases}; Bedenic B et al.; Many species of bacteria have inducible expression of beta-lactamase and the enzyme production in these bacteria is normally held at a low level by a repressor mechanism, but in the presence of a beta-lactamase inducer this repression is lifted and enzyme production is greatly increased . Inducible synthesis of beta-lactamase was first described for the gram-positive organism Bacillus licheniformis . After that inducible expression of beta-lactamase was discovered in gram-negative bacteria like Enterobacter cloacae, Citrobacter freundii, Pseudomonas aeruginosa, Morganella morganii, Proteus vulgaris, Serratia spp and Aeromonas spp . Inducible beta-lactamases belong into class I according to Richmond and Sykes . They are chromosomally mediated cephalosporinases . beta-lactam antibiotics differ in their inducing power . Cefoxitin and imipenem are among the strongest inducers . Induction of beta-lactamase caused by these substances can lead to antagonism with other beta-lactam antibiotics if they are used in combination . The most important clinical problem connected with inducible beta-lactamases is the emergence of multiple resistant strains which are associated with therapeutic failures . It is important to distinguish induction from derepression . Induction is a temporary phenomenon in which an inducer interacts with a functional AmpD protein, which consequently prevents complexing of the AmpD and ampR proteins . In contrast, genetic derepression is permanent and results from synthesis of a defective AmpD protein unable to complex with the AmpR protein.

J Bacteriol, 1995 Sep, 177(18), 5379 - 80
Cloning, sequencing, and expression of the araE gene of Klebsiella oxytoca 8017, which encodes arabinose-H+ symport activity; Shatwell KP et al.; Southern analysis of the genomic DNA from species of the family Enterobacteriaceae, using a probe derived from the Escherichia coli araE gene, which encodes an arabinose-H+ symporter, detected araE in Salmonella, Citrobacter, Klebsiella, and Enterobacter spp . The Klebsiella oxytoca araE gene was cloned, sequenced, and expressed to compare its properties with those of araE from E . coli.

J Antibiot (Tokyo), 1995 Sep, 48(9), 1027 - 33
Effects of tazobactam on the frequency of the emergence of resistant strains from Enterobacter cloacae, Citrobacter freundii, and Proteus vulgaris (beta-lactamase derepressed mutants); Higashitani F et al.; When Enterobacter cloacae, Citrobacter freundii, and Proteus vulgaris were treated with piperacillin (PIPC) in combination with tazobactam (TAZ), the in vitro frequency of emergence of resistant strains (beta-lactamase producing mutants) was lower than with PIPC or ceftazidime (CAZ) treated bacteria . In a mouse intraperitoneal infection model caused by E . cloacae, beta-lactamase derepressed mutants were detected following therapy with PIPC or CAZ, although no derepressed mutants were detected after treatment with PIPC in combination with TAZ . This suppression of the selection of derepressed mutants, which produce large amounts of beta-lactamases, by the combination of TAZ and PIPC suggests that the combination delays the increase of resistant mutants compared with PIPC alone.

Jpn J Antibiot, 1995 Sep, 48(9), 1174 - 263
{Comparative studies on activities of antimicrobial agents against causative organisms isolated from urinary tract infections (1989) . III . Secular changes in susceptibility}; Kumamoto Y et al.; Susceptibilities of Citrobacter spp., Enterobacter spp., Escherichia coli, Klebsiella spp., Proteus mirabilis, Pseudomonas aeruginosa, Serratia spp . isolated from patients with urinary tract infections (UTIs) in 10 hospitals during June 1989 to May 1990 to various antimicrobial agents were compared with those in the same period of previous years according to a classification, uncomplicated UTIs, complicated UTIs without indwelling catheter, and complicated UTIs with indwelling catheter . As for Citrobacter spp., P . mirabilis and Serratia spp., which were detected very few in 1989, their susceptibilities were not observed an obvious change . As for Enterobacter spp., the susceptible strains to flomoxef, cefixime, cefuzonam and ceftazidime increased in complicated UTIs with indwelling catheter . The susceptibilities of E . coli to penicillins increased slightly in complicated UTIs with indwelling catheter . Against Klebsiella spp., a good activity of minocycline or cephems was found . The susceptibilities of P . aeruginosa to ciprofloxacin and new quinolones increased in uncomplicated UTIs . These data should be considered in clinical treatment of various urinary tract infections.

Epidemiol Infect, 1995 Aug, 115(1), 61 - 70
Control of infection with multiple antibiotic resistant bacteria in a hospital renal unit: the value of plasmid characterization; Reed CS et al.; An outbreak of infections due to multiple antibiotic-resistant bacteria took place over a period of approximately 18 months in a renal unit . Strains of Escherichia coli, Enterobacter aerogenes, Klebsiella pneumoniae, Citrobacter spp . and Pseudomonas spp . were involved, and a variety of antibiotic resistances was encountered . Closely related plasmids encoding resistance to aztreonam, ceftazidime and piperacillin, possibly derived from an archetypal plasmid of 105 kb were found in the majority of isolates examined . After limiting the use of aztreonam the incidence of new patient isolates of multiple-resistant organisms was greatly reduced . This study demonstrated how molecular studies can contribute to the control of an outbreak situation in a hospital unit by providing an impetus to reduce the use of specific antibiotics.

J Bacteriol, 1995 Aug, 177(15), 4392 - 401
Biochemical and molecular characterization of the oxidative branch of glycerol utilization by Citrobacter freundii; Daniel R et al.; Glycerol dehydrogenase (EC 1.1.1.6) and dihydroxyacetone kinase (EC 2.7.1.29) were purified from Citrobacter freundii . The dehydrogenase is a hexamer of a polypeptide of 43,000 Da . The enzyme exhibited a rather broad substrate specificity, but glycerol was the preferred substrate in the physiological direction . The apparent Kms of the enzyme for glycerol and NAD+ were 1.27 mM and 57 microM, respectively . The kinase is a dimer of a polypeptide of 57,000 Da . The enzyme was highly specific for the substrates dihydroxyacetone and ATP; the apparent Kms were 30 and 70 microM, respectively . The DNA region which contained the genes encoding glycerol dehydrogenase (dhaD) and dihydroxyacetone kinase (dhaK) was cloned and sequenced . Both genes were identified by N-terminal sequence comparison . The deduced dhaD gene product (365 amino acids) exhibited high degrees of homology to glycerol dehydrogenases from other organisms and less homology to type III alcohol dehydrogenases, whereas the dhaK gene product (552 amino acids) revealed no significant homology to any other protein in the databases . A large gene (dhaR) of 1,929 bp was found downstream from dhaD . The deduced gene product (641 amino acids) showed significant similarities to members of the sigma 54 bacterial enhancer-binding protein family.

J Clin Microbiol, 1995 Aug, 33(8), 2064 - 8
Genetic and biochemical characterization of Citrobacter rodentium sp . nov; Schauer DB et al.; An unusual bacterial pathogen of laboratory mice has been previously classified as an atypical biotype of Citrobacter freundii . Designated C . freundii biotype 4280, this bacterium is the etiologic agent of transmissible murine clonic hyperplasia . An eaeA gene has been shown to be present in this organism and to be necessary for virulence in laboratory mice . However, other biotypes of C . freundii lack DNA homology with the eaeA gene . Because of the recent reclassification in which five named species and three unnamed species, all previously considered C . freundii, were described, we determined the taxonomic status of C . freundii biotype 4280 . With a battery of biochemical tests and DNA relatedness studies, three isolates of C . freundii biotype 4280 were shown to be members of an unnamed Citrobacter species, designated species 9 . In total, six isolates of Citrobacter species 9, but none of the type strains of the other eight named species or of the two remaining unnamed species of Citrobacter, were shown to possess DNA homology with both the eaeA and the eaeB genes . Species 9 was named Citrobacter rodentium sp . nov.

Diagn Microbiol Infect Dis, 1995 Jul, 22(3), 261 - 6
Beta-D-Glucuronidase activity among prototrophic and auxotrophic variants of Escherichia coli and other Enterobacteriaceae commonly implicated in urinary tract infections; Tapsall JW et al.; 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.

Pediatr Infect Dis J, 1995 Jul, 14(7 Suppl), S77 - 83
Ceftibuten: a review of antimicrobial activity, spectrum and other microbiologic features; Jones RN; Ceftibuten is a new, orally administered cephalosporin with exceptional beta-lactamase stability and potency against commonly isolated Gram-negative pathogens . More than 90% of recent Enterobacteriaceae clinical isolates were inhibited by < or = 8 micrograms/ml of ceftibuten . In only five enteric species (Citrobacter freundii, Enterobacter aerogenes, Enterobacter cloacae, Morganella morganii, Serratia marcescens) were more than 15% of strains resistant (minimal inhibitory concentrations (MIC, with percent of strains inhibited in subscript numbers) > 16 micrograms/ml) to ceftibuten . Enteritis-producing bacteria such as Salmonella, Shigella, Escherichia coli and Yersinia were very ceftibuten-susceptible (MIC50 < or = 0.13 microgram/ml) . Fastidious Gram-negative species causing respiratory tract or genital infections had very low ceftibuten MICs, including beta-lactamase-positive Haemophilus influenzae (MIC90 0.06 to 2 micrograms/ml), Moraxella catarrhalis (MIC90 0.25 to 4 micrograms/ml), and Neisseria gonorrhoeae (MIC90 0.015 to 0.5 microgram/ml) . Beta-hemolytic streptococci and penicillin-susceptible pneumococci were also inhibited by ceftibuten . Staphylococci, enterococci, Pseudomonas species and Gram-negative anaerobic bacteria were generally resistant to ceftibuten . Ceftibuten has demonstrated bactericidal activity against susceptible pathogens, has high affinity for several lethal penicillin-binding proteins and possesses stability to common plasmid- or chromosomal-mediated beta-lactamases, including those enzymes that hydrolyze parenteral third generation cephalosporins . The microbiologic features for ceftibuten indicate its clinical potential as chemotherapy for community-acquired respiratory tract infections.

J Chemother, 1995 Jun, 7 Suppl 2, 31 - 44
The changing nature of aminoglycoside resistance mechanisms and the role of isepamicin--a new broad-spectrum aminoglycoside . The Aminoglycoside Resistance Study Groups; Miller GH et al.; Aminoglycoside resistance mechanisms from recent studies were compared with those found in earlier studies in the USA and Europe for three pathogen groups . Among Citrobacter-Enterobacter-Klebsiella, four single mechanisms (AAc(3)-II, AAC(3)-I, ANT(2")-I and AAC(6')-I were found in all studies, but the most recent studies showed a significant increase in combinations of AAC(6')-I with the other common mechanisms . Since AAC(6')-I confers resistance to tobramycin, netilmicin and amikacin, combinations of it with the other gentamicin modifying enzymes conferred broad-spectrum resistance to all clinically available aminoglycosides except isepamicin . Similar changes occurred in Escherichia-Morganella-Proteus-Salmonella-Shigella except that the frequency of combinations was much lower and two additional single mechanisms - AAC(3)-IV and permeability - were also found frequently . Among aminoglycoside-resistant Pseudomonas, three mechanisms, AAC(6')-II, ANT(2")-I and permeability, were always common and remained common . However, combinations of the three mechanisms with each other and with other mechanisms were more common in the recent surveys . Different genes which produce different proteins with the same aminoglycoside-modifying activity are now known . The results of hybridisation studies with two aac(3)-I, 2 aac(6')-II and 4 aac(6')-I gene probes are presented . The most commonly occurring genes were: aac(3)-Ia, aac(3)-IIa, aac(6')-IIa, aac(6')-Ib and, in Serratia, aac(6')-Ic . The activity of isepamicin against amikacin resistant strain which produce AAC(6')-I can be related to differences in the structure of these two similar aminoglycosides at Position 3" . Amikacin may form a stable complex with AAC(6')-I enzymes via binding interaction at Position 3 and 3" . Isepamicin, which has a secondary amino group at Position 3", may only be able to interact at Position 3 and enzyme-isepamicin complexes are likely to be less stable.

Appl Environ Microbiol, 1995 Jun, 61(6), 2335 - 40
Effect of oxytetracycline-medicated feed on antibiotic resistance of gram-negative bacteria in catfish ponds; DePaola A et al.; The effect of oxytetracycline-medicated feeds on antibiotic resistance in gram-negative bacteria from fish intestines and water in catfish ponds was investigated . In experiments in the fall and spring, using ponds with no previous history of antibiotic usage, percentages of tetracycline-resistant bacteria in catfish intestines obtained from medicated ponds increased significantly after 10 days of treatment . In the fall, resistance of the intestinal and aquatic bacteria returned to pretreatment levels within 21 days after treatment . In the spring, resistance declined after treatment but remained higher than pretreatment levels for at least 21 days in intestinal bacteria and for 5 months in aquatic bacteria . Plesiomonas shigelloides, Aeromonas hydrophila, and Citrobacter freundii were isolated frequently in both spring and fall; Escherichia coli, Klebsiella pneumoniae, Edwardsiella tarda, and Enterobacter spp . were isolated primarily in the spring . Oxytetracycline treatment did not affect the distribution of bacterial species in the fall but may have accelerated a shift toward greater prevalence of members of the family Enterobacteriaceae in the spring . Multiple antibiotic resistance did not appear to be elicited by oxytetracycline treatment.

Epidemiol Infect, 1995 Jun, 114(3), 441 - 50
Verotoxinogenic Citrobacter freundii associated with severe gastroenteritis and cases of haemolytic uraemic syndrome in a nursery school: green butter as the infection source; Tschape H et al.; A summer outbreak of severe gastroenteritis followed by haemolytic uraemic syndrome (HUS) and thrombotic thrombocytopenic purpura in a nursery school and kindergarten is described . Sandwiches prepared with green butter made with contaminated parsley were the likely vehicle of infection . The parsley originated from an organic garden in which manure of pig origin was used instead of artificial fertilizers . Clonally identical verotoxinogenic Citrobacter freundii were found as causative agents of HUS and gastroenteritis and were also detected on the parsley.

Pediatrics, 1995 Jun, 95(6), 803 - 6
No lumbar puncture in the evaluation for early neonatal sepsis: will meningitis be missed?
Wiswell TE, Baumgart S, Gannon CM, Spitzer AR.
OBJECTIVE . We performed this investigation to assess whether selective approaches to performing lumbar puncture (LP) in the early neonatal period will result in a missed or delayed diagnosis of bacterial meningitis . DESIGN . A retrospective review was conducted of the medical records of all neonates born in US Army hospitals from 1988 through 1992 who developed culture-positive meningitis during the first 72 hours of life . RESULTS . In total, 169,849 infants were born during the 5-year study period . The incidence of meningitis in the first 72 hours of life was 0.25 per 1000 live births . Forty-three infants had organisms isolated from their cerebrospinal fluid (30, group B streptococcus; 10, Escherichia coli; 1, Listeria monocytogenes; 1, Streptococcus pneumoniae; and 1, Citrobacter diversus) . The median age of infants at evaluation was 12 hours, and the mean gestational age was 38.8 weeks (7 < 37 weeks), whereas mean birth weight was 3163 g (7 < 2500 g) . If we had used currently advocated selective criteria as the basis for not performing an LP, the diagnosis of bacterial meningitis would have been missed or delayed in 16 of 43 infants (37%): 5 infants born prematurely with suspected respiratory distress syndrome, 3 asymptomatic infants born at term with positive blood cultures, and 8 infants born at term with no central nervous system symptoms and negative blood cultures . CONCLUSIONS . If LPs are omitted as part of the early neonatal sepsis evaluation, the diagnosis of bacterial meningitis occasionally will be delayed or missed completely.

Microbiology, 1995 May, 141 ( Pt 5), 1085 - 92
The signal transducer encoded by ampG is essential for induction of chromosomal AmpC beta-lactamase in Escherichia coli by beta-lactam antibiotics and 'unspecific' inducers; Schmidt H et al.; Chemical mutagenesis of the AmpC beta-lactamase-hyperinducible Escherichia coli strain SN0301/pNu305 carrying the cloned ampC and ampR genes from Citrobacter freundii OS60 gave four independent mutants in which beta-lactamase was no longer inducible, or was inducible only to a low level, by beta-lactam antibiotics . The genes ampC, ampR, ampD and ampE, which were essential for beta-lactamase induction, were functional in these mutants . In all four mutants, the sites of mutation were mapped to 9.9 min on the E . coli chromosome . Complementation with wild-type ampG restored inducibility of beta-lactamase to wild-type levels . The nucleotide sequence of all four mutant ampG alleles (ampG1, ampG3, ampG4 and ampG5) was determined . In three of the mutants, a single base exchange led to an amino acid change from glycine to aspartate at different sites in the deduced amino acid sequence . In the fourth mutant (ampG4), with low-level inducibility, the nucleotide sequence was identical to wild-type ampG . Spontaneous back-mutation of the chromosomal ampG1 mutant resulted in restoration of wild-type inducibility and a return to the wild-type ampG sequence . Unspecific induction by components of the growth medium was also dependent on intact ampG function.

Epidemiol Mikrobiol Imunol, 1995 May, 44(2), 57 - 64
{Biochemical indicators of Citrobacter sedlakii}; Aldova E et al.; The authors describe eight strains identified biochemically as a new species of C . sedlakii in clinical material, surface water and the cutting surface of a melon . The majority of strains was isolated in the Czech and Slovak Republic.

Res Microbiol, 1995 May, 146(4), 279 - 90
Taxonomic diversity of anaerobic glycerol dissimilation in the Enterobacteriaceae; Bouvet OM et al.; A total of 1,123 strains representing 128 taxa in the Enterobacteriaceae (named species or subspecies and genomic species) were screened for the presence of glycerol dehydrogenases and 1,3-propanediol dehydrogenase . Only eight taxa, Citrobacter freundii sensu stricto, C . youngae, C . braakii, C . werkmanii, Citrobacter genomospecies 10 and 11, Enterobacter gergoviae and Klebsiella pneumoniae subsp . pneumoniae could grow fermentatively on glycerol and possessed both glycerol dehydrogenase type I (induced by glycerol and dihydroxyacetone) and 1,3-propanediol dehydrogenase which are typical enzymes of the anaerobic glycerol dissimilation pathway . Six other species, C . koseri, E . aerogenes, E . intermedium, K . oxytoca, K . planticola and K . terrigena could not grow fermentatively on glycerol and possessed a glycerol dehydrogenase type I but no 1,3-propanediol dehydrogenase . Other glycerol dehydrogenases types were found: type II (induced by glycerol and hydroxyacetone), type III (induced by glycerol only) and type IV (induced by hydroxyacetone only) . They were widely distributed among the Enterobacteriaceae . Classification and identification may take advantage of tests exploring the dissimilation of glycerol.

J Appl Bacteriol, 1995 May, 78(5), 507 - 20
Development and evaluation of two novel oligonucleotide probes based on 16S rRNA sequence for the identification of Salmonella in foods; Lin CK et al.; DNA sequence in the V3 to V6 region of the 16S rRNA gene of Salmonella enteritidis was determined . By comparison of this sequence with those of Escherichia coli and Proteus vulgaris obtained from GenBank/EMBL database, three oligonucleotides termed as 16S I, 16S II and 16S III were synthesized . Hybridization of these oligonucleotides with 325 Salmonella isolates and some non-Salmonella isolates including the Salmonella closely related species of the family of Enterobacteriaceae showed that 16S II could not be used as a Salmonella specific-probe . 16S I and 16S III hybridized with all the Salmonella isolates tested, the former also hybridizing with Citrobacter spp . and the latter hybridizing with Klebsiella pneumoniae as well as Serratia marcescens . Since enrichment of the target cells in food samples was usually required prior to the DNA hybridization assay, the interference from those non-Salmonella isolates could be prevented by enrichment by culturing in lactose-combined tetrathionate (CTET) broth followed by Gram-negative (GN) broth at 37 degrees C and/or 43 degrees C . Such a culture step could inhibit the growth of Klebsiella spp., Ser . marcescens and/or Citrobacter spp . and allowed the specific detection of Salmonella.

Eur J Biochem, 1995 Apr 15, 229(2), 540 - 9
Site-directed mutagenesis of His343-->Ala in Citrobacter freundii tyrosine phenol-lyase . Effects on the kinetic mechanism and rate-determining step; Chen H et al.; His343 in Citrobacter freundii tyrosine phenol-lyase is conserved in all known sequences of both tyrosine phenol-lyase and tryptophan indole-lyase; it is located near the active-site Lys257 in C . freundii tyrosine phenol-lyase {Antson, A . A., Demidkina, T . V., Gollnick, P., Danter, Z., Von Tersch, R.L., Long, J., Berezhnoy, S . N., Phillips, R . S., Harutyunyan, E . H . & Wilson, K . S . (1993) Biochemistry 32, 4195--4206} . In order to evaluate the role of His343 in the reaction mechanism of tyrosine phenol-lyase and tryptophan indole-lyase, we have mutated it to Ala; the former mutant is referred to as {H343A}tyrosine phenol lyase . All substrates for alpha, beta-elimination (except S-ethyl-L-cysteine) exhibited lower kcat (10-30%) and kcat/Km (1-10%) values with {H343A}tyrosine phenol-lyase than with the wild-type enzyme . The mutant also shows slower rates of deuterium isotope exchange for L-phenylalanine and L-methionine than does the wild type . The pH-dependent behavior in the reaction of 3-fluoro-L-tyrosine with wild-type tyrosine phenol-lyase is identical to that of L-tyrosine described previously {Kiick, D . M . & Phillips, R . S . (1988) Biochemistry 27, 7333-7338} . The pH profile of kcat/Km for this reaction exhibits two pKa values with an average of 7.7 +/- 0.2, indicating that the catalytic mechanism requires two essential basic groups . The pH profile of kcat/Km for 3-fluoro-L-tyrosine with {H343A}tyrosine phenol-lyase also exhibits two pKa values with an average of 7.8 +/- 0.3 . However, kcat for 3-fluoro-L-tyrosine is pH-dependent for the mutant, exhibiting two pKa values with an average of about 7.8, whereas it is pH-independent for the wild type . Steady-state kinetic isotope effects on the reactions with wild-type and {H343A}tyrosine phenol-lyase were examined at various pH values . For the wild type, the values of the isotope effects on kcat and kcat/Km for 3-fluoro-L-{alpha-2H}-tyrosine are independent of pH and equal to 3.9 +/- 0.2 and 2.2 +/- 0.3, respectively, while the corresponding values for {H343A}tyrosine phenol-lyase are 5.4 +/- 0.2 and 3.8 +/- 0.3, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)

J Indian Med Assoc, 1995 Apr, 93(4), 132 - 5
Bacteria in surface infections of neonates; Ghosh S et al.; A bacteriological work on surface infections was done among live births (study group I) and neonates admitted in hospital (study group II) . Out of 134 cases of conjunctivitis in group I Gram-negative bacilli predominated (48.5%) with Escherichia coli accounting for 29 (14.9%) cases, Klebsiella species 15 (11.2%) cases, Citrobacter freundii 3 (2.2%) cases, Pseudomons aeruginosa 18 (13.4%) cases and Aeromonas hydrophila 3 (2.2%) amongst pure isolates (73.9%) . Gonococcus was noted in 2 (1.5%) cases . In group II, 41.7% were Staphylococcus aureus in pure growth (75%), compared to only 9.0% in group I . Skin infections were caused by both Staphylococcus aureus and Staphylococcus epidermidis . Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae and Pseudomonas aeruginosa were the principal insolates from umbilical sepsis . Pseudomonas aeruginosa was isolated as pure growth from local site of noma neonatorum . Anaerobic cultures were negative in all except in 2 cases of umbilical sepsis with tetanus neonatorum revealing Clostridium tetani which however proved to be non-toxigenic . Blood cultures were positive in 4 out of 14 cases bearing 50% correlation with bacteria from surface infections . A source study established partial correlation with the cases of pseudomonas conjunctivitis . Phage typing of Staphylococcus aureus and biochemical typing failed to detect any definite marker of clinical entities, except that the skin infections were caused by group III phages predominantly (65.0%).

Eur J Biochem, 1995 Apr 1, 229(1), 299 - 307
Structural elucidation of the biological repeating unit of O-specific polysaccharide from Citrobacter serotype O41; Ravenscroft N et al.; Sugar analysis of the O-specific polysaccharide produced by Citrobacter serotype O41 revealed the presence of a hexasaccharide repeating unit, which includes the unusual 3-amino-N-(D-3'-hydroxybutyryl)-3,6-dideoxy-D-galactosyl residue (Fuc3NAcyl) . The structure of the repeating unit was determined by extensive use of homo- and heteronuclear two-dimensional NMR spectroscopy, including the application of long-range 1H-13C correlation experiments and NOE studies to establish the sequence of sugar units . Indentification of the Glc beta 1-->2Fuc3NAcyl beta 1-->6GlcNAc alpha 1-->sequence at the non-reducing terminus establishes the biological repeating unit of this O-specific polysaccharide as: -->2Glc beta 1-->Fuc3NAcyl-beta 1-->6GlcNAc alpha 1-->(Glc alpha 1-->2) 4Gal beta 1-->3GalNAc beta 1--> . This structure is similar to that found for the O-specific polysaccharide isolated from Hafnia alvei strain 1211 {Katzenellenbogen, E., Romanowska, E., Dabrowski, U . & Dabrowski, J . (1991) Eur . J . Biochem . 200, 401-407}, which differs in having an acetyl substituent at O4 of the Fuc3NAcyl residue and a branch point of GalNAc alpha substituted by Glc beta at O3; these differences are responsible for the only weak serological cross-reactivity of the two strains.

J Bacteriol, 1995 Apr, 177(8), 2151 - 6
Purification of 1,3-propanediol dehydrogenase from Citrobacter freundii and cloning, sequencing, and overexpression of the corresponding gene in Escherichia coli; Daniel R et al.; 1,3-Propanediol dehydrogenase (EC 1.1.1.202) was purified to homogeneity from Citrobacter freundii grown anaerobically on glycerol in continuous culture . The enzyme is an octamer of a polypeptide of 43,400 Da . When tested as a dehydrogenase, the enzyme was most active with substrates containing two primary alcohol groups separated by one or two carbon atoms . In the physiological direction, 3-hydroxypropionaldehyde was the preferred substrate . The apparent Km values of the enzyme for 3-hydroxypropionaldehyde and NADH were 140 and 33 microM, respectively . The enzyme was inhibited by chelators of divalent cations but could be reactivated by the addition of Fe2+ . The dhaT gene, encoding the 1,3-propanediol dehydrogenase, was cloned, and its nucleotide sequence (1,164 bp) was determined . The deduced dhaT gene product (387 amino acids, 41,324 Da) showed a high level of similarity to a novel family (type III) of alcohol dehydrogenases . The dhaT gene was overexpressed in Escherichia coli 274-fold by using the T7 RNA polymerase/promoter system.

Antimicrob Agents Chemother, 1995 Mar, 39(3), 629 - 37
Characterization of an LysR family protein, SmeR from Serratia marcescens S6, its effect on expression of the carbapenem-hydrolyzing beta-lactamase Sme-1, and comparison of this regulator with other beta-lactamase regulators; Naas T et al.; Serratia marcescens S6 produces a chromosomally encoded carbapenem-hydrolyzing class A beta-lactamase, Sme-1 (T . Naas, L . Vandel, W . Sougakoff, D . M . Livermore, and P . Nordmann, Antimicrob . Agents Chemother . 38:1262-1270, 1994) . Upstream from smeA we identified a second open reading frame (EMBL accession number Z30237) . This encodes a 33.1-kDa protein, SmeR, which has a high degree of homology with NmcR, the LysR regulatory protein of the only other sequenced carbapenem-hydrolyzing class A beta-lactamase, NmcA from Enterobacter cloacae NOR-1 . It is weakly related to AmpR of the chromosomal cephalosporinase regulatory systems described in E . cloacae, Yersinia enterocolitica, Citrobacter freundii, and Pseudomonas aeruginosa and is very weakly related to other LysR-type regulators of class A beta-lactamases . SmeR is a weakly positive regulator for Sme-1 expression in the absence of or in the presence of beta-lactam inducers . The -35 and -10 regions of smeR are in the opposite orientations and are face-to-face relative to the smeA promoter . SmeR acts similarly to NmcR and not as the AmpR regulators described for class C beta-lactamase systems . SmeR is a weak inducer in the absence or presence of beta-lactams . As was found for the AmpC-AmpR and NmcA-NmcR systems, a putative SmeR-binding site was present upstream from the beta-lactamase gene promoter regions . beta-Galactosidase activity from a smeR-lacZ translational fusion was expressed constitutively and decreased in the presence of SmeR from a coresident plasmid, suggesting that SmeR is autogeneously controlled . Finally, beta-lactams did not affect the expression of SmeR, which is the second regulator of a class A carbapenem-hydrolyzing beta-lactamase to be identified.

Diagn Microbiol Infect Dis, 1995 Mar, 21(3), 141 - 51
National survey of the in vitro spectrum of piperacillin-tazobactam tested against more than 40,000 aerobic clinical isolates from 236 medical centers; Baron EJ et al.; 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.

Eur J Clin Microbiol Infect Dis, 1995 Mar, 14(3), 244 - 52
Multicentre comparative study on the antibacterial activity of FK-037, a new parenteral cephalosporin; Martinez-Beltran J et al.; The in vitro antibacterial activity of FK-037, a new parenteral cephalosporin structurally related to cefpirome and cefepime, was compared with that of cefotaxime, ceftazidime, aztreonam, cefpirome, cefepime, imipenem and meropenem against 1,837 clinical isolates obtained from three Spanish hospitals . FK-037 inhibited 90% of Enterobacteriaceae isolates at < or = 0.25 microgram/ml, with the exception of Enterobacter aerogenes (MIC90 1 microgram/ml), Enterobacter cloacae and Citrobacter freundii (MIC90 8 micrograms/ml) . In cefotaxime- and ceftazidime-resistant Klebsiella pneumoniae strains producing SHV-2 and SHV-6 beta-lactamases, the activity of FK-037, cefpirome and cefepime was similar (MIC range 0.25-32 micrograms/ml) . In Enterobacteriaceae strains hyper-producing chromosomally inducible beta-lactamases, FK-037 (MIC90 range, 0.25-8 micrograms/ml) was 8- to 16-fold more active than cefotaxime and ceftazidime but two- to eightfold less active than cefpirome and cefepime . FK-037 and cefpirome were twofold more active than ceftazidime and cefepime against Pseudomonas aeruginosa isolates, with MIC90 values of 16 micrograms/ml . The activity of FK-037, cefpirome and cefepime was two- to eightfold lower in ceftazidime-resistant derepressed Pseudomonas aeruginosa mutants . FK-037 (MIC range, 0.12-2 micrograms/ml) and the other beta-lactam agents tested were active against methicillin-susceptible staphylococci; however, only cefpirome and, particularly, FK-037 (MIC90 of 32 micrograms/ml) displayed some activity against methicillin-resistant strains . In penicillin-susceptible, -intermediate and -resistant Streptococcus pneumoniae isolates, the MIC90s of FK-037 were 0.03, 0.5 and 1 microgram/ml, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Microbiology, 1995 Feb, 141 ( Pt 2), 419 - 30
GcvA, a LysR-type transcriptional regulator protein, activates expression of the cloned Citrobacter freundii ampC beta-lactamase gene in Escherichia coli: cross-talk between DNA-binding proteins; Everett M et al.; Escherichia coli JRG582 is an ampD ampE deletion derivative of strain HfrH and accordingly it is derepressed for expression of the cloned inducible beta-lactamase gene of Citrobacter freundii, carried on plasmid pNU305 . Following chemical mutagenesis of JRG582(pNU305) a cefotaxime sensitive mutant was isolated, CS51(pNU305), which produced low levels of beta-lactamase due to a mutation in the host chromosome . Two recombinant plasmids containing genomic DNA from E . coli HfrH, namely pUB5608 and pUB5611, were isolated as a consequence of their ability to restore the beta-lactam resistant phenotype to CS51(pNU305) . This ability was due to direct transcriptional activation of the beta-lactamase gene, ampC, rather than complementation of the CS51 mutation . Transposon mutagenesis and subcloning showed that restoration of ampicillin resistance to CS51(pNU305) was the function of a single gene, which maps at 60.3 min on the E . coli chromosome . The gene encodes a 33 kDa protein with significant homology to members of the LysR family of bacterial activator proteins, in particular the AmpR protein from C . freundii . Homology is especially strong over the N-terminal region which includes the helix-turn-helix DNA-binding motif . This gene was shown to complement the gcvA1 mutation at 60.3 min on the E . coli chromosome, and the DNA sequence agrees exactly with the published sequence of gcvA which encodes the transcriptional activator of the inducible glycine cleavage enzyme system . It is suggested that GcvA can activate transcription of ampC by binding to the AmpR binding region upstream of ampC so as to mimic the activated state of AmpR and hence provides an example of cross-talk between DNA-binding proteins of different inducible enzyme systems.

J Chemother, 1995 Feb, 7(1), 21 - 5
Comparative evaluation of orally active antibiotics against community-acquired pathogens: a multi-center study in five Mediterranean countries; Cullmann W; In 5 Mediterranean countries 7902 pathogens, all isolated in 1992 and 1993 from community-acquired infections, were studied for susceptibility to the following orally active antibiotics: penicillin G, ampicillin, ampicillin + sulbactam, amoxycillin + clavulanic acid (both 2:1 ratio), cefalexin, cefaclor, cefuroxime, cefetamet, doxycycline and erythromycin . Ten centers in Italy, 4 centers in Greece, 3 centers in Spain, and 1 center in Lebanon and Saudi Arabia contributed to this study; all centers used performed standardized microtiter panels (Sceptor, BBL, Heidelberg, FRG) . The most frequently isolated pathogens were Escherichia coli (n = 1267), Proteus mirabilis (n = 843), Klebsiella pneumoniae (n = 771), enteric Salmonella spp . (n = 629), Enterobacter cloacae (n = 486), Citrobacter freundii (n = 383), Streptococcus agalactiae (n = 346), Haemophilus influenzae (n = 298), Streptococcus pyogenes (n = 294), Streptococcus pneumoniae (n = 246), Klebsiella oxytoca (n = 243), and Shigella spp . (n = 185) . Statistical analysis was performed for each of the above countries and for all pooled data available . The penicillin antibiotics were the most active compounds against the gram-positive cocci, exceeding the MIC90 values 2- to 8-fold over all cephalosporins . Regarding the gram-negatives (above all Klebsiella spp.) cefetamet was by far the most active compound (MIC90 = 1 mg/l) . Regarding the percentage of resistant isolates, there were no striking discrepancies between the centers and countries involved in this study . There was, however, complete cross-resistance in penicillin-resistant S . pneumoniae isolates (MIC90 = 2 mg/l) . By far the majority of the penicillin-resistant pneumococci showed additional resistance to doxycycline and erythromycin.(ABSTRACT TRUNCATED AT 250 WORDS)

Biochem Biophys Res Commun, 1995 Jan 26, 206(3), 942 - 7
Transformation of microorganisms with the plasmid vector with the replicon from pAC1 from Acetobacter pasteurianus; Grones J et al.; A number of gram-negative and gram-positive bacteria species was screened for the expression of the gram-negative plasmid pACK5 and pACT72 with replicon of pAC1 plasmid from Acetobacter pasteurianus . As was described previously, both plasmids were expressed in Escherichia coli, Acetobacter pasteurianus, Acetobacter aceti, Shigella spp . and Citrobacter spp . Expressions of plasmids were successful in twelve species tested, Comamonas terrigena, Salmonella typhimurium, Serratia marcescens, Bacillus cereus, Bacillus megatericum, Bacillus subtilis, Lactobacillus helveticus, Micrococcus luteus, Sarcina lutea, Staphylococcus aureus, Staphylococcus epidermidis, Streptoccocus feacalis, and the stability of plasmid DNA was tested after cultivation in non-selective conditions.

Microb Drug Resist, 1995 Winter, 1(4), 327 - 30
Five-year survey of cefotaxime resistance in Spain; Colom K et al.; During 1991-1995 a Spain collaborative study group surveyed the resistance to cefotaxime both in community as well as in hospital isolates of bacteria . The isolates tested during the study period of 5 years were 813, 875, 3631, 3184, and 3050 strains, respectively . Antimicrobial activity of cefotaxime was assayed by broth or agar microdilution, in accordance with criteria of the National Committee of Clinical Laboratory Standards (NCCLS) . Cefotaxime resistance included 2.5% of all isolates: 2.6% Enterobacteriaceae, 1.7% Streptococcus pneumoniae, 0.5% Haemophilus influenzae, 0.0% Haemophilus spp., and 0.0% Moraxella catarrhalis . The overall incidence of resistance to cefotaxime decreased fro member of Enterobacteriaceae from 3.6% in 1991 to 2.5% in 1995 . The incidence of resistance varied with the species and was highest in Enterobacter and in Citrobacter freundii.

Microb Drug Resist, 1995 Winter, 1(4), 285 - 91
Inducible expression of the chromosomal cdiA from Citrobacter diversus NF85, encoding an ambler class A beta-lactamase, is under similar genetic control to the chromosomal ampC, encoding an ambler class C enzyme, from Citrobacter freundii OS60; Jones ME et al.; This study aimed to characterize the molecular basis of beta-lactamase induction in Citrobacter diversus . The chromosomal beta-lactamase encoding region from C . diversus, strain NF85, was cloned and expressed in Escherichia coli . The cloned region was sequenced and open-reading frames encoding a class A beta-lactamase, designated cdiA, and a putative LysR-type transcriptional regulator protein, divergently transcribed from the beta-lactamase gene and designated cdiR, were identified . The nucleotide sequence of the NF85 cdiA was identical to that of the published C . diversus ULA27 ampC sequence . A putative helix-turn-helix DNA-binding motif was located at the N-terminus of CdiR, and homology with enterobacterial AmpR proteins was noted . CdiR was demonstrated to bind to the C . diversus cdiAR intergenic region but not to the C . freundii ampCR intergenic region . A putative CdiR binding motif was identified in the cdiAR intergenic region . The cloned cdiAR region was inducible in E . coli strains SNO3 and HfrH . The inducible phenotype was dependent on the E . coli ampD and ampG gene products . We conclude that the molecular basis of inducible cdiA expression in C . diversus is similar to that of C . freundii ampC.

Med Dosw Mikrobiol, 1995, 47(3-4), 155 - 68
{Evaluation of the usefulness of tests for production of Beta-D-glucuronidase and propylene glycol utilization for the differentiation of enterobacteriaceae rods}; Kaluzewski S et al.; The aim of the study was to inquire about the diagnostic usefulness of determining the activity of glucuronidase and utilisation of propylene glycol in Enterobacteriaceae rods . The study included 1511 strains: 411- E . coli, 278 - Klebsiella, 231 - Salmonella, 159 - Yersinia, 97 - Citrobacter, 75 - Shigella and 260 strains representing 6 other kinds of enteric rods . Determination was performed in a liquid medium containing in 1 ml 25 mcg MUG and 100 mcg ONPG . Propylene glycol (PG) utilisation was observed in peptone water with 2% of the substrate and with the Andrade indicator . In comparative tests Rambach commercial medium and MacConkey agar from the Fluorocult series were used . In the test with MUG a positive result was obtained from 81.8% E . coli, 65% - Shigella and 13% - Salmonella subgenus I . Only exceptionally was this test positive with Providencia, Enterobacter and Yersinia strains (1-5%) but negative with Citrobacter, Klebsiella, Serratia, Hafnia, Proteus and Morganella strains . Glucuronidase production is not sufficiently characteristic of E . coli strains isolated from humans to be the only basis for the preliminary differentiation of these rods from other Enterobacteriaceae . The test with ONPG was positive from 95-100% E . coli, Yersinia, Citrobacter, Klebsiella, Enterobacter and Hafnia strains; 61% - Shigella, 9% - Salmonella and 3% - Providencia, but negative with Serratia, Proteus and Morganella strains . Propylene glycol was decomposed by 74% Salmonella strains of subgenus I, 65-94% - Klebsiella, Yersinia and Citrobacter . Shigella, Enterobacter, Serratia, Proteus, Providencia and Morganella rods did not decompose propylene glycol . Evidence that among strains non-decomposing propylene glycol were all the studied S . typhi, S . paratyphi A, S . paratyphi C, S . choleraesuis, S . virchow and S . gallinarum strains as well as a significant percentage of strains representing 8 other Salmonella serotypes frequently detected allows to believe that the use of activity against propylene glycol even simultaneously with the test for galactosidase as basis for the differentiation of Salmonella rods of subgenus I from other Enterobacteriaceae can lead to errors already at the onset of diagnostic procedure.

Acta Univ Palacki Olomuc Fac Med, 1995, 139, 33 - 5
Resistance to antibiotics in gram-negative rods from clinical material of hospital and community origin; Kolar M et al.; The authors examined the resistance to 13 antibiotics and chemotherapeutic agents by the diluting micromethod and the routine disk method in the group of 5375 gram-negative strains of 7 genera which prevailed in clinical material of an hospital (FN Olomouc) and in material of community (OHS Olomouc) provenance during the years 1992 and 1993 . In the majority of cases, the resistance was more frequent in the strains isolated from the hospital material . The most remarkable differences were found in A cinetobacter sp . (9-76%), Enterobacter sp . (10-60%) and Citrobacter sp . (18-58%) . In other examined species, the differences varied in the ranges 3-40% in E.coli, 2-33% in Klebsiella sp., 0-28% in P . vulgaris, 2-28% in P.mirabilis, 0-16% in M.morganii, and 0-13% in P . aeruginosa.

Biosci Biotechnol Biochem, 1995 Jan, 59(1), 93 - 9
Purification and properties of crystalline 3-methylaspartase from two facultative anaerobes, Citrobacter sp . strain YG-0504 and Morganella morganii strain YG-0601; Kato Y et al.; 3-Methylaspartase (3-methylaspartate ammonia-lyase, EC 4.3.1.2) from two facultative anaerobes from soil, Citrobacter sp . strain YG-0504 and Morganella morganii strain YG-0601, were purified and crystallized from their crude extracts . Both of the Citrobacter and Morganella enzymes appeared to be a dimer of subunits of M(r) 40,000 and 44,000, respectively . The enzymes had similar enzymological properties: optimum pH for the deamination reaction of (2S,3S)-3-methylaspartic acid, substrate specificity, inhibitor, divalent and monovalent cation requirement, and N-terminal amino acid sequence homology . However, some differences were detected in pH and temperature stability, optimum pH for the amination reaction of mesaconic acid, optimum temperature, specific activity, and stability during electrophoresis . Both enzymes had similar enzymological properties to the known 3-methylaspartase from an obligate anaerobic bacterium, Clostridium tetanomorphum H1, except kinetic constants and substrate specificities.

J Clin Microbiol, 1995 Jan, 33(1), 242 - 5
Ability of commercial identification systems to identify newly recognized species of Citrobacter; O'Hara CM et al.; The genus Citrobacter was recently determined to contain 11 genetically distinct species . In addition, the International Committee on Systematic Bacteriology no longer recognizes C . diversus and has, instead, validated the name C . koseri in its place . The 11 species are C . freundii, C . koseri, C . amalonaticus, C . farmeri, C . youngae, C . braakii, C . werkmanii, C . sedlakii, and three unnamed groups, genomospecies 9, 10, and 11 . To determine the ease with which some identification systems could respond to these changes, we evaluated five systems for their potential ability to recognize current species in the genus Citrobacter . A simple dichotomous key using conventional biochemicals is presented that may be helpful to presumptively identify Citrobacter strains.

Gesundheitswesen, 1994 Dec, 56(12), 690 - 3
{Enterobacteria in mineral, spring and table water}; Schindler PR; Whereas coliform bacteria rank first, non-coliform enterobacteria can be isolated second in frequency from bottled mineral, spring and table waters in South Bavaria . Escherichia coli were found in 8, coliforms in 44 and non-coliform enterobacteria in 41 of 54 manufactories with hygienic problems . 11 different bacterial species were determined from coliforms with Enterobacter cloacae occurring most numerous in 19, Citrobacter freundii in 12 and Enterobacter amnigenus in 7 manufactories . From non-coliform enterobacteria 18 different species could be determined . Again Enterobacter cloacae was found most frequently in 17 manufactories followed by Enterobacter amnigenus in 11 and Serratia liquefaciens in 7 manufactories . The investigation of total Enterobacteriaceae is suggested as a suitable quality criterion for bottled mineral, spring and table waters.

Appl Environ Microbiol, 1994 Dec, 60(12), 4608 - 11
Isolation of three hexahydro-1,3,5-trinitro-1,3,5-triazine-degrading species of the family Enterobacteriaceae from nitramine explosive-contaminated soil; Kitts CL et al.; Three species of the family Enterobacteriaceae that biochemically reduced hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) were isolated from nitramine explosive-contaminated soil . Two isolates, identified as Morganella morganii and Providencia rettgeri, completely transformed both RDX and the nitroso-RDX reduction intermediates . The third isolate, identified as Citrobacter freundii, partially transformed RDX and generated high concentrations of nitroso-RDX intermediates . All three isolates produced 14CO2 from labeled RDX under O2-depleted culture conditions . While all three isolates transformed HMX, only M . morganii transformed HMX in the presence of RDX.

J Antimicrob Chemother, 1994 Dec, 34(6), 943 - 54
Production of a plasmid mediated AmpC-like beta-lactamase by a Klebsiella pneumoniae septicaemia isolate; Pornull KJ et al.; A septicaemia Klebsiella pneumoniae isolate from Greece (Ath1) was shown to be resistant to third generation cephalosporins, aztreonam and cefoxitin and this resistance was not decreased in the presence of clavulanic acid . The gene coding for the resistance phenotype, associated with a beta-lactamase showing cephalosporinase activity and a pI of 8.6, could be transferred into Escherichia coli K-12 by conjugation and transformation . DNA-hybridisation showed that this gene was located on two different plasmids, 7.8 and 80 kb respectively . The larger, conjugative plasmid also carried genes coding for another beta-lactamase (pI 6.6) and resistance to aminoglycosides, tetracycline, chloramphenicol and trimethoprim . Under highly stringent conditions the gene coding for the pI 8.6 beta-lactamase hybridised with chromosomal DNA from Citrobacter freundii OS60 but not from E . coli or Enterobacter cloacae . Furthermore, the restriction map of this beta-lactamase gene corresponded to that of ampC from C . freundii OS60 . The regulatory ampR gene could not be detected in the plasmid DNA from the Ath1 K . pneumoniae by DNA hybridisation . The relationship between a plasmid-mediated extended spectrum beta-lactamase in a K . pneumoniae septicaemia isolate and chromosomal AmpC beta-lactamase from C . freundii was demonstrated.

Infect Immun, 1994 Nov, 62(11), 4838 - 43
Comparison of the antibody responses to the 77 Klebsiella capsular types in ankylosing spondylitis and various rheumatic diseases; Sahly H et al.; The production of antibodies to Klebsiella capsular polysaccharides was measured in sera from either HLA-B27-positive (HLA-B27+) or HLA-B27-negative (HLA-B27-) patients with classical ankylosing spondylitis (n = 54) . These sera were compared with sera from patients with various rheumatic diseases (n = 82) and HLA-B27+ or HLA-B27- healthy individuals (n = 85) . All sera were analyzed by means of an enzyme-linked immunosorbent assay specific to each of the 77 Klebsiella serotypes . The sera from HLA-B27+ patients with ankylosing spondylitis showed a significantly higher antibody frequency to the capsular types K26, K36, and K50 than the sera from HLA-B27- ankylosing spondylitis patients, patients with psoriatic arthritis, systemic lupus erythematosus, rheumatoid arthritis, or reactive arthritis after Yersinia enterocolitica infection, or healthy controls (P < 0.02) . The antibodies were of the immunoglobulin G type . No significant antibody response to the other 74 Klebsiella serotypes, noncapsulated mutants of K26, K36, and K50, or preparations of Citrobacter, Serratia, Hafnia, or Morganella spp . or Streptococcus pneumoniae could be detected . The results might suggest a specific association between these capsular types and HLA-B27+ ankylosing spondylitis and might imply their predominance in this disease.

Antimicrob Agents Chemother, 1994 Nov, 38(11), 2583 - 9
Postantibiotic effect of meropenem on members of the family Enterobacteriaceae determined by five methods; MacKenzie FM et al.; The postantibiotic effect (PAE) of meropenem was determined for 11 strains, both clinical isolates and reference strains of members of the family Enterobacteriaceae . The study compares PAE results obtained by five methods used to monitor bacterial regrowth, including viable counting, alone and in combination with impedance; bioluminescence, alone and in combination with impedance; and a morphological technique . After exposure of the test organisms to meropenem (0.1 x to 100 x MIC) for 2 h, concentration-dependent differences in counts by bioluminescence and viable counts were observed, the latter always being lower . The differences varied with the test organism . For example, after exposure of Providentia stuartii NCTC 10318 to 0.1 x MIC, the counts were 5.5 x 10(5) and 2.0 x 10(5) whereas after exposure of Citrobacter freundii MR76 to 0.1 x MIC of meropenem the counts were 2.3 x 10(6) and 6.8 x 10(3) by bioluminescence and viable counting, respectively . The discrepancies were probably due to the relative inability of the viable counting procedure to detect fragile aberrant morphologies and resulted in differences in the calculated PAE values . With methods which do not detect fragile morphologies, the PAE may be underestimated . A general trend was observed for the order of magnitude of the PAEs by the following methods (in order of decreasing magnitude of PAE): (i) morphological technique, (ii) bioluminescence technique alone, (iii) bioluminescence in combination with impedance, (iv) viable counting in combination with impedance, and (v) viable counting alone . It is our opinion that of the methods examined in this study, bioluminescence in combination with impedance best reflects the true values for PAEs, and these results were examined more closely.

Nihon Kyobu Shikkan Gakkai Zasshi, 1994 Nov, 32(11), 1078 - 82
{A case of severe pneumonia in an elderly man, caused by Citrobacter freundii suspected to have a low susceptibility to imipenem/cilastatin sodium}; Matsui S; An 80-year-old blind man with lepromatous leprosy suffered from right femoral neck and humeral neck fractures on July 9, 1993 . Because of fever (38.6 degrees C), difficult expectoration and diffuse bilateral perihilar infiltrates with consolidation in the left lower lung field on his chest radiograph, severe pneumonia was diagnosed . With intravenous hyperalimentation, imipenem/cilastatin (IPM/CS), ceftazidime, minocycline, gentamicin (GM), and human immunoglobulin were administrated . On July 29, hip screw-plate fixation was done . Citrobacter freundii was isolated from the sputum and its susceptibility was IPM/CS+, GM3+ . Multi-drug therapy with GM and other antibiotics improved the patients' condition, but Citrobacter freundii were still detected and 43 days of medication were needed . According to a report by the Ministry of Health and Welfare in 1992, the resistance rate of IPM/CS against Citrobacter freundii is only 0.7%, and IPM/CS is more effective than beta-Lactams . This is a very rare case of severe pneumonia in an elderly patient caused by Citrobacter freundii that was suspected to have low susceptibility to IPM/CS.

Infection, 1994 Nov-Dec, 22(6), 423 - 4
Bifocal cervical spondylodiscitis due to Citrobacter diversus; Sotto A et al.; Spondylodiscitis due to Citrobacter diversus is rare . An unusual case of bifocal cervical spondylodiscitis following transurethral prostatectomy is reported . C . diversus was detected by urinary cultures and locally by intervertebral disc puncture . Satisfactory recovery occurred after treatment with imipenem and amikacin, and then imipenem alone . Citrobacter infections are still rare in adults but are increasing in immunocompromised patients and sometimes occur in healthy subjects following surgery.

Proc Natl Acad Sci U S A, 1994 Oct 11, 91(21), 10227 - 31
Intergeneric transfer and recombination of the 6-phosphogluconate dehydrogenase gene (gnd) in enteric bacteria; Nelson K et al.; The gnd gene, encoding 6-phosphogluconate dehydrogenase (EC 1.1.1.44), was sequenced in 87 strains of 15 species assigned to five nominal genera of the Enterobacteriaceae, including 36 isolates of Salmonella enterica and 32 strains of Escherichia coli . In S . enterica, the effective (realized) rate of recombination of horizontally transferred gnd sequences is only moderately higher than the rates for other chromosomal housekeeping genes . In contrast, recombination at gnd has occurred with such high frequency in Escherichia coli that the indicated evolutionary relationships among strains are not congruent with those estimated by sequence analysis of other genes and by multilocus enzyme electrophoresis . E . coli and S . enterica apparently have not exchanged gnd sequences, but those of several strains of E . coli have been imported from species of Citrobacter and Klebsiella . The relatively frequent exchange of gnd within and among taxonomic groups of the Enterobacteriaceae, compared with other housekeeping genes, apparently results from its close linkage with genes that are subject to diversifying selection, including those of the rfb region determining the structure of the O antigen polysaccharide.

Diagn Microbiol Infect Dis, 1994 Oct, 20(2), 93 - 8
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; Jones RN et al.; 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.

Pathol Biol (Paris), 1994 Oct, 42(8), 754 - 60
{In vitro antibacterial activity of cefpirome: a new cephalosporin; results of a multicenter study}; Soussy CJ et al.; The in vitro activity of cefpirome, a new injectable cephalosporin was studied against 1,082 clinical isolates in a multicentre study . Minimum inhibitory concentrations were determined using an agar dilution method . Cefpirome was active against Enterobacteriaceae . On naturally non-producing beta-lactamase species, cefpirome was active on most of the strains with MICs < or = 0.5 mg/l, including strains producing an acquired penicillinase: E . coli 0.03-0.06, P . mirabilis 0.06-0.12, Salmonella spp . 0.06-0.06, Shigella spp . 0.016-0.03 . MICs of K . pneumoniae (0.06-4) ranged from 0.016 to 32:MICs were high against expanded-spectrum betalactamase producers strains . Against species producing cephalosporinase, cefpirome was also active on most of the strains with MICs < or = 0.5: E . cloacae 0.06-2, Citrobacter spp . 0.03-1, S . marcescens 0.06-0.5, P . indol + 0.06-0.25, P . stuartii 0.12-0.5 . Cefpirome was less active on Pseudomonas aeruginosa, (8-32) and on A . baumanii (16-32) . MICs of cefpirome were low against Haemophilus spp . betalactamase producing or not (0.01-0.03) and M . catarrhalis (0.6-4) . Activity of cefpirome on methicillin-sensitive staphylococci, was higher than other third generation cephalosporins (0.25-2) comparable to that of cephalotin and cefamandol . Methicillin-resistant strains should be considered as resistant . Pneumococci (0.01-0.03), except for penicillin-R strains, and streptococci A, B, C, G (0.01-0.06) were very susceptible . Enterococci were of low sensitivity or resistant . Among anaerobes, C . perfringens appeared often susceptible, and Bacteroides spp . resistant.

Biochem Biophys Res Commun, 1994 Sep 30, 203(3), 1882 - 8
Cloning and sequencing of the gene for the lactose carrier of Citrobacter freundii; Lee JI et al.; The gene coding for the lactose carrier of Citrobacter freundii was cloned into the plasmid pBR322 . The gene was sequenced and the amino acid sequence was found to be 70% identical to the lactose carrier of E . coli . All of the charged residues in the membrane spanning region were conserved . The sugar specificity is somewhat different from that of E . coli . The C . freundii carrier has less activity for lactose and more activity for o-nitrophenyl-galactose (ONPG) than the carrier of E . coli.

J Med Microbiol, 1994 Sep, 41(3), 209 - 14
Heterogeneity at the beta-lactamase structural gene ampC amongst Citrobacter spp . assessed by polymerase chain reaction analysis: potential for typing at a molecular level; Jones ME et al.; Considerable biochemical diversity and polynucleotide sequence variation have been reported amongst strains of Citrobacter spp . However, sequence heterogeneity has not been investigated at gene loci of clinical relevance . In this study, sequence heterogeneity in the beta-lactamase structural gene, ampC, amongst 91 clinical isolates of Citrobacter spp . that showed resistance to various third-generation cephalosporins was investigated . Variation was examined by high-stringency polymerase chain reactions (PCR) with primers homologous to the known ampC sequences of C . freundii strains OS60 and I113, and C . diversus NF85 . If an isolate contained an ampC gene homologous to one of these three characterised ampC genes, a single PCR band of a predictable size was generated with the appropriate primer set; 50 (60%) of isolates gave a PCR product of the expected size with the OS60 primer set and nine (10%) gave a product with the I113 primer set . All these 59 isolates were identified as C . freundii by API-20E strips . Six isolates (7%) gave a product with the C . diversus NF85 primer set but only four of these were identified as C . diversus in API-20E tests; the other two isolates were identified as C . freundii . Of the 91 isolates, 28 (31%), were identified as either C . freundii or C . diversus, but gave no PCR product with any primer set tested . Five of these showed no homology to any of the reference strain ampC PCR products in hybridisation tests . Nevertheless, all showed beta-lactamase activity . Overall, this method allowed the identification of novel ampC gene loci, which may serve as a basis for the identification of Citrobacter spp . rapidly at a molecular level.

Chemotherapy, 1994 Sep-Oct, 40(5), 311 - 6
In vitro activity of cefpirome against beta-lactamase-inducible and stably derepressed Enterobacteriaceae; Binfiglio G et al.; Most members of the Enterobacteriaceae and Pseudomonas aeruginosa possess an inducible chromosomal class I beta-lactamase . Bacterial strains which produce high levels of beta-lactamase constitutively can be isolated from infections; these derepressed mutants are responsible for resistance to third-generation cephalosporins and ureidopenicillins . Minimum inhibitory concentrations of cefpirome, a fourth-generation cephalosporin, and other beta-lactam antibiotics were determined for a series of mutants of Citrobacter freundii, Enterobacter cloacae, Morganella morganii, Proteus vulgaris and Serratia marcescens with inducible stably derepressed or basal expression of chromosomal class I beta-lactamases . All the antibiotics tested were almost equally active against beta-lactamase-inducible organisms and their basal mutants . Imipenem and cefpirome showed better activity against derepressed mutants than third-generation cephalosporins and ureidopenicillins.

Gan To Kagaku Ryoho, 1994 Sep, 21(13), 2233 - 6
{Analysis of cases with liver abscess following transcatheter arterial chemoembolization (TAE) for malignant hepatic tumors}; Ishikawa H et al.; A total of 182 TAE procedures were carried out in 98 patients with malignant hepatic tumors during the last five years . Liver abscess following TAE occurred in 3 cases (3.1%) and in 3 procedures (1.7%) . All cases were discharged after successful percutaneous transhepatic abscess drainage . One case had hepatocellular carcinoma . Another case had undergone total gastrectomy and esophagojejunostomy with Roux-Y reconstruction for gastric leiomyosarcoma . The other had undergone right hemicolectomy and pancreatoduodenectomy for colon cancers and carcinoma of the ampulla of Vater . Communication between the abscess and the intrahepatic bile duct was recognized in 2 cases . In the abscess culture, E . coli and Citrobacter freundii were detected . These results suggest the major factor leading to abscess formation is biliary infection . Therefore, a previous bilio-enteric anastomosis should be regarded as a risk factor for liver abscess following TAE.

Diagn Microbiol Infect Dis, 1994 Sep, 20(1), 49 - 55
Cefquinome (HR 111V) . In vitro evaluation of a broad-spectrum cephalosporin indicated for infections in animals; Murphy SP et al.; 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.

Antimicrob Agents Chemother, 1994 Sep, 38(9), 2207 - 9
Nucleotide sequence of a plasmid-mediated cephalosporinase gene (blaLAT-1) found in Klebsiella pneumoniae; Tzouvelekis LS et al.; The nucleotide sequence of the gene encoding a novel cephalosporinase (LAT-1), carried by a non-self-transferable plasmid from Klebsiella pneumoniae, has been determined . It was found that the sequence shares a high degree of homology with the Citrobacter freundii OS60 ampC structural gene.

Antimicrob Agents Chemother, 1994 Sep, 38(9), 2150 - 7
Gene sequence and biochemical characterization of FOX-1 from Klebsiella pneumoniae, a new AmpC-type plasmid-mediated beta-lactamase with two molecular variants; Gonzalez Leiza M et al.; Klebsiella pneumoniae BA32, a clinical isolate from Buenos Aires, Argentina, was found to produce a plasmid-encoded beta-lactamase (FOX-1) which conferred resistance to broad-spectrum cephalosporins and cephamycins . Resistance could be transferred by conjugation or transformation into Escherichia coli K-12 via a 48.5-kb plasmid (pGLK1) that produced two FOX-1 molecular variants with isoelectric points of 6.8 and 7.2 and apparent molecular sizes of 37 and 35 kDa, respectively . The kinetic study revealed that the two variants had very similar substrate and inhibition profiles . These values resemble those of chromosomally mediated class C (group 1) cephalosporinases . The structural gene of FOX-1 (blaFOX-1) was cloned into a 2,270-bp PstI-PstI fragment and was expressed in E . coli TG1 . The deduced 382-amino-acid sequence of FOX-1 exhibited a high degree of similarity with chromosomally encoded AmpC beta-lactamases of Pseudomonas aeruginosa, Serratia marcescens, Enterobacter cloacae, E . coli, and Citrobacter freundii . These findings suggest that FOX-1 is a plasmid-mediated AmpC-type beta-lactamase that is encoded by a single gene and that has two molecular variants.

Nihon Kyobu Shikkan Gakkai Zasshi, 1994 Sep, 32(9), 851 - 5
{Clinical bacteriology and empiric therapy for hospital-acquired pneumonia in the elderly at a national leprosarium}; Matsui S et al.; Hospital-acquired pneumonia is one of the most important fatal respiratory diseases in the elderly . Prompt and precise empiric therapy is essential for recovery . Fourteen isolates from twelve elderly lepromatous leprosy patients (9 men, 3 women, mean age of 75.8 years) with hospital-acquired pneumonia were studied . Subsequently, empiric therapy with gentamicin and beta-lactams for nosocomial pneumonia in the elderly was examined . Fourteen types of bacteria isolated from expectorated sputum specimens consisted mainly of ten strains of gram-negative bacilli (71%) six of Klebsiella pneumoniae, one each of Citrobacter freundii, Enterobacter agglomerans, Serratia liquefaciens, and Aeromonas hydrophilia and four strains of gram-positive cocci (29%) two of Staphylococcus sp., one each of Streptococcus sp . and Streptococcus pneumoniae . Methicillin-resistant Staphylococcus aureus and Pseudomonas sp . were not detected . Resistance rates of the etiologic agents to the antibiotics showed that gentamicin was 7.7%, ceftazidime 0%, and cefmetazole 23.1% . Cephalosporins were superior to penicillins . As a result of empiric therapy, six elderly leprosy patients with nosocomial pneumonia were cured and one improved temporarily . This study shows the necessity of specific empiric therapy for hospital-acquired pneumonia in a hospital with many elderly patients . The combination of gentamicin and beta-lactams is of value as an initial antibiotic therapy for hospital-acquired pneumonia in the elderly.

FEMS Microbiol Lett, 1994 Aug 15, 121(2), 141 - 6
Phosphatase-mediated heavy metal accumulation by a Citrobacter sp . and related enterobacteria; Macaskie LE et al.; A Citrobacter sp . was reported previously to accumulate heavy metals as cell-bound heavy metal phosphates . Metal uptake is mediated by the activity of a periplasmic acid-type phosphatase that liberates inorganic phosphate to provide the precipitant ligand for heavy metals presented to the cells . Amino acid sequencing of peptide fragments of the purified enzyme revealed significant homology to the phoN product (acid phosphatase) of some other enterobacteria . These organisms, together with Klebsiella pneumoniae, previously reported to produce acid phosphatase, were tested for their ability to remove uranium and lanthanum from challenge solutions supplemented with phosphatase substrate . The coupling of phosphate liberation to metal bioaccumulation was limited to the metal accumulating Citrobacter sp.; therefore the participation of species-specific additional factors in metal bioaccumulation was suggested.

Jpn J Antibiot, 1994 Aug, 47(8), 1053 - 64
{Antibacterial activities of isepamicin against fresh clinical isolates of gram-negative bacilli}; Deguchi K et al.; To investigate antibacterial activities of isepamicin (ISP), MICs of ISP as well as other aminoglycosides (AGs) were determined against many strains of Gram-negative bacilli that were clinically isolated in 1993 . 1 . No ISP-resistant strains were observed among isolates of Escherichia coli, Citrobacter diversus, Klebsiella spp., Enterobacter spp . or Proteus mirabilis . 2 . ISP-resistant strains were observed among isolates of Citrobacter freundii, Serratia spp., Proteus vulgaris, Morganella morganii, Providencia spp . and Pseudomonas aeruginosa . The frequency of resistant strains in each species, however, was lower for ISP than other AGs . 3 . When MIC90s were compared, antibacterial activities of ISP determined in this study were quite similar to those determined during the drug's development period (1980's) in Japan, suggesting no increase in the number of ISP-resistant strains over the years . 4 . The number of clinically isolated Gram-negative bacilli resistant to multiple drugs are increasing in from year to year Japan . Our results in this study suggest that antibacterial activities of ISP may be potent enough against such Gram-negative bacilli resistant to multiple drugs.

Jpn J Antibiot, 1994 Aug, 47(8), 1041 - 52
{Clinical laboratory approach to evaluate administration dose of arbekacin . Reevaluation of in vitro MIC break points in the disk susceptibility test}; Matsuo K et al.; Antimicrobial activities of arbekacin (ABK) against various clinical isolates, 335 strains obtained in 1991, were determined and the reliability of the ABK disk susceptibility test in estimating approximate values of MICs was studied . In addition, clinical significance of two different systems for the interpretation of the disk tests was evaluated as to which system would be more useful for the evaluation of clinical efficacy of ABK . The 4 category system used in Japan, and the system proposed by the Japanese Society for Antimicrobial Chemotherapy were studied . In this study, MICs were determined using the Mueller-Hinton agar containing 50 mg/L of Ca and 25 mg/L of Mg at an inoculum level of 10(6) CFU/ml . MIC90 values of ABK against Staphylococcus aureus (MSSA and MRSA) and Staphylococcus epidermidis were both 3.13 micrograms/ml . Those against Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis and Proteus vulgaris, Enterobacter spp., and Citrobacter spp., were also < or = 3.13 micrograms/ml . MIC90 values against Pseudomonas aeruginosa and Serratia spp . were both 50 micrograms/ml . The disk susceptibility test was carried out according to the instruction in the Showa disk manual . The inhibition zones obtained with the disk method were compared with MICs . Results of ABK disk susceptibility test with 30, 10, 5 or 2 micrograms disks were well correlated with MICs, showing the reliability of the disk method in estimating approximate values of MICs (r = -0.627 approximately -0.724, P < 0.01) . In the 4 category classification system currently used, break points in MIC values of ABK proposed are (+ + +) MIC < 3 micrograms/ml, (++) MIC > 3-10 micrograms/ml, (+) MIC > 10-50 micrograms/ml and (-) MIC > 50 micrograms/ml . The results obtained with Showa 30 micrograms disks showed false positive in 13.4%, and false negative in 3.9% of the samples . With 10 micrograms disks, false positive and false negative were 8.1%, and 3.9%, respectively . Similarly, those with 5 micrograms and 2 micrograms disks were 6.9% and 7.2%, and 3.0% and 14.6%, respectively . In the break point classification system of Japanese Society for Antimicrobial Chemotherapy, the MIC break point for ABK proposed is 2 micrograms/ml . It appeared to be difficult to make out this break point on the inhibition zone diameters obtained with various disks used, since there were no significant difference in the inhibition zone diameters against strains with MIC values ranging 0.39-3.13 micrograms/ml . A pharmacokinetic examination with the recommended dose schedule for ABK (100 mg i.m . or i.v.) showed that plasma levels of ABK reached 3.7-11.3 micrograms/ml.(ABSTRACT TRUNCATED AT 400 WORDS)

FEMS Microbiol Rev, 1994 Aug, 14(4), 351 - 67
Enzymically accelerated biomineralization of heavy metals: application to the removal of americium and plutonium from aqueous flows; Macaskie LE et al.; A biological process for the removal of heavy metals from the aqueous flows is described . Metals are precipitated on the surface of immobilized cells of a Citrobacter sp . as cell-bound metal phosphates . This uses phosphate liberated by the activity of a cell-bound phosphatase . Some radionuclides (e.g . 241americium) form metal phosphates readily; efficient removal of 241Am on a continuous basis is demonstrated . At low phosphatase activities, the efficiency of uranium removal correlates with enzyme activity . High phosphatase activities are not realised as an increase in metal removal, suggesting that chemical events become rate-limiting . Studies have suggested that maximal metal uptake occurs only after nucleation and the formation of precipitation foci . A model is presented to illustrate how nucleation and crystallization processes could enhance the removal of plutonium and neptunium from dilute solutions.

Zh Mikrobiol Epidemiol Immunobiol, 1994 Aug-Sep, Suppl 1, 106 - 9
{Experimental and clinical studies of the effect of pectin on the causative agents of acute intestinal infections}; Potievskii EG et al.; The effect of alimentary pectin obtained from different raw materials (apple, tomato, cotton-wool) on the causative agents of infectious diarrheal diseases, belonging to the genera Shigella, Salmonella, Klebsiella, Enterobacter, Proteus and Citrobacter, was studied under experimental conditions and in clinical observations . The study revealed that pectin, irrespective of raw material used for its production, produced an inhibiting effect on these microorganisms . This effect was most pronounced, both under experimental and clinical conditions, when 5% pectin solution was used . A rapid suppress of diarrhea and other manifestations of the infectious syndrome was observed in patients . The patients treated with pectin stayed in the hospital, on the average, 2-3 days less than the control group of patients treated with antibiotics and other chemotherapeutic preparations.

Eur J Clin Microbiol Infect Dis, 1994 Aug, 13(8), 675 - 9
In vitro antimicrobial activity of cefpirome against ceftazidime-resistant isolates from two multicenter studies; Sader HS et al.; The in vitro activity of cefpirome against ceftazidime-resistant (MIC > 16 mg/l) isolates from two multicenter studies was analyzed . The first investigation carried out in the USA, was an in vitro comparison of cefpirome and five third-generation cephalosporins in which more than 6,000 isolates were evaluated, including 97 Enterobacteriaceae and 1,509 staphylococci resistant to ceftazidime . The second study was a multicenter international study (> 5,000 strains total) in which 160 ceftazidime-resistant gram-negative bacilli and 509 staphylococci from five countries (Australia, France, Germany, Italy and UK) were tested against cefpirome . The results from the US trial indicated that only 0.8% of enteric bacilli were resistant to cefpirome compared to 4.9% and 4.7% resistant to ceftazidime and cefoperazone, respectively . In the international trial, cefpirome was also active against ceftazidime-resistant, class I beta-lactamase producing enteric bacilli (75% susceptibility, MIC50 of 4 mg/l) especially against Citrobacter spp., Enterobacter spp . and Morganella morganii . Cefpirome was 8- to 64-fold more active than ceftazidime against seven different staphylococcal species . The antimicrobial activity of cefpirome against routine clinical isolates and those organisms resistant to third-generation cephalosporins was highly consistent within a nation (USA) and among various developed countries.

Biophys Chem, 1994 Jul, 51(1), 1 - 7
O-acetylation affects the binding properties of the carboxyl groups on the Vi bacterial polysaccharide; Bystricky S et al.; The capsular polysaccharide of Salmonella typhi and Citrobacter freundii (Vi) is a linear homopolymer of (1-->4)-linked 2-acetamido-2-deoxy-alpha-D-galactopyranosyluronic acid partially O-acetylated at the C-3 position . The physico-chemical properties of the carboxyl groups of the Vi polysaccharide, as a function of different degrees of O-acetylation, were studied by potentiometric titration, circular dichroism, and their reaction with the bulky nucleophile 2-nitro-phenylhydrazine (NPH) . Potentiometric titrations with K+ and Ca2+ hydroxides showed that the difference in the free energy of binding between the two cations (delta GKCa) was inversely proportional to the degree of O-acetylation . Similar cationic effects were found when measuring circular dichroism . Moreover there was also an inverse relation between the degree of O-acetylation and the extent of binding of NPH to the carboxyl groups . These data all indicate that O-acetyl groups hinder the association of carboxyls with cations and nucleophilic reagents . This provides a possible explanation for the importance of the O-acetyl and the relative unimportance of the carboxyl groups in contributing to the immunologic properties of the Vi.

Intensive Care Med, 1994 Jul, 20 Suppl 3, S10 - 3
Evolution of beta-lactamase inhibitors; Livermore DM; beta-Lactamases present the greatest single challenge to beta-lactam antibiotics, including piperacillin . beta-Lactamase-mediated resistance to supposedly beta-lactamase stable agents such as second- and third-generation cephalosporins is now emerging and inhibitor combinations provide an alternative strategy to overcome this problem . The success of this strategy depends on 1) how efficiently the inhibitor inhibits important beta-lactamases, 2) on how much beta-lactamase the bacteria produce, 3) on the drug that is to be protected, 4) on the permeability and intrinsic susceptibility of the organisms and 5) on the conditions, notably the pH . Tazobactam inhibits most of the clinically important beta-lactamases that give piperacillin resistance, except for the Class I types . Piperacillin itself is a relatively easy drug to protect, particularly against the TEM-type enzymes . The result is that tazobactam greatly extends the activity of piperacillin, notably against enterobacteria, but also against staphylococci and anaerobes . The survey confirmed the very broad spectrum of activity of piperacillin/tazobactam . Resistance occurred in about 17% of the Enterobacter, Citrobacter, Serratia group, where we believe it to have been caused by derepressed Class I enzymes since these strains were cross-resistant to third-generation cephalosporins . Otherwise, resistance was largely confined to such organisms as E . faecium and methicillin-resistant staphylococci, which have piperacillin insensitive penicillin-binding proteins . Finally, some question remains on the antistaphylococcal activity of piperacillin/tazobactam, where MIC tests gave a more favorable impression than disc tests . Nevertheless, early clinical results against staphylococcal infection appear good, with a response rate of nearly 90% {15}.

Lett Appl Microbiol, 1994 Jul, 19(1), 32 - 6
Evaluation of the Vitek Immunodiagnostic Assay System (VIDAS) for the detection of Salmonella in foods; Blackburn CW et al.; A Salmonella Assay using the Vitek Immunodiagnostic Assay System (VIDAS) was compared with a conventional cultural method (CCM) for the detection of salmonellas in 141 samples of artificially and naturally contaminated foods . There was an overall agreement of 92.9% between the methods . The productivity of the VIDAS Salmonella Assay (VSA) was not improved using an alternative enrichment protocol for the detection of Salmonella in 12 raw meat samples . The sensitivity and specificity of the VSA was assessed using pure cultures of salmonellas and non-salmonellas . The detection limit was 1.8 x 10(6) salmonellas ml-1 in M-broth and some Citrobacter freundii strains gave false-positive results . Using an immunomagnetic separation (IMS) technique and an abbreviated cultural enrichment, the VSA results could be obtained a day earlier than the standard VSA method.

Antimicrob Agents Chemother, 1994 Jun, 38(6), 1374 - 7
Interaction of oxyimino beta-lactams with a class C beta-lactamase and a mutant with a spectrum extended to beta-lactams; Nukaga M et al.; The class C beta-lactamase of Citrobacter freundii GN346 is a typical cephalosporinase comprising 361 amino acids, and substitution of the glutamic acid at position 219 in the enzyme by lysine was previously shown to broaden its substrate spectrum to oxyimino beta-lactams (K . Tsukamoto, R . Ohno, and T . Sawai, J . Bacteriol . 172:4348-4351, 1990) . To clarify this spectrum extension from the kinetic point of view, the interactions of cefuroxime, ceftazidime, and aztreonam with the wild-type and mutant enzymes were analyzed . In addition to aztreonam, known as a progressive inhibitor of class C beta-lactamases, cefuroxime and ceftazidime were found to act as progressive inhibitors of the wild-type enzyme . On the other hand, only aztreonam showed weak progressive inhibition of the mutant enzyme . On the basis of kinetic parameters, a minimum scheme for interaction of the oxyimino beta-lactams with the wild-type enzyme was proposed, and the rate-limiting step of the hydrolysis of unfavorable substrates was indicated to be conversion of the stable acyl-enzyme intermediate to the unstable intermediate . In aztreonam hydrolysis by the mutant enzyme, the reaction rate at the rate-limiting step was 2,000 times that of the wild-type enzyme . These results indicate that the mutation at position 219 disturbs the stabilization of the stable intermediate.

Appl Environ Microbiol, 1994 Jun, 60(6), 1789 - 97
Comparison of the taxonomy, serology, drug resistance transfer, and virulence of Citrobacter freundii strains from mammals and poikilothermic hosts; Toranzo AE et al.; In this study, the phenotypic, antigenic, and virulence characteristics of 32 Citrobacter freundii strains of fish, human, and veterinary origin were comparatively analyzed . In addition, the spread of drug resistance factors by conjugation was investigated . Regardless of the source of isolation, the strains exhibited variable reactions mainly for arginine dihydrolase, ornithine decarboxylase, and fermentation of sucrose, melibiose, amygdalin, and salicin . Total fatty acid methyl ester analysis by gas chromatography proved to be useful for an intratypic differentiation within the C . freundii strains studied . In fact, although all of the isolates exhibited similar fatty acid methyl ester profiles, significant differences in the major fatty acids 16:1 and 16:0 and in the 17:0 delta region were observed between the isolates from salmonids and the remaining strains . Serological studies using agglutination tests, analysis of lipopolysaccharides (LPS), and the corresponding immunoblots with 13 antisera indicated a great antigenic diversity among the strains . Common LPS patterns were shared only by some isolates showing high cross-agglutination titers . In contrast, although all strains exhibited very similar surface protein patterns, only two common outer membrane proteins of 54 and 58 kDa were immunologically related . Infectivity trials performed in mice and rainbow trout indicated that all of the C . freundii strains were not pathogenic for mice (50% lethal dose of > 5 x 10(7)) . Although the isolates displayed a low degree of virulence for trout, inoculated strains were always recovered from the survivors in pure culture.(ABSTRACT TRUNCATED AT 250 WORDS)

J Appl Bacteriol, 1994 Jun, 76(6), 553 - 8
Identification of infant food as a vehicle in a nosocomial outbreak of Citrobacter freundii: epidemiological subtyping by allozyme, whole-cell protein and antibiotic resistance; Thurm V et al.; A total of 38 Citrobacter freundii strains was isolated from patients and their environment at a neonatal intensive care unit of a large hospital where colonization and clinical diseases due to the agent had been observed . Epidemiological investigations were carried out by subtyping, for which a combination of allozyme, whole-cell protein and resistance pattern analysis was used . Infant formula was identified as a vehicle of nosocomial spread . This shows that the role of foods in the transmission of hospital infections should not be underestimated . The combination of methods applied, in particular a limited enzyme set, is recommended also for epidemiological investigations of food-borne infections and establishment of their causes.

J Trop Med Hyg, 1994 Jun, 97(3), 161 - 6
Transferable drug resistance (R-factor) among the enterobacteriaceae in urinary tract infections: a study at an urban hospital in Bangladesh; Chowdhury MA et al.; Prevalence and patterns of drug resistance were studied among Enterobacteriaceae, isolated from the cases of urinary tract infection (UTI) attending at an outpatient department of an urban hospital in Dhaka . Out of 90 Enterobacteriaceae isolated, 95.5% were resistant to the different antimicrobials tested . The most common resistance pattern observed was against ampicillin (A), trimethoprim (Tm), sulphamethoxazole (S), tetracycline (T) and chloramphenicol (C) in all four genera of Enterobacteriaceae . Transferable drug resistance (R(+)-factor) was detected in 68.5% Escherichia coli (E . coli), 60% Klebsiella, 66.6% Proteus and 50% Citrobacter strains . By using a resistance transfer factor (RTF) mobilizing strain, resistance factors were transferred from 3 (11.3%) out of 26 non-autotransferable strains . The study revealed that transferable drug resistance is common in organisms isolated from UTI in Bangladesh.

Aust N Z J Med, 1994 Jun, 24(3), 307 - 11
Septic arthritis: a second decade of experience; Youssef PP et al.; AIMS: To review the diagnostic features, treatment and outcomes of all cases of septic arthritis presenting to a major Australian rheumatology unit, between 1982 and 1991 . These were compared with the previous decade's experience . METHODS: The medical records of all cases of septic arthritis presenting to the Combined Centre for Rheumatic Diseases, The Rachel Forster Hospital between 1982 and 1991 were reviewed and compared with the experience of the previous decade (1971-1981) . RESULTS: Twenty-seven episodes of septic arthritis were diagnosed in 27 patients . There were 18 females and nine males . The average age was 62 (21-83) with three patients less than 30 . Their rheumatological diagnoses were: rheumatoid arthritis (RA) in 15, osteoarthritis in five, gout in two, and one each of mixed connective disease, sarcoid, tenosynovitis of the forearm, seronegative spondyloarthropathy, and non specific polyarthritis . Eleven patients were on oral corticosteroids . Four patients had intra-articular injections within two months of the onset of the septic episode . Sixteen out of 19 aspirates on the wards were positive . The organisms identified were Staphylococcus aureus in ten (one multiply resistant S . aureus (MRSA), Streptococcus four, Mycobacterium three (one atypical), two Pseudomonas and one each of Citrobacter, Enterobacter, Gram negative bacillus . Five patients did not have a causative organism identified . The site of involvement and the causative organisms were similar in both decades . All patients received intravenous antibiotics for at least two weeks and oral antibiotics for at least another four weeks . Twenty-two per cent had regular aspirations on the wards and 26% had surgical drainage performed . Only 59% of all joints returned to good function . Fifty per cent of infected arthroplasties required arthrodesis and only a third of these returned to acceptable function . CONCLUSION: Septic arthritis in subjects with previous rheumatic disease continues to have a poor prognosis, especially in cases of infected arthroplasties . There has been no change in the types of causative organisms or sites of involvement over the last two decades.

Biol Pharm Bull, 1994 Jun, 17(6), 794 - 7
Characterization of the lactose transport system in Citrobacter freundii; Okazaki N et al.; The lactose transport system of Citrobacter freundii was characterized . Both the lactose transport system and beta-galactosidase were induced with either lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG), the latter being the better inducer . The Km values for methyl-beta-D-thiogalactopyranoside (TMG) transport and lactose transport were 0.61 mM and 1.1 mM, respectively, and the Vmax values were 53 nmol/min/mg cell protein and 12 nmol/min/mg cell protein, respectively . Thus, TMG is a better substrate than lactose . Thiogalactopyranoside (TDG) was a very potent competitive inhibitor . Neither Na+ nor Li+ had a significant effect on the TMG transport or the lactose transport . Proton/substrate cotransport (symport) via this system was observed.

Mol Microbiol, 1994 Jun, 12(6), 893 - 901
Unique fimbriae-like structures encoded by sefD of the SEF14 fimbrial gene cluster of Salmonella enteritidis; Clouthier SC et al.; The SEF14 gene cluster of Salmonella enteritidis was recently shown to contain three genes, sefABC, encoding a unique fimbrin, and proteins homologous to fimbrial chaperones and outer membrane proteins (ushers), respectively . A fourth open reading frame, designated sefD, was found immediately downstream of sefABC and overlapping sefC . The translated protein sequence of sefD was unique, but the composition was similar to that of other bacterial fimbriae . SefD was produced in abundance by wild-type S . enteritidis as shown by Western blot analysis using antibodies raised to affinity-purified, recombinant SefD . Furthermore, unusually long, thin, fimbriae-like structures were evident on S . enteritidis and Escherichia coli by immunoelectron microscopy, but in other bacterial species SefD was expressed as amorphous material . Therefore, in S . enteritidis and E . coli, SefD is the predominant structural subunit of SEF18 . The SEF18 fimbriae-like structures were shown to be serologically distinct from the three known S . enteritidis fimbriae SEF14, SEF17 and SEF21 . Furthermore, SEF18 was still produced in sefA insertion mutants, indicating that SEF14 and SEF18 were structurally distinct . Thus, the SEF14 gene cluster is the first example in the Enterobacteriaceae of a gene cluster that encodes two fimbrin-like proteins, which are assembled into two distinct cell-surface structures, SEF14 and SEF18 . DNA hybridization and Western blot analyses showed that SefD was widely distributed among the Enterobacteriaceae and was present in E . coli, Shigella, Enterobacter, Citrobacter, Erwinia, Hafnia, Klebsiella, Providencia, and Proteus but not in the non-Enterobacteriaceae Gram-negative bacteria Pseudomonas and Aeromonas, or in Gram-positive bacteria Bacillus or Staphylococcus.(ABSTRACT TRUNCATED AT 250 WORDS)

FEMS Microbiol Lett, 1994 May 15, 118(3), 237 - 42
Salmonella typhimurium enterotoxin epitopes shared among bacteria; Chopra AK et al.; The deduced amino acid sequence of the cloned Salmonella enterotoxin gene (stn) was used to prepare anti-peptide antibodies . These antibodies were then employed to screen isolates of this enteric pathogen for the synthesis of protein enterotoxin (Stn) . Cell lysates of all Salmonella isolates tested displayed a prominent immunoblot band of approximately 29 kDa, which was consistent with the size of the cloned stn gene product . Among other Gram-negative bacteria examined, isolates of Klebsiella, Enterobacter, and Citrobacter exhibited a similar-sized protein that reacted strongly with the Stn antibodies . Since the stn gene was located opposite the hydrogenase regulatory genes (hydHG) required for hydrogen metabolism in bacteria, our data suggested that only in Salmonella and some other members of the family Enterobacteriaceae had the DNA sequence evolved, presumably through point mutations, into an expressed gene product of similar size.

Infect Immun, 1994 May, 62(5), 1835 - 42
Characterization of the C-terminal domains of intimin-like proteins of enteropathogenic and enterohemorrhagic Escherichia coli, Citrobacter freundii, and Hafnia alvei; Frankel G et al.; Surface proteins called intimins (Int), which are homologous to the invasin protein (Inv) of Yersinia spp., play a role in inducing brush border damage, termed attachment and effacement, which follows infection by enteropathogenic and enterohemorrhagic Escherichia coli, Citrobacter freundii biotype 4280, and Hafnia alvei . Maltose-binding protein (MBP) fusions containing the C-terminal 280 amino acids of Int-like proteins of strains of enteropathogenic E . coli, enterohemorrhagic E . coli, H . alvei, and C . freundii biotype 4280 and of Yersinia pseudotuberculosis Inv were constructed and purified . The 3' end of the gene for the H . alvei Int-like protein was sequenced and showed homology to corresponding regions of other Int-encoding genes . Binding of MBP-Int-like and MBP-Inv fusion proteins to HEp-2 cells was demonstrated by immunofluorescence microscopy and by fluorescence-activated cell sorting . MBP-Inv induced attachment and spreading of HEp-2 cells to plastic-coated wells, but MBP-Int-like fusion proteins did not . Preincubation of HEp-2 cells with MBP-Inv, but not with MBP-Int-like fusion proteins, inhibited MBP-Inv-induced cell attachment . Fixed staphylococci and fluorescent polymer microspheres coated with both MBP-Int-like and MBP-Inv fusion proteins showed enhanced adhesion to HEp-2 cells . These fusion proteins will facilitate studies of the role of intimin in the pathogenesis of diarrhea associated with members of the family Enterobacteriaeceae that induce attachment and effacement.

Int J Food Microbiol, 1994 May, 22(2-3), 217 - 22
Modes of inhibition of foodborne non-Salmonella bacteria by selenite cystine selective broth; Chen H et al.; Various cell densities of six common foodborne non-Salmonella bacteria were exposed to selenite cystine (SC) Salmonella selective medium . The insensitivity of Pseudomonas aeruginosa and Proteus vulgaris to SC was confirmed . Selenite cystine selective medium was effective against the sensitive bacteria up to certain cell densities, beyond which the bacteria survived . As judged from the minimum cell number required for survival in SC, Staphylococcus aureus was the most sensitive to SC, followed by Bacillus cereus, Escherichia coli and Citrobacter freundii . When sensitive bacteria were grown in SC, they enriched resistant variants which exhibited no or reduced sensitivity to SC . The change in density of sensitive cells after exposure to SC suggested that bacterial sensitivity to SC depended on the efficiency of killing and growth inhibition by SC as well as the fraction of resistant variants in the bacterial population . Since Salmonella samples generally contain unknown numbers and types of sensitive bacteria, it is difficult to predict the effectiveness of their selective inhibition by SC.

Antimicrob Agents Chemother, 1994 May, 38(5), 1182 - 5
Cloning and sequence analysis of blaBIL-1, a plasmid-mediated class C beta-lactamase gene in Escherichia coli BS; Fosberry AP et al.; The extended-spectrum, plasmid-borne beta-lactamase gene blaBIL-1, which was discovered in Escherichia coli, has been cloned . Unusually for a plasmid-borne beta-lactamase, blaBIL-1 encodes a novel class C enzyme and appears to have originated from the chromosomal ampC gene of Citrobacter freundii.

Jpn J Antibiot, 1994 May, 47(5), 502 - 20
{Antibacterial activity of aztreonam against clinical isolates}; Igari J; To evaluate the antibacterial activity of a monobactam antibiotic, aztreonam (AZT), MICs of AZT and other antibiotics against clinical isolates collected at 36 participating institutions after January 1992 were determined using the agar plate dilution method (size of inoculum, 10(6) cfu/ml) according to the Japan Society of Chemotherapy standard . The antibiotics that were tested along with AZT included piperacillin (PIPC), cefoperazone (CPZ), and amikacin (AMK) . AZT was found to be more active against Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Proteus vulgaris, Morganella morganii, and Providencia rettgeri than CPZ, PIPC, and AMK . AZT was also more active against Serratia marcescens than the other antibiotics, but about 6% of the strains tested were resistant to AZT with MIC of 12.5 micrograms/ml . Against Enterobacter cloacae and Citrobacter freundii, however, was more active AMK than AZT . AZT showed a normal activity distribution with a single peak at the MIC of 3.13-6.25 micrograms/ml against Pseudomonas aeruginosa, and 35% of inoculum was resistant with high MIC values (MIC > or = 25 micrograms/ml) . Activities of AZT against Haemophilus influenzae and Neisseria gonorrhoeae were comparable to those of CPZ.

Enferm Infecc Microbiol Clin, 1994 May, 12(5), 246 - 50
{Development of a selective differential medium for the isolation of Corynebacterium urealyticum (group D-2)}; Munoz JL et al.; BACKGROUND: We have developed a new selective culture medium for the isolation of C . urealyticum, with the aim of improving and making easier the isolation and identification of this microorganism . MATERIAL AND METHODS: The medium is based on components similar to other media, usually used for diagnosis of urinary tract infections, also containing glucose, urea, phenol red, polysorbate 80 (Tween 80), polymixin B, amphotericin B, nalidixic acid and lyncomycin . The medium was tested in 65 clinical isolates and three type strains, and in 533 clinical samples of urine . RESULTS: All 65 clinical strains and 3 reference strains of C . urealyticum tested grew faster on this medium than on blood agar . E . coli, Klebsiella spp., Enterobacter spp., Citrobacter spp., M . morganii, Proteus spp., S . aureus, Enterococcus spp . and Corynebacterium spp . did not grow in the selective agar . S . epidermidis and Streptococcus spp . grew in less than 10% of cases, P . aeruginosa in 29.2% and Serratia spp . in 41.7% . The colonies were always easily differentiated from C . urealyticum . In the studies performed on 553 clinical samples from patients with urinary tract infections, six infections by C . urealyticum were detected . The growth of all strains being faster on this medium than on blood-agar . CONCLUSIONS: This new selective medium may be a valuable tool for diagnosis of UTIs caused by C . urealyticum in patients with retard risk factors.

Appl Environ Microbiol, 1994 May, 60(5), 1495 - 9
Vitamin B12 production by Citrobacter freundii or Klebsiella pneumoniae during tempeh fermentation and proof of enterotoxin absence by PCR; Keuth S et al.; The influence of some fermentation parameters on vitamin B12 formation by strains of Citrobacter freundii and Klebsiella pneumoniae isolated from Indonesian tempeh samples during tempeh fermentation was investigated . A decrease in fermentation temperature from 32 to 24 degrees C led to a decrease in vitamin B12 formation . Inoculation of soybeans with different numbers of cells of C . freundii at the beginning of solid-substrate fermentation showed that only the velocity of vitamin formation and not the final amount of vitamin formed depended on the number of cells . The addition of cobalt and 5,6-dimethylbenzimidazole increased the vitamin B12 content of tempeh . Nevertheless, levels of incorporation of the two precursors into the vitamin B12 molecule were very low . Neither C . freundii nor K . pneumoniae possessed the genes encoding the enterotoxins Shiga-like toxin SLT IIA, heat-labile enterotoxin LT Ih, and heat-stable enterotoxin ST Ih, as indicated by PCR . This result supports the suggested use of these two strains to form vitamin B12 during tempeh fermentation in Indonesia.

Gene, 1994 Apr 8, 141(1), 85 - 9
Cloning and analysis of translational control for genes encoding the Cfr9I restriction-modification system; Lubys A et al.; The complete type-II Cfr9I restriction-modification (R-M) system of Citrobacter freundii strain RFL9, recognizing the DNA sequence CCCGGG, has been cloned and expressed, and functionally active enzymes have been produced in Escherichia coli . Both the methyltransferase (MTase; M.Cfr9I) and restriction endonuclease (ENase; R.Cfr9I) were found to be encoded on a 2.3-kb cloned fragment in the same transcriptional orientation, but differing in translational phases . The last codon (underlined) (ATGA) of the MTase-encoding gene (Cfr9IM) overlaps with the start codon for the ENase-encoding gene (overlined) (cfr9IR) . A nucleotide sequence complementary to a predicted Shine-Dalgarno sequence preceding cfr9IR is within this gene . Predicted free energy (delta G) for formation of the mRNA secondary structure involving these complementary sequences was found to be -16.1 kcal/mol . Amino-acid sequence homology of 80% was found between R.Cfr9I and R.XcyI.

Int J Food Microbiol, 1994 Apr, 22(1), 23 - 31
Formation of B-vitamins by bacteria during the soaking process of soybeans for tempe fermentation; Denter J et al.; The formation of B-vitamins (nicotinic acid and nicotinamide, thiamine, vitamin B6 and vitamin B12) during the soaking of soybeans by bacteria, isolated from tempe, was investigated . Among the isolates examined no vitamin B6 producer was found . After inoculation of the soaking soybeans with Citrobacter freundii, Klebsiella pneumoniae, Pseudomonas fluorescens and Streptococcus spp . the concentrations of vitamin B12 increased significantly . For the first time it was found that soakings inoculated with C . freundii showed an increased vitamin B12 content . Nicotinic acid and nicotinamide were produced by Lactobacillus spp . and C . freundii . C . freundii synthesized thiamine as well.

J Clin Microbiol, 1994 Apr, 32(4), 931 - 4
Evaluation of RapID onE system for identification of 379 strains in the family Enterobacteriaceae and oxidase-negative, gram-negative nonfermenters; Kitch TT et al.; The ability of the RapID onE system (Innovative Diagnostic Systems, Inc., Norcross, Ga.) to identify 364 strains in the family Enterobacteriaceae and 15 oxidase-negative, gram-negative, nonfermentative rods was evaluated . Kits were inoculated with no . 2 McFarland standard suspensions, and reactions were interpreted after 4 h of incubation at 35 degrees C . Overall, the method correctly identified (to the species level or to the genus level for salmonellas and non-Shigella sonnei Shigella species) 363 strains (95.8%) without additional tests . For four strains (1.0%), additional tests were required to delineate the correct identification from a range of two or more possibilities; these included one Serratia liquefaciens (Serratia marcescens or Serratia liquefaciens), one Serratia rubidaea (Serratia rubidaea or Serratia odorifera), one Salmonella typhi (Leminorella richardii or Salmonella sp.) and one Yersinia enterocolitica (Yersinia frederiksenii, Yersinia intermedia, or Yersinia enterocolitica) . Twelve strains (3.2%) were misidentified or yielded codes with no identification; these comprised one Citrobacter amalonaticus (no identification), three Enterobacter hormaechei (not in the RapID onE database; two Enterobacter amnigenus, one Enterobacter sp.), one Serratia liquefaciens (Enterobacter cloacae), one Serratia rubidaea (no identification), four Serratia fonticola (not in RapID onE database; two Enterobacter aerogenes, one Serratia marcescens, one not identified), one Proteus mirabilis (Proteus penneri), and one Proteus vulgaris (Providencia rustigianii) . If the seven strains not included in the database had been excluded, correct identification rates would have risen to 97.6% without additional tests and 98.7% with additional tests, with misidentification rates dropping to 1.3% . The RapID onE system is easy to set up and the results are easy to read, and the system provides an accurate, nonautomated commercially available method for the same-day identification of members of the family Enterobacteriaceae and oxidase-negative, gram-negative nonfermenters.

J Antimicrob Chemother, 1994 Mar, 33(3), 453 - 64
Comparative in-vitro activity of biapenem against enterobacteria with beta-lactamase-mediated antibiotic resistance; Chen HY et al.; The effects of enterobacterial beta-lactamases were studied for biapenem (L627), a new carbapenem . Susceptibility tests were performed for isogenic mutant series of Citrobacter freundii, Enterobacter cloacae, Morganella morganii, Serratia marcescens and Proteus vulgaris which varied only in chromosomal beta-lactamase expression . beta-Lactamase-derepressed organisms in these series were as susceptible as beta-lactamase-inducible strains to biapenem; beta-lactamase-basal mutants were up to eight-fold more susceptible . Similar patterns of relative activity against the different expression types were noted for imipenem and biapenem . These data were related to direct induction and hydrolysis assays: biapenem, like imipenem, was a strong inducer of several Class I enzymes and of the P . vulgaris cefuroximase and, like the other carbapenems, was only very slowly hydrolysed by these enzymes . Moreover, like meropenem, biapenem reversibly deactivated these beta-lactamases . Piperacillin and the cephalosporins, tested as comparators, were more labile than carbapenems to the Class I enzymes, were weak inducers below their MICs and lacked deactivator function . In consequence their MICs were higher for derepressed organisms than for those with inducible or basal beta-lactamase expression . Unlike the carbapenems, they selected derepressed mutants from inducible populations . Biapenem, like imipenem and meropenem, retained full activity against most transconjugants of Escherichia coli K-12 that produced plasmid-mediated beta-lactamases, including extended-spectrum TEM mutants . Only production of OXA-10 (previously PSE-2) enzyme gave a slight reduction in susceptibility to the new carbapenem . Biapenem resistance (MIC 16 mg/L) did, however, occur in S . marcescens S6, which produced a chromosomal carbapenemase . This enzyme hydrolysed biapenem . Overall, our findings indicate that biapenem shares the favourable properties of imipenem and meropenem in its interactions with the most important beta-lactamases of enterobacteria.

Drugs, 1994 Mar, 47(3), 506 - 35
Piperacillin/tazobactam . A review of its antibacterial activity, pharmacokinetic properties and therapeutic potential; Bryson HM et al.; Combining tazobactam, a beta-lactamase inhibitor, with the ureidopenicillin, piperacillin, successfully restores the activity of piperacillin against beta-lactamase-producing bacteria . Tazobactam has inhibitory activity, and therefore protects piperacillin against Richmond and Sykes types II, III, IV and V beta-lactamases, staphylococcal penicillinase and extended-spectrum beta-lactamases . However, tazobactam has only species-specific activity against class I chromosomally-mediated enzymes . Resistant organisms include some Citrobacter spp., Enterobacter spp., Serratia spp., Xanthomonas maltophilia and Enterococcus faecium . Consistent with its in vitro activity, preliminary clinical data indicate that the fixed combination of piperacillin/tazobactam (dose ratio 8:1) is effective in the treatment of moderate to severe polymicrobial infections, including intra-abdominal, skin and soft-tissue and lower respiratory tract infections . In limited comparative trials, piperacillin/tazobactam demonstrated equivalent or better efficacy than standard comparator regimens in these infections . Piperacillin/tazobactam in combination with an aminoglycoside was effective in the empirical treatment of fever in patients with neutropenia and compared favourably with ceftazidime in combination with an aminoglycoside, although second-line therapy with a glycopeptide antibiotic may be indicated in unresponsive episodes . Data from phase III trials indicate that piperacillin/tazobactam has a tolerability profile typical of a penicillin agent . Piperacillin/tazobactam provides a broad spectrum of antibacterial activity in a convenient single formulation suitable for use in the treatment of polymicrobial infections . Possible limitations concern its restricted activity against class I beta-lactamases, enzymes that are becoming increasingly important in the nosocomial environment . Combined therapy with an aminoglycoside may be necessary in more serious infections.

Clin Neurol Neurosurg, 1994 Feb, 96(1), 52 - 7
Citrobacter meningitis in adults; Tang LM et al.; Citrobacter meningitis is an uncommon infection of neonates and young children . It is rarely seen in adults . We describe a 46-year-old man with a mixed bacterial meningitis caused by C . diversus and Klebsiella oxytoca and a 64-year-old woman with C . freundii meningitis . Review of the English-language literature revealed only 2 adult patients with C . diversus meningitis and another 2, with C . freundii meningitis . The ages of these 6 aforementioned patients ranged from 31 to 84 years . Multiple facial fractures, neurosurgical procedures, alcoholism and diabetes mellitus were predisposing conditions . Among the 5 patients whose outcome was known, antibiotic therapy was successful in 4 but failed in 1 . This study emphasizes that almost any of the gram-negative bacilli can cause serious infection of the central nervous system in adults in the proper setting.

Kansenshogaku Zasshi, 1994 Feb, 68(2), 217 - 25
{Coexisting respiratory tract infection and bacteremia or sepsis caused by the same bacterium}; Fujii T et al.; We experienced 12 patients who were suffering from bacteremia or sepsis and pneumonia or lung abscess coincidently caused by the same bacterium, during the 8 years from 1985 to 1992 a Hokusyo Central Hospital . All of them has some underlying diseases, and the average age was 73.6 years old . In ten out of 12 patients bacteremia or sepsis preceded the respiratory tract infection, and in 7 cases indwelling intravenous catheter was thought to be the port of entry of the bacteria . Respiratory tract infections were composed of pneumonia in 8 cases and lung abscess in 4 cases . Methicillin resistant Staphylococcus aureus (MRSA) was detected in 4 cases, Methicillin sensitive Staphylococcus aureus (MRSA), Klebsiella pneumoniae and Pseudomonas aeruginosa were in 2 cases respectively, Citrobacter freundii and Methicillin resistant Staphylococcus spp . were in one cases respectively . Eight out of 12 cases died in spite of current antibiotics therapy, suggesting poor prognosis.

Pediatr Emerg Care, 1994 Feb, 10(1), 23 - 5
Plantar cellulitis; Shukla PC; An unusual case of acute cellulitis of the foot in a child is reported . The child failed to respond to standard treatment even after removal of an occult foreign body . Wound cultures revealed Klebsiella oxytoca and Citrobacter freundii . This is the first documented report on Klebsiella oxytoca in cellulitis . The cellulitis resolved after being treated with the appropriate antibiotics.

Eur J Biochem, 1994 Jan 15, 219(1-2), 653 - 61
The structure of the O-specific polysaccharide of Citrobacter O16 containing glycerol phosphate; Kocharova NA et al.; The O-specific polysaccharide, obtained by mild acid degradation of Citrobacter O16 lipopolysaccharide, consists of D-glucose, D-galactose, 2-acetamido-2-deoxy-D-galactose, glycerol and phosphate in the ratios 2:2:2:1:1 . Selective cleavage of the polysaccharide was carried out by Smith degradation, N-deacetylation-deamination and dephosphorylation with 48% hydrofluoric acid, which was accompanied by unexpected splitting of one of the glycosidic linkages . The structures of the oligosaccharides thus obtained were established using 1H- and 13C-NMR spectroscopy, including one-dimensional NOE, two-dimensional rotating-frame NOE, homonuclear and heteronuclear 13C, 1H correlation spectroscopy, and, for the Smith degradation product, positive- and negative-ion-mode fast-atom-bombardment MS and MS/MS with collision-induced dissociation . On the basis of these data and the results of methylation analysis, it was concluded that the O-specific polysaccharide has the following repeating unit structure: {formula: see text}

Mol Microbiol, 1994 Jan, 11(1), 77 - 86
A new class of proteins regulating gene expression in enterobacteria; Mikulskis AV et al.; YmoA and Hha are highly similar bacterial proteins downregulating gene expression in Yersinia enterocolitica and Escherichia coli, respectively . The phenotype of ymoA mutants evokes that of mutants affected in some histone-like proteins . This paper describes complementation of a ymoA mutation in Y . enterocolitica by the hha gene from E . coli . We show that YmoA and Hha are not only very similar proteins but that they are functionally interchangeable . Genetic experiments indicate that Hha can also stimulate transposition events in vivo . By Southern blot analysis we detected hha-homologous genes at least in Citrobacter diversus, Shigella flexneri, Shigella dysenteriae, Klebsiella pneumoniae and Salmonella typhimurium . We suggest that both YmoA and Hha belong to a new family of proteins downregulating gene expression in different enterobacteria.

Infection, 1994, 22 Suppl 3, S152 - 60
The antimicrobial activity of cefotaxime: comparative multinational hospital isolate surveys covering 15 years; Jones RN; The "third-generation" cephalosporins (3GC) have emerged as one of the most significant therapeutic entities in the last 15 years . These 3GC compounds (using cefotaxime as a model) have generally maintained their potency and spectrum of activity against important pathogens . However, the continuing popularity of this class associated with local, regional, or national-level use or abuse has led to efficacy reduction against some organism populations associated with selection of Class I cephalosporinase, stably derepressed mutants predominantly among Citrobacter and Enterobacter spp.; emergence of extended-spectrum beta-lactamase producing Enterobacteriaceae (usually Klebsiella spp.), as well as some isolates mimicking Class I-type resistance patterns; and lastly, altered PBP-mediated resistances among pneumococci, Haemophilus influenzae and pathogenic Neisseria spp . Some of these resistance patterns had been present prior to the clinical introduction of 3GCs and have only significantly threatened their use in the last 5 years . Prudent application of these 3GC drugs should be the goal for this decade as follows: 1) use as monotherapy at appropriate doses and frequencies only for organisms with low potential for mutational events; 2) use combination therapy routinely for organisms such as Citrobacter, Enterobacter, some indole-positive protease and Pseudomonas aeruginosa, to minimize emerging resistance clones; 3) use conservatively in high risk patients to minimize "super-colonization" by emerging problem bacteria (e.g . vancomycin-resistant enterococci, Xanthomonas maltophilia etc.); 4) use only those agents among 3GCs that have documented safety, broad clinical applications to all age groups, acceptable pharmacokinetic features and clear cost-saving potential; and 5) use in prophylaxis (surgical procedure, selective decontamination), should be focused toward single-dose or short-course regimens to reduce total hospital-wide exposure to broad-spectrum beta-lactam drugs.(ABSTRACT TRUNCATED AT 250 WORDS)

Eur J Clin Microbiol Infect Dis, 1994, 13 Suppl 1, S2 - 11
Genetics of extended-spectrum beta-lactamases; Jacoby GA; Bacteria have adapted to the introduction of aztreonam, cefotaxime, ceftazidime, ceftriaxone and other oxyimino-beta-lactams by altering existing plasmid-mediated class A and class D beta-lactamases so as to expand their spectrum of activity . In the TEM and SHV families of extended-spectrum beta-lactamases, relative activity toward oxyimino-substrates increases with the number of amino acid substitutions but at the price of lowered intrinsic efficiency, so that compensatory up-promoter events are often associated with increased enzyme expression . Another new mechanism of resistance is the capture on plasmids of normally chromosomal genes from Enterobacter cloacae, Citrobacter freundii or Pseudomonas aeruginosa, which upon transfer can provide Klebsiella pneumoniae or Escherichia coli with resistance to alpha-methoxy-beta-lactams, such as cefoxitin or cefotetan, as well as to oxyimino-beta-lactams.

Biochemistry, 1993 Nov 30, 32(47), 12685 - 93
Structural characterization of gangliosides from resting and endotoxin-stimulated murine B lymphocytes; Portner A et al.; B lymphocytes from CBA/J mice were stimulated in splenocyte cultures for 72 h with various endotoxins . Bisphosphoryl lipid A from Escherichia coli had the highest stimulatory effect followed by LPS of Citrobacter freundii and Salmonella minnesota as measured by {3H}thymidine uptake . Gangliosides of stimulated B cells (metabolically labeled with D-{1-14C}galactose and D-{1-14C}glucosamine) and unlabeled gangliosides from resting B cells (prepared from spleens without stimulus) were analyzed by high-performance TLC, DEAE anion-exchange HPLC, and immunostaining procedures . Contents of ganglioside-derived sialic acids, quantified by HPLC as their fluorescent derivatives, decreased from stimulated to resident B lymphocytes in the following order: LPS S . minnesota > LPS C . freundii > bisphosphoryl lipid A E . coli > resting B cells . Gangliosides of resting B cells contained more N-glycolyl- than N-acetylneuraminic acid, whereas inverse ratios were found in activated cells, indicating a shift from N-glycolyl- to N-acetylneuraminic acid due to stimulation . Furthermore, a higher disialoganglioside content was characteristic for activated B cells . Fast atom bombardment mass spectrometry was performed with permethylated mono- and disialoganglioside fractions of LPS S . minnesota and LPS C . freundii stimulated B cells . Major gangliosides were GM1a and GD1a beside minute amounts of GD1b . The structural heterogeneity in the gangliosides was caused by (a) N-substitution of the sialic acids with either acetyl or glycolyl groups, (b) variation in the long-chain base (sphingosine, sphinganine), and (c) substitution of the ceramide moiety by fatty acids of different chain length and degree of unsaturation (C16:0, C24:0,24:1).2+ p6

Biochemistry, 1993 Nov 2, 32(43), 11591 - 9
Binding of phenol and analogues to alanine complexes of tyrosine phenol-lyase from Citrobacter freundii: implications for the mechanisms of alpha,beta-elimination and alanine racemization; Chen H et al.; We have examined the interaction of Citrobacter freundii tyrosine phenol-lyase with both L- and D-alanine . This enzyme catalyzes the racemization of alanine as a side reaction, in addition to the physiological beta-elimination of L-tyrosine to give phenol and ammonium pyruvate . The steady-state kinetic parameters for alanine racemization, kcat and Km, for D-alanine are 0.008 S-1 and 32 mM, respectively, while those for L-alanine are 0.03 S-1 and 11 mM . Incubation of tyrosine phenol-lyase with either L- or D-alanine forms a quinonoid complex that exhibits a strong peak at 500 nm . The presence of K+ increases the intensity of the 500-nm absorption with L-alanine, but decreases the intensity of the peak with D-alanine . Rate constants for the formation of these quinonoid intermediates and the effects of phenol and analogues on the reaction with either L- or D-alanine have been studied by rapid-scanning and single-wavelength stopped-flow spectrophotometry . Phenol binds to all the intermediates of tyrosine phenol-lyase with L- and D-alanine, but most strongly to the external aldimine complex, resulting in a decrease in the absorbance at 500 nm at equilibrium . Pyridine N-oxide binds selectively to the quinonoid complex of alanine, and thus causes an increase in the absorbance at 500 nm at equilibrium . 4-Hydroxypyridine causes a decrease in absorbance at 500 nm during the fast phase, but an increase in absorbance at 502 nm in a subsequent slow relaxation.(ABSTRACT TRUNCATED AT 250 WORDS)

Infect Immun, 1993 Nov, 61(11), 4654 - 61
The eae gene of Citrobacter freundii biotype 4280 is necessary for colonization in transmissible murine colonic hyperplasia; Schauer DB et al.; Transmissible murine colonic hyperplasia is characterized by proliferation of anchored stem cells in the mucosa of the descending colon of laboratory mice and is caused by Citrobacter freundii biotype 4280 . This bacterium produces attaching and effacing lesions in the descending colon prior to the onset of gross hyperplasia . By mutational analysis, the chromosomal eae gene of C . freundii biotype 4280 was shown to be necessary for colonic colonization . Conversely, bacteria cured of a 65-kb plasmid, which was identified in C . freundii biotype 4280, were not attenuated for colonic colonization or for the induction of colonic hyperplasia.

J Appl Bacteriol, 1993 Nov, 75(5), 427 - 34
Formation of vitamins by pure cultures of tempe moulds and bacteria during the tempe solid substrate fermentation; Keuth S et al.; The formation of water soluble vitamins (vitamin B12, vitamin B6, riboflavin, thiamine, nicotinic acid and nicotinamide) during the tempe solid substrate fermentation was investigated . The role of several strains of Rhizopus oligosporus, R . arrhizus, and R . stolonifer and the role of several bacteria in the vitamin formation process were checked . All fungal and bacterial strains were isolated from Indonesian tempe and soaking water samples . The Rhizopus strains formed riboflavin, nicotinic acid, nicotinamide and vitamin B6 . The final concentrations of these substances depended on the different strains involved and on the fermentation time . Isolates of R . oligosporus were generally the best vitamin formers . The moulds did not produce physiologically active vitamin B12 . The thiamine content decreased during fermentation . The addition of bacteria, which had been selected in a screening for vitamin B12 production, resulted in an increase of physiologically active vitamin B12 . Citrobacter freundii and Klebsiella pneumoniae showed the best formation capabilities . Furthermore, the bacteria produced riboflavin and vitamin B6 in addition to the moulds . The influence of Rhizopus on the vitamin B12 formation of Cit . freundii was also investigated . The vitamin content of tempe that was fermented with the mould and the bacterium was three times as high as a control fermentation with Cit . freundii only.

J Gen Microbiol, 1993 Nov, 139 ( Pt 11), 2715 - 22
Dependence of induction of enterobacterial AmpC beta-lactamase on cell-wall peptidoglycan, as demonstrated in Proteus mirabilis and its wall-less protoplast L-form; Tolg M et al.; The mobilizable plasmid pMD101 (ampR, ampC) was constructed by inserting cloned ampC, the structural gene for the chromosomal AmpC beta-lactamase of Citrobacter freundii, and the closely linked ampR encoding the transcriptional regulator essential for enzyme induction, into the broad host-range plasmid pKT231 . Plasmid pMD101 was transconjugated into Proteus mirabilis VI and its isogenic, cell-wall-less protoplast L-form LVI . AmpC beta-lactamase was expressed constitutively from cloned ampR and ampC in bacteria and in some L-form protoplasts . However, induction of the enzyme by beta-lactam antibiotics occurred only in bacterial cells and not in the cell-wall- and peptidoglycan-deficient L-form . In agreement with current models, induction of AmpC beta-lactamase is thought to be initiated by an induction signal arising from the metabolic disturbance of cell-wall peptidoglycan.

J Antimicrob Chemother, 1993 Nov, 32 Suppl B, 31 - 53
In-vitro activity of cefepime and other antimicrobials: survey of European isolates; Thornsberry C et al.; Cefepime, a new cephalosporin which has a broad-spectrum of activity was tested in vitro against 1961 Gram-positive and Gram-negative clinical isolates obtained from European hospitals . Cefepime was highly active against Gram-negative organisms, inhibiting over 94% of strains tested . The overall susceptibility rate for cefepime against all isolates was 81% . Cefepime was more active than any of the third-generation cephalosporins tested against species capable of producing type I beta-lactamases, e.g . Enterobacter cloacae, Citrobacter freundii, and Enterobacter aerogenes . The activity of cefepime against Pseudomonas aeruginosa was similar to that of ceftazidime and substantially greater than those of cefotaxime and ceftriaxone . All cephalosporins except ceftazidime exhibited high activity against methicillin-susceptible Staphylococcus aureus but poor activity was observed against methicillin-resistant strains . Overall susceptibility to cefepime is lower in Europe than it is in North America.

Mol Microbiol, 1993 Nov, 10(3), 555 - 65
Interactions of wild-type and mutant AmpR of Citrobacter freundii with target DNA; Bartowsky E et al.; The AmpR transcriptional activator for the chromosomal ampC beta-lactamase gene of Citrobacter freundii was found to interact with an operator sequence located in the 5' half of the 38 bp region protected by AmpR in DNase I footprinting experiments . AmpR binding was associated with significant DNA bending of target DNA . A glycine to glutamic acid alteration at position 102 in AmpR converts AmpR into a transcriptional activator even in the absence of beta-lactam inducer . AmpRG102E interacted with the operator binding sequence and induced DNA bending . A glycine to lysine alteration at residue 102 completely abolished the ability of AmpR to transcriptionally affect the ampC promoter, i.e . to repress in the absence of beta-lactam inducer and induce in the presence of beta-lactam . Nevertheless, AmpRG102K could repress the oppositely orientated ampR promoter . AmpRG102K could also specifically interact with the operator but the resulting complex migrated faster in gel retardation experiments and no significant DNA bending was observed.

FEBS Lett, 1993 Oct 11, 332(1-2), 93 - 8
A survey of a functional amino acid of class C beta-lactamase corresponding to Glu166 of class A beta-lactamases; Nukaga M et al.; The class C beta-lactamase of Citrobacter freundii GN346 is a typical cephalosporinase comprising 361 amino acids . The aspartic acid at position 217 and glutamic acid at position 219 in this beta-lactamase were, respectively, previously shown not to be the counterpart of Glu166 (ABL166) in class A beta-lactamases, even though sequence alignment of class A and C enzymes strongly suggested this possibility {(1990) FEBS Lett . 264, 211-214; (1990) J . Bacteriol . 172, 4348-4351} . We tried again to assign candidates for the counterpart of Glu166 through sequence alignment based on other criteria, the glutamic acids at positions 195 and 205 in the class C beta-lactamase being selected . To investigate this possibility, these two glutamic acids were changed to glutamine, lysine or alanine, respectively . All the mutant enzymes showed more than 50% of the activity of the wild-type enzyme, indicating that the possibility was ruled out . These results strongly suggested the possibility that the class C beta-lactamase lacks a functional acidic residue corresponding to Glu166 in class A enzymes.

Virology, 1993 Oct, 196(2), 712 - 21
The bacteriophage Mu middle operon: essential and nonessential functions; Mathee K et al.; Transcription during the bacteriophage Mu lytic cycle occurs in three phases: early, middle, and late . Previous DNA sequence analysis of the middle operon revealed five potential open reading frames (ORFs) with lengths of 39, 42, 72, 120, and 140 amino acids . The distal 140-amino-acid ORF encodes C, the activator of late transcription . Expression of the middle operon under the control of a T7 promoter and T7 RNA polymerase resulted in production of two polypeptides of 15 (ORF 120) and 16.5 kDa (C) . Introduction of a linker containing a translation terminator into ORF120 resulted in the production of a truncated form of the ORF120 polypeptide . When the ORF120 linker mutation and several middle operon deletion mutations were assayed for their effect on Mu growth in Escherichia coli K12, the deletions caused 6- to 22-min delays in lysis, and two resulted in a smaller plaque morphology, but all gave normal plating efficiencies and burst sizes . The plating efficiencies for all the mutants were also similar to that of wild-type Mu on alternate hosts E . coli C, Citrobacter freundii, Shigella sonnei, and Shigella flexneri . These results indicate that, with the exception of C, the middle operon ORFs are not essential for phage development.

J Antimicrob Chemother, 1993 Oct, 32(4), 595 - 603
Characterization of ceftriaxone-resistant Enterobacteriaceae: a multicentre study in 26 French hospitals . Vigil'Roc Study Group; Goldstein FW et al.; During a multicentre study performed in 26 French hospitals, 287 (3.2%) of 9038 Enterobacteriaceae isolated, mainly Enterobacter spp., Serratia spp., Citrobacter spp . and Klebsiella spp . were classified as ceftriaxone resistant on the basis of an MIC > 4 mg/L or the presence of an extended-spectrum beta-lactamase . Extended-spectrum beta-lactamase was present mainly in Klebsiella pneumoniae (65 strains, 10.2%) and very rarely in Escherichia coli, Proteus mirabilis, Klebsiella oxytoca, Citrobacter spp . and Enterobacter spp . The extended-spectrum beta-lactamases conferred low-level resistance to ceftriaxone in nearly 60% of the strains harbouring them, emphasizing the need for routine testing for the presence of these enzymes . Among transconjugants three types of extended-spectrum beta-lactamase were identified . Those resembling TEM-3 were the most common, but TEM-21, and SHV-4 were also found . Clavulanate and to a lesser extent sulbactam inhibited all the extended-spectrum beta-lactamases encountered in this study.

Mol Cell Probes, 1993 Oct, 7(5), 345 - 8
Oligonucleotide probe determination of tetracycline-resistant bacteria isolated from catfish ponds; DePaola A et al.; Oligonucleotide probes for class A, B and C tetracycline resistance determinants were synthesized and tested individually and in combination for hybridization with Gram-negative isolates previously characterized by restriction fragment DNA probes . Similar hybridization patterns were observed with either probe type . Gram-negative catfish pond bacteria were characterized by these oligonucleotide probes . The most common bacteria were Plesiomonas shigelloides (47.7%), Aeromonas hydrophila (28.8%) and Citrobacter freundii (13.3%) . The class A (46.4%) tetracycline resistance determinant was more prevalent than B (19.8%) and C (0.3%) . Some tetracycline-resistant isolates of each species, including most A . hydrophila (62.1%) isolates, failed to hybridize with any of the probes.

Jpn J Antibiot, 1993 Oct, 46(10), 860 - 76
{Antimicrobial activity of cefodizime against fresh clinical isolates}; Deguchi K et al.; In order evaluate antimicrobial activities of cefodizime (CDZM), minimum inhibitory concentrations (MIC's) of CDZM and other control drugs were determined against various clinical isolates, that were sent to our center from nation-wide medical institutions or were isolated and identified in our laboratory from various specimens of infected patients . The followings are a summary of the results: 1 . Bacterial species with no or few strains resistant to cephems including CDZM included Streptococcus pyogenes, Haemophilus influenzae, Citrobacter diversus, most of Klebsiella pneumoniae and Proteus mirabilis . Some strains of Klebsiella oxytoca were resistant to cephems increases in beta-lactams resistant Streptococcus pneumoniae and cephem resistant Escherichia coli seemed likely . Among Citrobacter freundii, Enterobacter spp., Serratia marcescens, Proteus vulgaris, Morganella morganii and Providencia spp . belonging to a category of so-called "mildly toxic bacteria", high portions of the strains examined were resistant to cephems including CDZM and these strains were also resistant to new quinolones, thus they showed multiple drug resistance . 2 . MIC90's of CDZM against Streptococcus spp., H . infleunzae, Moraxella subgenus Branhamella catarrhalis, E . coli, Klebsiella spp . and P . mirabilis, frequently found in daily treatment of infections, were less than < or = 0.025 to 1.56 micrograms/ml . This indicates that CDZM would be expected to have enough antibiotic activity in infections caused by above mentioned bacteria . However, cautions are needed in the treatment of infections by beta-lactam resistant S . pneumoniae, cephem resistant E . coli and cephem resistant K . oxytoca with CDZM . 3 . Among the above mentioned "mildly toxic bacteria", many multiple drug resistant strains exist . Therefore, we evaluated an usefulness of concomitant use of CDZM with aminoglycosides in the treatment of infections by these bacteria, using other reports which indicates the usefulness in vitro and in vivo . 4 . Antibacterial activities of CDZM we observed in this study seem to indicate that CDZM concentrations in infected areas are maintained at above MIC levels for relatively long periods of time.

Int J Syst Bacteriol, 1993 Oct, 43(4), 645 - 58
Classification of citrobacteria by DNA hybridization: designation of Citrobacter farmeri sp . nov., Citrobacter youngae sp . nov., Citrobacter braakii sp . nov., Citrobacter werkmanii sp . nov., Citrobacter sedlakii sp . nov., and three unnamed Citrobacter genomospecies; Brenner DJ et al.; DNA relatedness studies (hydroxyapatite method) were done on 112 strains of citrobacteria . By using the recommended definition of a genomospecies 11 genomospecies were identified in the genus Citrobacter . These genomospecies were separable by their biochemical profiles . Citrobacter koseri (Citrobacter diversus) and Citrobacter amalonaticus proved to be homogeneous species, as previously described . C . amalonaticus biogroup 1, as described by Farmer et al . (J . Clin . Microbiol . 21:46-76, 1985), was shown to be a separate homogeneous species, which was named Citrobacter farmeri sp . nov . The Citrobacter freundii complex was quite heterogeneous . C . freundii sensu stricto, as represented by the type strain, contained only 9 of 66 strains in this complex . The remaining 57 strains were members of seven genomospecies . Genomospecies 5, containing 21 strains, was named Citrobacter youngae sp . nov . Genomospecies 6, containing 15 strains, was named Citrobacter braakii sp . nov . Genomospecies 7 and 8, each containing six strains, were named Citrobacter werkmanii sp . nov . and Citrobacter sedlakii sp . nov., respectively . Genomospecies 9, 10, and 11, each containing three strains, were not named.

J Bacteriol, 1993 Sep, 175(18), 5882 - 9
Cloning and sequence analysis of a gene (pchR) encoding an AraC family activator of pyochelin and ferripyochelin receptor synthesis in Pseudomonas aeruginosa; Heinrichs DE et al.; Pseudomonas aeruginosa K372 is deficient in the production of both the 75-kDa ferripyochelin receptor protein and pyochelin . A 1.8-kb EcoRI-SalI fragment which restored production of both the receptor protein and pyochelin was cloned . Nucleotide sequencing of the fragment revealed an open reading frame of 888 bp, designated pchR (pyochelin), capable of encoding a 296-amino-acid protein of a 32,339-Da molecular mass . By using a phage T7-based expression system, a protein of ca . 32 kDa was produced off the 1.8-kb fragment, confirming that this open reading frame was indeed expressed . A region exhibiting homology to the consensus Fur-binding site of Escherichia coli was identified upstream of the pchR coding region overlapping a putative promoter . In addition, the C-terminal 80 amino acid residues of PchR showed approximately 50% homology (identity, 31%; conserved changes, 19%) to the carboxy terminus of AraC, a known transcriptional activator of gene expression in E . coli, Salmonella typhimurium, Citrobacter freundii, and Erwinia chrysanthemi . Within the C-terminal region of PchR, AraC, and a number of other members of the AraC family of transcriptional activators, there exists a highly conserved 17-residue domain where, in fact, two residues are strictly maintained and two others exhibit only conserved changes, suggesting a common functional significance to this region in all of these proteins . These data are consistent with a role for PchR as a transcriptional activator of pyochelin and ferripyochelin receptor synthesis in P . aeruginosa . In agreement with this, a PchR mutant obtained by in vitro mutagenesis and gene replacement was deficient in production of the ferripyochelin receptor and pyochelin.

Zentralbl Hyg Umweltmed, 1993 Sep, 194(5-6), 540 - 52
{Clonal spreading of a multidrug resistance Citrobacter freundii strain at a neonatal intensive care unit}; Gericke B et al.; This paper reports on the epidemic spreading of a multiresistant Citrobacter (C.) freundii strain at a neonatal intensive care unit . A premature baby of the 27th week of pregnancy died from septic shock caused by this strain . According to the result of a statistical analysis of risk factors a connection between the colonization of neonates and the feeding with an enteral feeding tube was probably . This suspicion could be confirmed by the detection of the multiresistant strain in the infant formula . Plasmid analysis, examination of outer membrane proteins and lipopolysaccharides of these C . freundii strains demonstrated the identity of all isolates . The resistance to antibiotics of this multiresistant C . freundii clone was determined by a plasmid belonging to incompatibility group C with a molecular weight of 110 MDa . This plasmid was involved also in other nosocomial outbreaks . It persisted more than 7 years in the hospital flora of the intensive care unit.

Jpn J Antibiot, 1993 Sep, 46(9), 801 - 17
{The antimicrobial activity of cefteram against recently clinically detected and isolated strains}; Deguchi K et al.; In order to examine antibiotic activities of cefteram (CFTM), its minimum inhibitory concentrations (MIC's) and those of other cephem drugs were determined against clinically isolated strains received from July 1990 to June 1991 and from July 1992 to February 1993 from medical facilities throughout the country and against clinically isolated strains detected in our laboratory in samples from patients with various infectious diseases . The obtained results are summarized below . 1 . No CFTM-resistant strains were found among beta-streptococci, Klebsiella spp., Proteus mirabilis, Haemophilus influenzae, and Neisseria gonorrhoeae or even when found, they were present in extremely low proportions . 2 . It appeared that Streptococcus pneumoniae insensitive or resistant to beta-lactams, as well as cephems-resistant strains of Escherichia coli were increasing . The former included benzylpenicillin (PCG)-insensitive S . pneumoniae (PISP) of PCG-resistant S . pneumoniae (PRSP), and the presence of the latter suggests the possibility of the existence of "Extended broad-spectrum beta-lactamase" producing strains . The MIC's of beta-lactams against the above PISP or PRSP, and against cephems-resistant E . coli tended to be high, but those of CFTM were relatively low (in most cases) . 3 . Proportions of strains resistant to cephems, including CFTM among Citrobacter spp., Enterobacter spp., Serratia marcescens, Proteus vulgaris, Morganella morganii, and Providencia rettgeri were high, and in addition, the existence of these cephem resistant species suggests an increase in multiple drug resistant strains that show resistance to new quinolone drugs . 4 . As mentioned above, CFTM is by no means a perfect drug or utility drug and its antimicrobial activities do not cover some recent isolates with multiple drug resistance . Except problems encountered with so-called "attenuated" strains of bacteria, increases in resistance can only be observed at a level of MIC90's, and as far as MIC80's are concerned, CFTM still is as active as before and may be used in the treatment of most infections we encounter in normal medical practices.

Clin Infect Dis, 1993 Sep, 17(3), 437 - 40
Recurring ventriculitis due to Citrobacter diversus: clinical and bacteriologic analysis; Eppes SC et al.; Neonatal meningitis due to Citrobacter diversus is usually accompanied by the development of brain abscesses and tends to have high rates of morbidity and mortality . We report a case of a newborn with C . diversus meningitis and brain abscesses who relapsed after initial antibiotic therapy and from whom C . diversus was recovered from cerebrospinal fluid 4 years later during a neurosurgical procedure . Genetic differences in the early and late isolates were discovered, and explanations for this phenomenon are suggested . Possible mechanisms for prolonged persistence in the central nervous system are explored . This unusual case and the analysis of the organisms illustrate the unique features of C . diversus as a pathogen and underscore the need for developing optimal therapeutic strategies.

Intern Med, 1993 Sep, 32(9), 719 - 21
Spontaneous bacterial peritonitis in an adult patient with nephrotic syndrome; Kato A et al.; Most cases of spontaneous bacterial peritonitis (SBP) in association with nephrotic syndrome are children . The complication of SBP in adults with nephrotic syndrome is extremely rare . Herein, we report a 25-year-old man with nephrotic syndrome and chronic renal failure who suffered from SBP . Citrobacter freundii was isolated from ascites . Irreversible deterioration of renal function followed the development of SBP, though the peritonitis was cured with antibiotic treatment . This case suggests that SBP is a rare, but serious complication of adult nephrotic syndrome with ascites.

J Clin Microbiol, 1993 Sep, 31(9), 2263 - 73
DNA-based diagnostic tests for Salmonella species targeting agfA, the structural gene for thin, aggregative fimbriae; Doran JL et al.; Salmonella enteritidis 27655-3b and a few diarrheagenic Escherichia coli strains produce morphologically and antigenically related, thin, aggregative fimbriae, collectively named GVVPQ fimbriae (S . K . Collinson, L . Emody, T . J . Trust, and W . W . Kay, J . Bacteriol . 174:4490-4495, 1992) . To determine whether GVVPQ fimbriae are common to Salmonella spp . and other enteropathogenic members of the family Enterobacteriaceae, 113 isolates were phenotypically screened for Congo red binding and aggregative colony morphology . Presumptive positive and representative negative strains were examined by Western blotting (immunoblotting) by using antiserum to SEF 17, the native GVVPQ fimbria of S . enteritidis . Only four S . enteritidis strains and six E . coli isolates possessed substantial amounts of GVVPQ fimbriae after 24 h of incubation on T medium . Following 5 days of incubation, 56 of 93 Salmonella isolates (60%) and 1 of 7 additional E . coli clinical isolates possessed detectable levels of GVVPQ fimbriae . Since variable expression of GVVPQ fimbriae was observed among Salmonella isolates and some E . coli strains produced scant amounts, as revealed by immunoelectron microscopy, the ability to produce these fimbriae was evaluated by genotypic screening . The structural gene for the SEF 17 fimbrin, agfA, was amplified by the polymerase chain reaction, cloned, and sequenced to provide a characterized DNA probe . An agfA DNA fragment hybridized strongly to 603 of 604 (99.8%) Salmonella isolates but very weakly to 31 of 266 other members of the family Enterobacteriaceae including 26 of 137 E . coli strains, 3 of 14 Citrobacter spp., and single isolates of Shigella sonnei and Enterobacter cloacae . The agfA DNA probe proved to be a valuable diagnostic tool for Salmonella isolates arrayed on hydrophobic grid membrane filters . Unique agfA sequences were targeted in the development of a polymerase chain reaction assay specific for Salmonella spp.

J Trop Pediatr, 1993 Aug, 39(4), 230 - 3
Bacteriology of neonatal septicaemia in a rural referral hospital in south India; Rao PS et al.; Out of 640 suspected cases of neonatal septicaemia studied, bacteraemia was detected in 255 (40 per cent) of the infants . Gram negative organisms were predominant (56 per cent) with Pseudomonas, Citrobacter, and Klebsiella as the commonest pathogens . Among the Gram positive organisms both Staphylococcus aureus and Staphylococcus epidermidis were equally prevalent . Staphylococci were mainly responsible for early onset infections, whereas Salmonella typhimurium and Pseudomonas were the main organisms in late onset infections . Group B streptococcal infection was not encountered in this part of the country.

Appl Environ Microbiol, 1993 Aug, 59(8), 2713 - 6
Monoclonal antibodies that react with live Listeria spp; Torensma R et al.; Seven monoclonal antibodies (MAbs) against Listeria spp . that were reactive with live Listeria spp . were developed . Two of these MAbs (55-8 and 55-37) were members of the immunoglobulin M class, and all other MAbs were members of the immunoglobulin G class . MAb 55-23 reacted with 148 of 157 strains tested . MAb 34-51 reacted with serotype 1/2a, 1/2b, and 1/2c strains and exhibited a scattered reaction pattern with strains belonging to other serotypes . MAb 55-44 reacted with all of the strains belonging to serotype 4b tested . MAb 55-4 reacted with all of the serotype 1/2a isolates tested, although reactivity with other isolates also was observed . The other MAbs exhibited scattered reaction patterns . No correlation of reactivity pattern with serotype was found . Marked differences were observed between the reactivities of MAbs as determined by a magnetic immunoluminescence assay and a whole-cell enzyme-linked immunosorbent assay . Only MAb 55-23 exhibited minor reactivity with three Streptococcus spp . isolates, while no reactivity was observed with six Bacillus spp . strains, one Escherichia coli strain, and one Citrobacter sp . strain . In Western blots (immunoblots) MAbs 55-23, 55-44, and 34-9 exhibited reactivity; all other MAbs were negative in this assay.

Appl Environ Microbiol, 1993 Aug, 59(8), 2602 - 6
Lysine-mannitol-glycerol agar, a medium for the isolation of Salmonella spp., including S . typhi and atypical strains; Cox JM; An agar medium for the isolation of Salmonella spp . is described . The medium, lysine-mannitol-glycerol agar, has features of both xylose-lysine-deoxycholate agar and mannitol-lysine-crystal violet-brilliant green agar, but glycerol is added for the differentiation of Salmonella and Citrobacter spp . The medium facilitates the detection of strains having atypical fermentation patterns, such as the lactose- or sucrose-positive salmonellae . The medium also detects Salmonella typhi after enrichment.

Appl Environ Microbiol, 1993 Aug, 59(8), 2531 - 7
Copper-resistant enteric bacteria from United Kingdom and Australian piggeries; Williams JR et al.; Thirty-three enteric isolates from Australian (Escherichia coli only) and United Kingdom (U.K.) (Salmonella sp., Citrobacter spp., and E . coli) piggeries were characterized with respect to their copper resistance . The copper resistance phenotypes of four new Australian E . coli isolates were comparable with that of the previously studied E . coli K-12 strain ED8739(pRJ1004), in that the resistance level in rich media was high (up to 18 mM CuSO4) and resistance was inducible . Copper resistance was transferable by conjugation from the new Australian isolates to E . coli K-12 recipients . DNA similarity between the new Australian isolates and the pco copper resistance determinant located on plasmid pRJ1004 was strong as measured by DNA-DNA hybridization; however, the copper resistance plasmids were nonidentical as indicated by the presence of restriction fragment length polymorphisms between the plasmids . DNA-DNA hybridization and polymerase chain reaction analysis demonstrated DNA homology between the pco determinant and DNA from the U.K.E . coli, Salmonella sp., and Citrobacter freundii isolates . However, the copper resistance level and inducibility were variable among the U.K . strains . Of the U.K . E . coli isolates, 1 demonstrated a high level of copper resistance, 4 exhibited intermediate resistance, and 16 showed a low level of copper resistance; all of these resistances were expressed constitutively . A single U.K . C . freundii isolate, had a high level of copper resistance, inducible by subtoxic levels of copper . Transconjugants from one E . coli and one C . freundii donor, with E . coli K-12 strain UB1637 as a recipient, showed copper resistance levels and inducibility of resistance which differed from that expressed from plasmid pRJ1004.(ABSTRACT TRUNCATED AT 250 WORDS)

J Infect Dis, 1993 Aug, 168(2), 484 - 8
A multidrug resistance regulatory chromosomal locus is widespread among enteric bacteria; Cohen SP et al.; Constitutive expression of the mar operon (marRAB) in Escherichia coli produces a multiple antibiotic resistance phenotype mediated by the expression of multiple genetic loci in response to regulatory proteins in the operon . A mar-specific DNA probe was used to search for the operon in bacterial strains representing 53 species and 27 genera . Among these, 6 other Enterobacteriaceae, Salmonella, Shigella, Klebsiella, Citrobacter, Hafnia, and Enterobacter species, contained DNA sequences that hybridized to the probe under high-stringency conditions . By use of a selection protocol developed to obtain multiple antibiotic resistant mutants of E . coli, multiply resistant mutants that showed increased expression of mar-specific RNA were obtained from Enterobacter agglomerans and Salmonella species.

Mol Microbiol, 1993 Aug, 9(4), 703 - 15
AmpG, a signal transducer in chromosomal beta-lactamase induction; Lindquist S et al.; The chromosomal ampC beta-lactamase in Citrobacter freundii and Enterobacter cloacae is inducible by beta-lactam antibiotics . When an inducible ampC gene is introduced on a plasmid into Escherichia coli together with its transcriptional regulator ampR, the plasmid-borne beta-lactamase is still inducible . We have isolated mutants, containing alterations in a novel E . coli gene, ampG, in which a cloned C . freundii ampC gene is unable to respond to beta-lactam inducers . The ampG gene was cloned, sequenced and mapped to minute 9.6 on the E . coli chromosome . The deduced amino acid sequence predicted AmpG to be a 53 kDa, transmembrane protein, which we propose acts as a signal transducer or permease in the beta-lactamase induction system . Immediately upstream of ampG there is another 579-base-pair-long open reading frame (ORF) encoding a putative lipoprotein shown to be non-essential for beta-lactamase induction . We have found that ampG and this ORF form an operon, whose promoter is located in front of the ORF . Located closely upstream of the putative promoter is the morphogene bolA, which is transcribed in the opposite orientation . However, using transcription fusions, we have found that the ampG transcription is not regulated by bolA . In addition, we show that transcription is probably not regulated by either the starvation specific sigma factor RpoS, which controls bolA, or by AmpD the negative regulator for ampC transcription.

Antimicrob Agents Chemother, 1993 Aug, 37(8), 1696 - 700
Multicenter comparison of in vitro activities of FK-037, cefepime, ceftriaxone, ceftazidime, and cefuroxime; Washington JA et al.; In a multicenter study, the MICs of FK-037 for 90% of the strains tested (MIC90s) were < or = 1 microgram/ml for members of the family Enterobacteriaceae other than Citrobacter freundii, Enterobacter spp., and Serratia marcescens . Activity against Pseudomonas aeruginosa was variable, with a MIC50 and a MIC90 of 4 and 32 micrograms/ml, respectively . Relative to cefepime, however, FK-037 was less active against ceftazidime-resistant isolates of Enterobacter cloacae . The MIC90 of FK-037 for methicillin-resistant staphylococci was > 16 micrograms/ml.

Gene, 1993 Jul 15, 129(1), 77 - 81
Cloning and sequences of the genes encoding the CfrBI restriction-modification system from Citrobacter freundii; Zakharova MV et al.; The genes encoding the CfrBI restriction and modification (R-M) systems from Citrobacter freundii and recognizing the sequence 5'-CCWWGG-3' (W = A or T) were cloned in Escherichia coli McrBC- cells . The nucleotide (nt) sequences of the genes were determined . Two large open reading frames were found . Deletion analysis showed that one of them {1128 nt coding for 376 amino acids (aa)} corresponds to a methyltransferase (MTase)-encoding gene and the other (1065 nt coding for 355 aa) to a restriction endonuclease-encoding gene . The genes are oriented divergently and separated by 76 bp . A CfrBI site (5'-m4CCATGG) was found in the intergenic region of the cfrBIRM genes . Analysis of the deduced aa sequence of M.CfrBI made it possible to determine the typical features of a m4C-specific MTase . Limited homology between the M.CfrBI and R.CfrBI proteins was also found.

Antimicrob Agents Chemother, 1993 Jul, 37(7), 1547 - 51
In vitro antibacterial activity and beta-lactamase stability of CP-70,429 a new penem antibiotic; Minamimura M et al.; In in vitro susceptibility tests, the new penem CP-70,429 showed potent antibacterial activity against gram-positive and gram-negative bacteria except Pseudomonas aeruginosa and Xanthomonas maltophilia . CP-70,429 was stable to various types of beta-lactamases except for the enzyme from X . maltophilia and was 16- to 128-fold more active than the other compounds against beta-lactamase-producing strains of Enterobacter cloacae and Citrobacter freundii.

J Clin Microbiol, 1993 Jul, 31(7), 1946 - 8
Evaluation of the L-pyrrolidonyl-beta-naphthylamide hydrolysis test for the differentiation of members of the families Enterobacteriaceae and Vibrionaceae; Chagla AH et al.; A simple, rapid, and inexpensive spot test incorporating the substrate pyrrolidonyl naphthylamide was used to examine pyrrolidonyl peptidase activity among 800 bacterial strains belonging to the families Enterobacteriaceae and Vibrionaceae . The pyrrolidonyl naphthylamide test was found to be particularly useful in separating Citrobacter spp . (100% positive) from Salmonella spp . (0.4% positive) and Escherichia coli (0% positive) . Furthermore, it would appear to offer a safer alternative to the traditional potassium cyanide test for differentiating citrobacters from salmonellae.

J Clin Microbiol, 1993 Jul, 31(7), 1927 - 31
Whole-cell repetitive element sequence-based polymerase chain reaction allows rapid assessment of clonal relationships of bacterial isolates; Woods CR et al.; Repetitive element sequence-based polymerase chain reaction (rep-PCR) enables the generation of DNA fingerprints which discriminate bacterial species and strains . We describe the application of whole-cell methods which allow specimens from broth cultures or colonies from agar plates to be utilized directly in the PCR mixture . The rep-PCR-generated DNA fingerprints obtained with whole-cell samples match results obtained with genomic DNA templates . Examples with different gram-negative bacteria (e.g., Citrobacter diversus, Escherichia coli, and Pseudomonas aeruginosa) and gram-positive bacteria (e.g., Staphylococcus aureus and Streptococcus pneumoniae) are demonstrated . Rapid specimen preparation methods enable rep-PCR-based fingerprinting to be completed in several hours and, therefore, allows the timely analysis of epidemiological relationships.

Am J Clin Pathol, 1993 Jul, 100(1), 47 - 51
Emergence of antimicrobial resistance in gram-negative bacilli causing bacteremia during therapy; Siebert JD et al.; Treatment of serious infections caused by gram-negative bacilli with beta-lactam antimicrobial agents can induce Class I beta-lactamase production . This phenomenon can result in resistant microorganisms, and has been postulated to be a cause of therapeutic failure . The charts of patients bacteremic with Pseudomonas aeruginosa, Serratia marcescens, Enterobacter cloacae, Citrobacter freundii, Proteus vulgaris, and Providencia species (n = 120) during a 3-year period were reviewed to determine how common the emergence of resistance was, and to determine if in vitro susceptibility testing was a reliable therapeutic guide . Emergence of resistance was believed to occur when a subsequent bacteremic isolate showed at least a fourfold increase in minimum inhibitory concentration accompanied by a change of interpretive susceptibility category . In the group of patients who survived at least 48 hours that received beta-lactam therapy (n = 76), one case of emergence of resistance was identified (1.3%) . Emergence of resistance to beta-lactam antimicrobial agents did not commonly cause therapeutic failure at our institution, and susceptibility testing of gram-negative bacilli by usual methods was a reliable guide to antimicrobial therapy.

J Clin Pathol, 1993 Jul, 46(7), 637 - 41
Multipoint identification of Enterobacteriaceae: report of the British Society for Microbial Technology collaborative study; Winstanley TG et al.; AIMS--To evaluate the accuracy and reproducibility of multipoint identification schemes in a multicentre trial . METHODS--Forty two strains of Enterobacteriaceae were distributed to 22 laboratories for identification by routine multipoint methods . Analysis of results enabled inter- and intralaboratory reproducibility of a variety of tests, and the ability of laboratories to identify individual organisms to be determined . RESULTS--Interlaboratory reproducibility of most of the biochemical tests was acceptable . The least reproducible tests, both within and between laboratories, were citrate utilisation, production of urease and beta galactosidase, detection of motility, and decarboxylation of lysine and ornithine . Inconsistent results for these tests were often associated with misidentified strains . Most laboratories performed identifications satisfactorily . Most isolates (72.1%) were identified correctly to species level; 9.6% were incorrectly identified, and 6.4% could not be identified at all . The most difficult organisms to identify were Citrobacter freundii, Enterobacter cloacae, Hafnia alvei and Aeromonas hydrophila . Strains of Enterobacter, Serratia sp, and Providencia sp were difficult to speciate . Several laboratories could not identify organisms exhibiting at least one atypical biochemical reaction . CONCLUSION--This study emphasises the need for quality control of media and reagents for multipoint identification of Gram negative enteric bacilli.

Infect Immun, 1993 Jun, 61(6), 2486 - 92
Attaching and effacing locus of a Citrobacter freundii biotype that causes transmissible murine colonic hyperplasia; Schauer DB et al.; Citrobacter freundii biotype 4280 produces attaching and effacing (AE) lesions in the large intestine of laboratory mice and is the causative agent of transmissible murine colonic hyperplasia . AE lesions are also produced by enteropathogenic Escherichia coli in humans . Southern analysis revealed that biotype 4280, but not 20 other strains of C . freundii, contained DNA homologous to the eae (E . coli attaching and effacing) gene which is necessary for AE activity by enteropathogenic E . coli in vitro . We have cloned and determined the nucleotide sequence of the C . freundii eae homolog . Our findings suggest that the eae locus of C . freundii biotype 4280 is necessary for AE activity and has a role in the pathogenesis of transmissible murine colonic hyperplasia.

J Gen Microbiol, 1993 Jun, 139 ( Pt 6), 1245 - 52
beta-Lactamase of Lysobacter enzymogenes: cloning, characterization and expression of the gene and comparison of the enzyme to other lactamases; Boras GJ et al.; The gene for the periplasmic beta-lactamase of Lysobacter enzymogenes was isolated as part of a 1017 bp EcoRI fragment and the nucleotide sequence of the gene was determined . It has a G+C content of 71.5% and encodes a 27 amino acid signal sequence and the mature beta-lactamase of 276 amino acids which has a mass of 29,146 Da . The enzyme appears to be unique to L . enzymogenes but its amino acid sequence shows a high degree of homology with the amino acid sequences of the lactamase from Citrobacter diversus and other Class A beta-lactamases . The beta-lactamase gene of L . enzymogenes was expressed in Escherichia coli using pUC118 as the vector . The production of active beta-lactamase was highest after the active growth phase of the expression host and reached levels which were about three times higher than those obtained with L . enzymogenes.

Acta Paediatr, 1993 Jun-Jul, 82(6-7), 530 - 2
The use of imipenem-cilastatin in neonatal meningitis caused by Citrobacter diversus; Haimi-Cohen Y et al.; Citrobacter diversus is a cause of severe meningitis in neonates and infants . It is unique in its propensity to produce brain abscesses that play an important role in the poor prognosis associated with this condition . The recommended therapeutic regimen of third-generation cephalosporines, aminoglycosides and trimethoprim-sulfamethoxazole is usually disappointing . We describe the use of imipenemcilastatin in successfully treating Citrobacter diversus meningitis complicated by brain abscesses.

Mikrobiol Z, 1993 Jun-Aug, 55(4), 75 - 81
{The pathogenicity factors in different representatives of the genus Citrobacter}; Semenova EA et al.; No trustworthy distinctions were revealed under the comparative analysis of properties of bacteria of Citrobacter genus from 3 studied ecological groups (from patients with diarrhea, from healthy persons and patients with extraenteric pathological process) . It allows the finding of potentially partogenic bacteria of Citrobacter genus in patients with diarrhea to be considered the manifestation of dysbacteriosis.

Mol Phylogenet Evol, 1993 Jun, 2(2), 97 - 111
Phylogenetic tree and sequence similarity of beta-lactamases; Ogawara H; beta-Lactamases are the main cause of beta-lactam resistance in many pathogenic bacteria . These enzymes can be detected in a variety of pathogenic as well as non-pathogenic bacteria . The cyanobacteria are also known to produce a beta-lactamase . Recently, the amino acid sequences and the three-dimensional structures of some of these beta-lactamases have been clarified . On the basis of the amino acid sequences of 47 beta-lactamases and the computer-aided analysis, a phylogenetic tree is proposed in this paper . According to the tree, beta-lactamases are classified into six groups . Group 1 beta-lactamases are mainly composed of plasmid-mediated enzymes from gram-negative bacteria . However, chromosome-derived beta-lactamases from Klebsiella pneumoniae and Rhodopseudomonas capsulata take part in this group . Group 2 enzymes consist of a part of the chromosome-encoded beta-lactamases from Streptomyces, and chromosome-mediated enzymes from Yersinia enterocolitica, Citrobacter diversus, and Klebsiella oxytoca . Chromosome-encoded beta-lactamases from gram-negative bacteria form group 3 . Group 4 is composed of metalloenzymes, whereas group 5 consists of OXA type beta-lactamases . Chromosome-encoded beta-lactamases from gram-positive bacteria form group 6 . Comparison of the amino acid sequences among these groups confirmed the phylogenetic tree and the classification: the beta-lactamases in each group have its particular conserved amino acid sequences . In addition, the tree provides more detailed classification and time-scale mutual relationships and predicts new types of beta-lactamases that may be found . Furthermore, the classification deduced from the tree is generally in accord with the one based on the amino acid sequences reported previously . However, the class A beta-lactamases are clearly divided into three groups: groups 1, 2, and 6 . RDF2 analysis shows that some combinations between beta-lactamases and beta-lactam-interacting proteins as well as eukaryotic proteins have a low but significant evolutionary relatedness.

Appl Environ Microbiol, 1993 May, 59(5), 1473 - 9
Specific detection of Salmonella spp . by multiplex polymerase chain reaction; Way JS et al.; Three sets of oligonucleotide primers were used in the polymerase chain reaction (PCR) assay to detect Salmonella species . phoP primers specific to the phoP/phoQ loci of coliform pathogenic bacteria such as Salmonella, Shigella, Escherichia coli, and Citrobacter species served as presumptive indicators of enteric bacteria . In addition to the phoP primers, the Hin and the H-1i primers, which targeted a 236-bp region of hin/H2 and a 173-bp region of the H-1i flagellin gene, respectively, were used . Both Hin and H-1i primers are specific to motile Salmonella species and are not present in Shigella, E . coli, or Citrobacter species . Thus, by multiplex PCR amplification, Salmonella species including Salmonella typhi, Salmonella typhimurium, Salmonella paratyphi A, and Salmonella enteritidis can be specifically detected . Optimal reaction conditions have been described to demonstrate this specific, sensitive detection of Salmonella species . By using agarose gel electrophoresis for detection of the PCR-amplified products, the sensitivity of detection was 10(2) CFU after 25 cycles of PCR and 1 (10(0)) CFU after a 50-cycle double PCR . The efficacy of these primers was demonstrated on environmental isolates which had previously been confirmed as Salmonella species by the use of conventional cultural techniques . In addition, positive amplifications resulted from Salmonella species in environmental samples including soil and water.

Eur J Clin Microbiol Infect Dis, 1993 May, 12(5), 377 - 84
Comparative in vitro activity of biapenem, a new carbapenem antibiotic; Clarke AM et al.; Biapenem is a new carbapenem antibiotic with high stability to human renal dehydropeptidase I . Its in vitro activity was compared with that of imipenem, meropenem, ceftazidime, ceftriaxone, piperacillin and gentamicin against a total of 650 recent clinical isolates . MICs were determined by a standard agar dilution procedure and all isolates were tested at two inocula (10(4) and 10(6) cfu) . Biapenem inhibited 90% of isolates of Escherichia coli, Klebsiella spp., Proteus mirabilis, Proteus vulgaris, Morganella morganii, Providencia stuartii, Enterobacter spp., Citrobacter freundii, Serratia marcescens, Salmonella typhi, Shigella sonnei and Yersinia enterocolitica at < or = 2 mg/l and was as active as or two- to four-fold more active than imipenem against all these species, with the exception of Serratia marcescens, against which imipenem was two-fold more active . Biapenem was two-fold more active than imipenem against Pseudomonas aeruginosa (MIC90 4 mg/l) and had activity similar to that of imipenem against the Bacteroides fragilis group (MIC90 0.5 mg/l) but was two-fold less active than imipenem against methicillin-susceptible Staphylococcus aureus (MIC90 0.06 mg/l) and was, like imipenem, inactive against methicillin-resistant Staphylococcus aureus.

J Appl Bacteriol, 1993 May, 74(5), 583 - 7
Structure and antigenic properties of Citrobacter freundii lipopolysaccharides; Chart H et al.; Citrobacter freundii strain E69366 was detected in a colony immunoassay with a rabbit antiserum prepared to a strain of Escherichia coli belonging to serogroup O157 . Lipopolysaccharide (LPS) was shown to contain the epitope(s) involved in antibody binding . LPS prepared from strain E69366, with hot phenol contained core-LPS that migrated to the water-phase and long-chain LPS that separated into the phenol-phase . A total of 36 strains of Cit . freundii were analysed for their LPS profiles by SDS-PAGE . Sixteen could be allocated into five groups (A(3), B(7), C(2), D(2) and E(2)) based on similarities in LPS profile . The remaining strains either expressed unique SDS-PAGE profiles (18) or did not express long-chain LPS (2) . All strains with a profile 'A' reacted with the antiserum prepared to E . coli O157, whereas those with other profiles did not . The two strains of Cit . freundii expressing LPS profiles designated 'E' reacted with an antiserum prepared in rabbits to E . coli O45.

Biochemistry, 1993 Apr 27, 32(16), 4195 - 206
Three-dimensional structure of tyrosine phenol-lyase; Antson AA et al.; Tyrosine phenol-lyase (EC 4.1.99.2) from Citrobacter freundii has been cloned and the primary sequence deduced from the DNA sequence . From the BrCN digest of the NaBH4-reduced holoenzyme, five peptides were purified and sequenced . The amino acid sequences of the peptides agreed with the corresponding parts of the tyrosine phenol-lyase sequence obtained from the gene structure . K257 is the pyridoxal 5'-phosphate binding residue . Assisted by the sequence data, the crystal structure of apotyrosine phenol-lyase, a pyridoxal 5'-phosphate-dependent enzyme, has been refined to an R-factor of 16.2% at 2.3-A resolution using synchrotron radiation diffraction data . The tetrameric molecule has 222 symmetry, with one of the axes coincident with the crystallographic 2-fold symmetry axis of the crystal which belongs to the space group P2(1)2(1)2 with a = 76.0 A, b = 138.3 A, and c = 93.5 A . Each subunit comprises 14 alpha-helices and 16 beta-strands, which fold into a small and a large domain . The coenzyme-binding lysine residue is located at the interface between the large and small domains of one subunit and the large domain of a crystallographically related subunit . The fold of the large, pyridoxal 5'-phosphate binding domain and the location of the active site are similar to that found in aminotransferases . Most of the residues which participate in binding of pyridoxal 5'-phosphate in aminotransferases are conserved in the structure of tyrosine phenol-lyase . Two dimers of tyrosine phenol-lyase, each of which has a domain architecture similar to that found in aspartate aminotransferases, are bound together through a hydrophobic cluster in the center of the molecule and intertwined N-terminal arms.

J Gen Microbiol, 1993 Apr, 139 ( Pt 4), 881 - 7
Immunological relatedness of the LamB proteins among members of Enterobacteriaceae; Werts C et al.; We have studied the immunological relatedness of LamB proteins from a wide range of enterobacterial species, using antibodies directed against denatured Escherichia coli K12 and Klebsiella pneumoniae LamB proteins (LamBE.c . and LamBK.p., respectively), and anti-peptide antibodies directed against 10 distinct loops of LamB from E . coli K12 predicted to protrude either side of the outer membrane . We have shown that a protein immunologically related to LamBE.c . and LamBK.p . was present in all members of Enterobacteriaceae tested . A protein recognized by several anti-peptide antibodies was identified in E . coli, Shigella sonnei, Salmonella typhimurium and Kleb . pneumoniae, as well as in two Citrobacter species, two Enterobacter species and Kluyvera ascorbata . The recognition patterns obtained with the anti-peptide antibodies were in agreement with the LamB protein sequence data available . They indicated that the cell surface and also the periplasmic loops of LamB are subject to great antigenic variability.

Eur J Biochem, 1993 Apr 1, 213(1), 405 - 12
Overproduction, solubilization, purification and DNA-binding properties of AmpR from Citrobacter freundii; Bishop RE et al.; AmpR belongs to the LysR family of prokaryotic DNA-binding transcriptional regulators and controls induction of the enterobacterial ampC beta-lactamase gene . The ampR gene of Citrobacter freundii was deregulated by employing the polymerase chain reaction to introduce an efficient ribosome-binding sequence and suitable restriction enzyme sites for cloning into a chemically inducible tac-promoter expression vector . When induced in Escherichia coli, the modified ampR gene rapidly overproduced the AmpR protein as an insoluble aggregate . The AmpR protein could be solubilized with 1.32 M guanidine/HCl and remained soluble when dialyzed against 0.5 M NaCl . The solubility properties of AmpR were exploited to selectively precipitate and resolubilize the protein in a nearly homogenous state . AmpR was then purified by a single gel-filtration chromatography step which demonstrated that AmpR exists in solution as a monodisperse homodimeric protein . Several milligrams of purified AmpR could be obtained routinely from a 1-1 culture of induced bacteria . A DNA-binding assay buffer containing 300 mM potassium glutamate and 30% glycerol was found to stabilize AmpR and used to demonstrate sequence-specific DNA-binding . Additionally, purified AmpR binds a half-operator DNA with an inverted-repeat sequence which competes with binding by the wild-type operator . These findings are discussed in terms of the helix-turn-helix DNA-binding motif, whereby AmpR is proposed to interact with its wild-type operator as a dimer of dimers.

FEMS Microbiol Lett, 1993 Mar 15, 108(1), 81 - 5
Purification and characterization of three carboxylesterases from Enterobacteriaceae; Goullet P et al.; The carboxylesterases from Proteus vulgaris, Salmonella enterica and Citrobacter amalonaticus were purified 104-, 95- and 120-fold, respectively by chromatography . The enzymes had similar catalytic activities but differed considerably in their inactivation by heat, di-isopropyl fluorophosphate and Cd2+, Zn2+, Hg2+ and Cu2+ . Quantitative neutralization of hydrolytic activity with specific immunoglobulins indicated that the three enzymes were antigenically distinct.

J Antimicrob Chemother, 1993 Mar, 31(3), 345 - 62
The activity of cefpirome and ten other antibacterial agents against 2858 clinical isolates collected from 20 centres; Reeves DS et al.; Two thousand eight hundred and fifty-eight aerobic clinical isolates of Enterobacteriaceae, Pseudomonas species, staphylococci, streptococci and Haemophilus species were collected in 19 geographically separated centres in the UK and one in Ireland . The identity of each isolate was confirmed at Southmead Hospital and the MIC of cefpirome, cefotaxime, ceftazidime, cefuroxime, cephradine, amoxycillin, piperacillin, imipenem, gentamicin, ciprofloxacin and trimethoprim was determined by an agar dilution method . Against species of Enterobacteriaceae not associated with producing an inducible cephalosporinase, cefpirome had a similar degree of activity to cefotaxime and was more active than ceftazidime and earlier cephalosporins . Against species with a high prevalence of inducible beta-lactamase production, cefpirome was superior to other cephalosporins; imipenem was also active against these isolates . Cefpirome was active against Pseudomonas aeruginosa, methicillin-sensitive staphylococci, non-enterococcal streptococci and Haemophilus spp . When the isolates were exposed to 0.1 mg/L of imipenem in agar plus the test agent, cefpirome had superior activity compared with the other cephalosporins tested for the Enterobacteriaceae, except Proteus vulgaris and Proteus penneri . 8.7% of Enterobacter spp., 7% of Citrobacter spp . and 6.7% of Morganella morganii had susceptibilities to beta-lactams suggesting constitutive hyperproduction of chromosomal cephalosporinase; cefpirome, unlike the other cephalosporins tested, was active against these isolates, although to a lesser degree than against the wild-type inducible isolates . No isolates were thought to produce an extended-spectrum beta-lactamase.

J Infect, 1993 Mar, 26(2), 207 - 9
Citrobacter diversus ventriculitis and brain abscesses in an adult; Booth LV et al.; A case of Citrobacter diversus brain abscesses following urinary infection in an adult is described . The patient was treated with surgical drainage, netilmicin and cefotaxime . Citrobacter species CNS infection is discussed.

J Bacteriol, 1993 Mar, 175(5), 1508 - 13
Analysis of 7-substituted sialic acid in some enterobacterial lipopolysaccharides; Gamian A et al.; Sialic acid-containing lipopolysaccharides (LPS) were isolated from six bacterial strains of the family Enterobacteriaceae . Sialic acid was released from permethylated LPS by methanolysis, and partially O-methylated N-acetyl-N-methyl-neuraminic acid methyl ester methyl glycosides were analyzed by gas-liquid chromatography-electron ionization mass spectrometry . It was proved that all LPS contain N-acetylneuraminic acid (NeuAc) . The occurrence of 7-substituted NeuAc in Escherichia coli serotypes O24 and O56 and in Citrobacter freundii O37 LPS was documented . The LPS preparations also contained terminal NeuAc . LPS of E . coli O104 had exclusively 4-substituted sialic acid . The remaining LPS studied, namely, from Salmonella toucra O48 and Hafnia alvei 2, had 4-linked and terminally localized NeuAc residues.

J Clin Microbiol, 1993 Mar, 31(3), 760 - 1
Isolation of a Citrobacter freundii strain which carries the Escherichia coli O157 antigen; Bettelheim KA et al.; A biochemically typical strain of Citrobacter freundii which carries the Escherichia coli O157 antigen is described . The significance of differentiating such strains from typical E . coli O157 strains is stressed.

J Biol Chem, 1993 Feb 15, 268(5), 3690 - 7
Role of Ser-238 and Lys-240 in the hydrolysis of third-generation cephalosporins by SHV-type beta-lactamases probed by site-directed mutagenesis and three-dimensional modeling; Huletsky A et al.; A growing number of extended spectrum SHV-type beta-lactamases capable of hydrolyzing third-generation cephalosporins such as cefotaxime and ceftazidime have been reported . These new enzymes differ by a few amino acids from SHV-1, an enzyme incapable of hydrolyzing these drugs . Two of these substitutions, Gly-238-->Ser and Glu-240-->Lys, are in a key beta-strand of the catalytic site of class A beta-lactamases . To understand the structural basis of these new activities, we first subcloned the DNA region coding for SHV-1 and SHV-2 and did site-directed mutagenesis to create two mutant SHV-1 proteins containing Ser and Glu or Gly and Lys and two mutant SHV-2 proteins containing Gly and Glu or Ser and Lys in positions 238 and 240, respectively . Phenotypic analysis (antibiograms and minimum inhibitory concentrations) and activity spectra of mutant enzymes showed that Ser-238 is critical for cefotaxime hydrolysis whereas both Ser-238 and Lys-240 are needed for strong ceftazidime hydrolysis . A three-dimensional model for SHV beta-lactamase complexes was constructed using the crystallographic structure of the homologous Bacillus licheniformis beta-lactamase, the complex of cefotaxime with the Streptomyces sp . R61 D-alanyl-D-alanine peptidase, and the complex of aztreonam with the Citrobacter freundii beta-lactamase . The modeling of SHV beta-lactamase complexes showed that factors which are most likely to correlate with binding and kinetic data are the size of the relatively buried amino acid at position 238 and the electrostatic charge of the exposed group at position 240.

Int J Food Microbiol, 1993 Feb, 17(4), 281 - 8
Detection of Salmonella in animal protein by Rappaport-Vassiliadis broth using indirect impediometry; Donaghy JA et al.; Indirect impedance methodology for the detection of Salmonella was investigated using a rapid automated bacterial impedance technique (RABIT) system . Four commercially available Rappaport-Vassiliadis (RV) enrichment broths were evaluated for their sensitivity and selectivity in detecting Salmonella using this technique . The RV from Lab M and Oxoid (new) gave the shortest detection times and showed good correlation between Salmonella numbers and detection times . Using Lab M medium, the indirect impedance technique could distinguish between Salmonella spp . and the closely related genera, Proteus and Citrobacter . The impedance technique showed recoveries of Salmonella from processed animal protein and raw meats equivalent to, or better than, those obtained with RV used in a conventional Salmonella isolation procedure.

Pediatr Res, 1993 Feb, 33(2), 205 - 8
Interspecific plasmid transfer and modification of heat-stable enterotoxin expression by Klebsiella pneumoniae from infants with diarrhea; Alessio M et al.; We have previously described two heat-stable enterotoxins (ST) produced by Citrobacter freundii and by Klebsiella pneumoniae . To see whether transfer of toxigenic ability may occur between different bacterial species, conjugal mating experiments were performed using strains isolated from children . Donors and recipients were incubated together . ST production by the recipients was tested by the suckling mouse assay, which is the standard biologic method for detecting ST, and also by an ELISA test . The latter is based on MAb directed to Escherichia coli 18- or 19-amino acid toxin . Citrobacter and E . coli strains producing an ELISA-positive ST were used as plasmid donors . Recipients were E . coli, Klebsiella P89, and Klebsiella AL55 . The latter had been previously shown to be capable of producing an ELISA-negative ST, but had spontaneously lost this ability . After conjugation, all strains gave positive results in the suckling mouse assay . However, E . coli and Klebsiella P89 ST were positive in the ELISA test, whereas Klebsiella AL55 ST was negative . The same three strains were transformed by inserting the plasmid pSLM004 encoding an ELISA-positive ST . Klebsiella P89 and E . coli became capable of producing an ELISA-positive ST, whereas Klebsiella AL55 cells were positive in the suckling mouse assay but negative in the ELISA . To assess whether the toxigenicity acquired was itself transmissible, the new ST-producing strains were incubated with a nontoxigenic Citrobacter strain . The latter became capable of producing an ELISA-positive ST in all cases.(ABSTRACT TRUNCATED AT 250 WORDS)

Infect Immun, 1993 Feb, 61(2), 534 - 43
Shiga-like toxin II-related cytotoxins in Citrobacter freundii strains from humans and beef samples; Schmidt H et al.; By hybridizing colonies grown from 928 individual stool samples of patients suffering from diarrhea with oligonucleotide probes 772 and 849 complementary to Shiga-like toxin I (SLT-I) and SLT-II gene sequences, respectively, we identified two strains that hybridized with probe 849, which biochemical identification revealed as Citrobacter freundii . An additional five slt-II probe-positive isolates were screened from 81 beef samples . Polymerase chain reaction analysis and restriction of amplified products provided evidence for slt-II-related genes in all seven strains . From C . freundii LM 76, the genes encoding the A and B subunits were cloned in pUC 18 vectors and sequenced . The gene encoding the A subunit differed from that of Escherichia coli slt-IIvhc in 4 bases, resulting in two amino acid residue differences . In 11, 13, and 11 nucleotides, differentiation of slt-IIA, slt-IIcA, and vtx2haA, respectively, was found . These differences affected the predicted amino acid sequence as follows: there were six amino acid differences with SLT-IIA, five with SLT-IIcA, and four with VTx2haA . The nucleotide sequence of the gene encoding the B subunit is identical to slt-IIvhcB and differed from slt-IIcB and vtx2haB by only a single nucleotide base, but this resulted in a predicted amino acid sequence identical to that reported for these toxins . We therefore termed the toxin genes C . freundii slt-IIcA and slt-IIcB . Culture filtrates inoculated with material from the colonies from primary cultures were cytotoxic to Vero cells . Neutralization assays with antisera to E . coli SLT-I, SLT-II, and SLT-IIvhc revealed that antibodies against SLT-IIvhc reduced the C . freundii cytotoxic activity specifically and to the same degree as with the E . coli SLT-IIvhc control strain . In five of the seven strains tested, subcultivation on both a liquid or solid medium resulted in loss of cytotoxic activity . With polymerase chain reaction, we demonstrated that loss of cytotoxic activity ran parallel with the loss of slt genes . These data demonstrate the intergeneric occurrence of SLT-II-related toxins, which may well be a new marker of enteropathogenicity in C . freundii . Our findings that the toxin genes belong to the slt-II family plus their evident instability in the majority of strains should help pave the way to a better understanding of their role in diarrhea or food poisoning.

Antimicrob Agents Chemother, 1993 Feb, 37(2), 224 - 8
Sequences of wild-type and mutant ampD genes of Citrobacter freundii and Enterobacter cloacae; Kopp U et al.; The ampD gene product regulates the expression of AmpC beta-lactamase in gram-negative bacteria and is proposed to be involved in peptidoglycan metabolism . In this study, we sequenced the ampD wild type and three mutant genes of Enterobacter cloacae and Citrobacter freundii . They exhibited a high degree of homology with the corresponding gene of Escherichia coli except in the carboxy termini, where, in the wild-type genes of E . cloacae and C . freundii, four additional amino acids yielding the Ser-X-X-Lys motif were found . Evidence that this C-terminal region of the ampD gene product is necessary for activity was shown by constructing a deletion of the last 16 amino acids . The spontaneous mutation of ampD02 is an out-of-frame insertion and yields an inactive AmpD protein . The single-base-pair substitution of Gly for Asp-121 in ampD05 is responsible for a hyperinducible phenotype . These results demonstrate regions of the ampD gene and the corresponding protein which have functional importance for the induction of AmpC beta-lactamase in E . cloacae.

Enzyme Microb Technol, 1993 Feb, 15(2), 109 - 13
Assessment of macroporous polystyrene-based polymers for the immobilization of Citrobacter freundii; Griffiths MS et al.; POLYHIPE polymers represent a novel immobilization support possessing many desirable characteristics . These materials have been used to immobilize Citrobacter freundii in order to develop a continuous bioconversion system for the anaerobic production of trimethylene glycol from glycerol . Several immobilization methods were employed, of which the most successful involved culturing the organism with POLYHIPE in shake-flasks, transferring the contents into a column, and growing the cells in situ within the column . Cell activity was determined by the amount of TMG produced from a buffered glycerol solution . The latter was passed through the columns by either a recirculation system or the superior single-pass method . The optimal system was scaled up and showed potential for further large-scale investigations . Cell lifetime in the column could be improved by supplying the cells with diluted growth medium in place of the buffer.

J Antibiot (Tokyo), 1993 Jan, 46(1), 71 - 87
In vitro antibacterial activity of FK037, a novel parenteral broad-spectrum cephalosporin; Mine Y et al.; FK037 is a new parenteral cephalosporin, which offers some advantages over the commercially available parenteral cephalosporins . It demonstrated potent broad-spectrum activity against clinical isolates of Gram-positive bacteria including methicillin-resistant staphylococci, and Gram-negative bacteria including Pseudomonas aeruginosa . Against clinical isolates of aerobic Gram-positive bacteria, FK037, like cefpirome, demonstrated more potent activity than ceftazidime, cefoperazone and ceftizoxime . It is noteworthy that FK037, on the basis of the MIC90s, was the most active of all the cephalosporins tested against methicillin-resistant Staphylococcus aureus (MRSA) . It was similar in activity to cefpirome against methicillin-sensitive S . aureus (MSSA) . Against clinical isolates of aerobic Gram-negative bacteria, FK037, like cefpirome, was superior to cefoperazone, similar to ceftazidime and inferior to ceftizoxime in activity . Against P . aeruginosa, FK037 was superior to cefoperazone, similar or slightly superior to cefpirome and inferior to ceftazidime in activity . However, FK037 exhibited significant activity against Citrobacter and Enterobacter which were highly resistant to ceftazidime, cefoperazone and ceftizoxime . FK037 had an advantage in that its bactericidal activity against S . aureus, Escherichia coli and P . aeruginosa at sub-MICs (1/2 or 1/4 the MIC) was much stronger than those of cefpirome and ceftazidime . Moreover, it exhibited potent bactericidal activity against MSSA, MRSA and P . aeruginosa in a pharmacokinetic in vitro model simulating human plasma concentrations after intravenous dosage of 0.125, 1.0 and 1.0 g, respectively . FK037 inhibited essential penicillin-binding proteins (PBPs), 1, 2 and 3 of S . aureus with a 50% inhibitory concentration (I50) of 0.58 micrograms/ml or lower . Of essential PBPs 3, 1a and 1b of E . coli and P . aeruginosa, FK037 inhibited PBP 3 at the lowest I50 (0.03 and 0.04 micrograms/ml, respectively) and PBPs 1a and 1b with I50 values of 2.7 micrograms/ml or lower . FK037, like cefpirome, was highly stable to hydrolysis by various beta-lactamases except Ic cephalosporinase from Bacteroides fragilis, and had extremely low affinity for beta-lactamases . Therefore, FK037 was more potent than ceftazidime in activity against beta-lactamase-producing bacteria except P . aeruginosa and Serratia marcescens . The ability of FK037 to penetrate the outer membrane of E . coli was slightly higher than that of ceftazidime, but slightly lower than that of cefpirome.

Mikrobiyol Bul, 1993 Jan, 27(1), 62 - 70
{Nose and throat carriage in "food handlers"}; Hacibektasoglu A et al.; From December 1991 to February 1992, 450 personnel have been investigated for pathogen microorganisms in their nose and throat . The study was performed in the Infectious Diseases Section of Gulhane Military Medical Academy . Pathogen microorganisms have been isolated from 54 nose culture (12%) and 6 throat culture (1.33%) . In one person pathogen microorganisms have been isolated from his nose and throat . The difference between the two groups (The nose and throat cultures) was significant at p = 0.001 by Fisher's exact test (t = 6.414) . In the nose cultures the pathogen microorganisms were Staphylococcus aureus (85.2%), Proteus vulgaris (5.6%), Proteus mirabilis (3.7%), Klebsiella pneumoniae (1.8%), Citrobacter freundii (1.8%), Group C beta-hemolytic streptococcus (1.8%) and Pseudomonas aeruginosa (1.8%) . Only one (1.85%) had two different pathogen microorganisms (Staphylococcus aureus and Proteus vulgaris) in his nasal culture . In the throat cultures the pathogen microorganisms were Staphylococcus aureus (66.7%) and group A beta-hemolytic Streptococcus (50%) . Only one (16.6%) had two different pathogen microorganisms (S.aureus and group A beta-hemolytic Streptococcus) in his throat culture . The same pathogen microorganisms (S.aureus) has been isolated in an only one person's nasal and throat cultures (0.22%) . We treated 60 personnel who were nasal and throat carriers according to the results of antibiograms . After treatment, two still had previous pathogen microorganism (Staphylococcus aureus) . These two carriers were eradicated by repeating the treatment.

J Pediatr, 1993 Jan, 122(1), 15 - 21
Gram-negative enteric bacillary meningitis: a twenty-one-year experience; Unhanand M et al.; We reviewed our experience with gram-negative enteric bacillary meningitis in neonates and infants from 1969 through 1989 . Ninety-eight patients were identified . Their ages were from 1 day to 2 years with a median of 10 days . In 25 patients (26%), predisposing factors were identified, the most common of which were neural tube defects and urinary tract anomalies . The causative agents were Escherichia coli (53%), Klebsiella-Enterobacter species (16%), Citrobacter diversus (9%), Salmonella species (9%), Proteus mirabilis (4%), Serratia marcescens (3%), Bacteroides fragilis (3%), and Aeromonas species (2%) . At the time of diagnosis, Gram-stained smears of cerebrospinal fluid revealed gram-negative bacilli in 61% of patients . The causative organism was cultured from blood obtained from 55% of patients, and 21% had positive urine culture results . The cerebrospinal fluid leukocyte counts ranged from 0 to 80,600 cells/mm3, and the cerebrospinal fluid/serum glucose concentration ratio was less than 0.5 in 72% of patients . Antimicrobial regimens varied greatly . After initiation of antibiotic therapy, an average of 3 days was needed for eradication of bacteria from cerebrospinal fluid . The case-fatality rate was 17%, and 61% of survivors had long-term sequelae that included seizure disorders, hydrocephalus, physical disability, developmental delay, and hearing loss.

Microbios, 1993, 74(300), 147 - 54
Comparative antimicrobial activity of lomefloxacin, norfloxacin, ofloxacin, ciprofloxacin and enoxacin against > 500 bacterial isolates; Ismaeel NA et al.; An agar dilution technique was used to compare the antimicrobial activities of lomefloxacin, norfloxacin, ofloxacin, ciprofloxacin and enoxacin against 544 strains of bacterial isolates . Among the five quinolone agents tested, ciprofloxacin was the most active . Enoxacin was the most active after ciprofloxacin against Escherichia coli, Enterobacter aerogenes, Proteus mirabilis, Shigella spp., Yersinia enterocolitica, and Haemophilus influenzae with an MIC90 of < or = 0.25 micrograms/ml . Ofloxacin was the most active agent after ciprofloxacin against Klebsiella pneumoniae, Enterobacter cloacae, Citrobacter diversus, and Legionella pneumophila with an MIC of < or = 0.25 micrograms/ml . Ciprofloxacin inhibited Staphylococcus spp . and Streptococcus spp., at < or = 0.5 micrograms/ml and 2 micrograms/ml, respectively . Norfloxacin and enoxacin had the same antimicrobial activity (MIC90) against Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pyogenes, Streptococcus agalactiae and some other Gram-positive species, but these activities were weak when compared with ciprofloxacin . The results of this in vitro study show that ciprofloxacin is very active against Gram-negative and Gram-positive species.

Scand J Infect Dis Suppl, 1993, 91, 25 - 32
Cross-susceptibility of cefpirome and four other beta-lactams against isolates from haematology/oncology and intensive care units . International Study Group; Spencer RC; A multi-centre in-vitro study (8625 isolates) was conducted in 13 countries between May and November, 1992 to determine both the current bacterial epidemiology in intensive care and haematology/oncology units and the cross-susceptibility of the organisms to cefpirome, ceftazidime, ceftriaxone, imipenem and piperacillin . Bacterial species with 20 or more isolates resistant to one of the six antibiotics were examined for their susceptibility to the beta-lactams . Cefpirome and imipenem had the smallest total numbers of isolates . Bacteria resistant to ceftazidime or ceftriaxone were often susceptible (> 50%) to cefpirome . Conversely, cefpirome resistant isolates were frequently resistant (> 90%) to ceftazidime and ceftriaxone . P . aeruginosa was an exception, exhibiting cross-resistance to all cephalosporins . beta-lactamase producing Enterobacter, Citrobacter and Klebsiella spp . were especially resistant to piperacillin and ceftazidime but not cefpirome or imipenem . Two-thirds or more of coagulase-negative staphylococci resistant to any single agent, including imipenem, maintained their susceptibility to cefpirome . Cross-class resistance was not exhibited by imipenem and cefpirome against ciprofloxacin resistant isolates but was more evident for piperacillin, ceftazidime and ceftriaxone . Cefpirome was more active than ceftazidime against bacteria resistant to both piperacillin and gentamicin, especially coagulase-negative staphylococci (76% vs . 6%) and Enterobacter spp . (56% vs . 21%) . Many coagulase-negative staphylococci and Enterobacter spp . susceptible to cefpirome (50-89%) . These results suggest that cefpirome has a potential clinical advantage against gram-positive and gram-negative bacteria resistant to other beta-lactams, including imipenem.

Med Dosw Mikrobiol, 1993, 45(1), 115 - 8
{Susceptibility in vitro to certain quinolines of gram-negative bacteria and gram-positive cocci causing urinary tract infections}; Mroz E et al.; Susceptibility to norfloxacin, ofloxacin, pipemidic acid and nalidixic acid of 848 bacterial strains isolated from urine of patients treated in 1989-1992 in Wroclaw hospitals was investigated . The study, performed by the disc diffusion methods, concerned 568 Enterobacteriaceae strains, 147 Gram-negative non-fermenting bacteria and 133 strains of staphylococci . Highest percentage (90-100%) of susceptibility to all used antimicrobial agents was detected among Escherichia, Proteus, Morganella and Citrobacter . Less frequent susceptibility (30-70%) was observed among Klebsiella, Enterobacter and Serratia . Among strains of P . aeruginosa susceptible to norfloxacin and ofloxacin were, respectively, 61.4 and 22.2% isolates . Over 95% of strains of P . aeruginosa were resistant to nalidixic acid . Among other non-fermenting Gram-negative bacteria, over 50% were resistant to norfloxacin and ofloxacin . Both S . aureus and S . epidermidis were susceptible to ofloxacin and norfloxacin in 81-93% of tested strains . They were 2-3 times less frequently susceptible to pipemidic and nalidixic acid.

FEMS Microbiol Lett, 1992 Dec 15, 79(1-3), 281 - 5
Growth temperature-dependent activity of glycerol dehydratase in Escherichia coli expressing the Citrobacter freundii dha regulon; Daniel R et al.; Using the cosmid pWE15, a genomic library of Citrobacter freundii DNA in Escherichia coli ECL707 was prepared and screened for glycerol utilization . Six out of approximately 3000 clones were positive . One clone, harboring the recombinant cosmid pRD1, expressed glycerol dehydratase in high activity when grown at 28 degrees C but not at 37 degrees C . The growth temperature had little effect on the activity of the other enzymes encoded by the dha regulon . When the glycerol-containing medium was supplemented with corrinoids, the recombinant E . coli strain produced 1,3-propanediol in high amounts at 28 degrees C.

Biosci Biotechnol Biochem, 1992 Dec, 56(12), 1916 - 20
Characterization of membrane-bound spermidine dehydrogenase of Citrobacter freundii; Hisano T et al.; Spermidine dehydrogenase found in the membrane fraction of Citrobacter freundii IFO 12681 was solubilized with Triton X-100 and further purified to homogeneity . The properties of the membrane enzyme were almost identical to those obtained from the soluble fraction of the organism with respect to molecular and catalytic properties . Thus, binding properties of the enzyme to the bacterial membrane were checked . The ratio of enzyme activity found in the soluble fraction to the membrane fraction was dependent on salt concentration during cell disruption . A hydrophobic interaction was largely involved in anchoring the enzyme to the membrane fraction . Purified spermidine dehydrogenase from the soluble fraction was readily adsorbed into the membrane fraction in the presence of salt . Spermidine dehydrogenase appeared to be a membrane-bound enzyme localized in the cytoplasmic membranes in a manner that makes a partial release of the enzyme possible during mechanical cell disruption . When spermidine oxidation was done with the resting cells of C . freundii, a stoichiometric formation of two reaction products, 1,3-diaminopropane and gamma-aminobutyraldeyde, was observed without any lag time . These facts indicate that the enzyme is localized on the outer surface of the cytoplasmic membranes or in the periplasmic space of the organism.

FEMS Microbiol Lett, 1992 Dec 1, 78(2-3), 199 - 204
Characterization of the Citrobacter freundii phoE gene and development of C . freundii-specific oligonucleotides; Spierings G et al.; The phoE gene of Citrobacter freundii, encoding a pore-forming outer membrane protein, was cloned and its nucleotide sequence was determined . The homologies in terms of identical amino acids between the C . freundii PhoE protein and those of Escherichia coli, E . cloacae and Klebsiella pneumoniae were 90%, 86% and 84%, respectively . Two synthetic oligonucleotides, corresponding to hypervariable, cell surface-exposed regions of the protein, were tested for their specificity in polymerase chain reactions . They were specific for the species C . freundii, i.e., no reaction was detected with 35 non-C . freundii strains tested, including 17 Salmonella, two C . amalonaticus and three C . diversus strains, whereas all five C . freundii strains tested were correctly recognized.

J Clin Microbiol, 1992 Nov, 30(11), 2921 - 9
Analysis of relationships among isolates of Citrobacter diversus by using DNA fingerprints generated by repetitive sequence-based primers in the polymerase chain reaction; Woods CR Jr et al.; Oligonucleotide probes which match consensus sequences of the repetitive extragenic palindromic (REP) element hybridize to genomic DNA of diverse bacterial species . Primers based on the REP sequence generate complex band patterns with genomic DNA in the polymerase chain reaction (PCR), a technique named REP-PCR . We used REP-PCR with genomic DNA to fingerprint 47 isolates of Citrobacter diversus . Previously, 37 were assigned electrophoretic types (ETs) by multilocus enzyme electrophoresis and 35 were evaluated by using outer membrane protein profiles . Fingerprints were compared by visual inspection and by similarity coefficients (SimCs) based on the number of common bands versus total bands between two given isolates . DNA fingerprints were highly similar visually for patient pairs and outbreak-related sets . SimCs for these were > or = 0.952 . Fingerprints of isolates with different ETs generally were distinctive . Among 21 unrelated isolates representing 15 ETs, only 6 of 210 comparisons had SimCs of > or = 0.952 . REP-PCR rapidly generated DNA fingerprints which were highly similar for epidemiologically linked isolates of C . diversus and distinct for previously characterized strains within this species . The ability of this method to discriminate between C . diversus isolates with the same biotype was similar to that of multilocus enzyme electrophoresis and outer membrane protein profiles . REP-PCR may be useful in evaluation of apparent outbreaks of this or other bacterial species which possess these extragenic, repetitive elements.

J Infect Dis, 1992 Nov, 166(5), 1035 - 44
Interaction of Citrobacter diversus strains with HEp-2 epithelial and human umbilical vein endothelial cells; Woods CR Jr et al.; More than 75% of neonates with Citrobacter diversus meningitis develop brain abscesses . Interaction of C . diversus strains with HEp-2 and human umbilical vein endothelial cells (HUVEC) was studied to examine mechanisms related to brain abscess formation . Two of 9 strains invaded HEp-2 cells and 0 of 6 invaded HUVEC better than the others . C . diversus survived at least 20 h within HEp-2 cells (in decreasing numbers) . Adhesion to HEp-2 cells was increased in 3 of 4 strains expressing type 1 fimbriae, but this did not correlate with increased invasion . Inhibition of RNA or protein synthesis blocked invasion but not adhesion . Thus, invasion requires ongoing protein synthesis, and adhesion to and invasion of HEp-2 cells by type-1-fimbriated strains are independent steps . Invasion was inhibited by cytochalasin D . A 32-kDa protein found in cerebrospinal fluid isolates of C . diversus was not related to invasion of either cell line . Ability to invade HEp-2 cells was not increased among strains isolated from central nervous system sources.

J Antibiot (Tokyo), 1992 Nov, 45(11), 1738 - 45
Antifungal anthracycline antibiotics, spartanamicins A and B from Micromonospora spp; Nair MG et al.; Spartanamicins A and B, two antifungal antibiotics, were produced by a culture of Micromonospora spp . strain No . MSU-43097 (ATCC 53803), isolated from a potted soil containing asparagus (Asparagus officinalis L.) plants . The antibiotics were isolated from the mycelial cake using organic solvents . The structures of spartanamicins A and B were determined by spectral and chemical means . Spartanamicin B is more active as an antifungal compound than it's analogue, A . The minimum inhibitory concentration for spartanamicin B on Candida albicans and Aspergillus, Cladosporium, Cryptococcus, Rhodotorula and Staphylococcus spp . ranged from 0.2 to 1 microgram/ml . It was not active against Staphylococcus aureus, Escherichia coli and Citrobacter spp . but some strains of S . aureus were sensitive.

Equine Vet J, 1992 Nov, 24(6), 457 - 61
Evaluation of progesterone treatment to create a model for equine endometritis; Hinrichs K et al.; To investigate a model for equine endometritis, 12 mares with normal reproductive tracts were divided into 2 groups . All mares received progesterone in oil, 250 mg im, daily . At 5 days after initiation of progesterone administration, the uteri were inoculated with 10(6) colony forming units of Pseudomonas aeruginosa . The day of inoculation was designated Day 0 . On Day 6, endometrial swab samples yielded P . aeruginosa in 5 mares; samples from the other 7 mares yielded heavy growth of Escherichia coli, Streptococcus zooepidemicus, Klebsiella pneumoniae, Enterobacter spp., Citrobacter diversus, Staphylococcus epidermidis and Streptococcus morbillorum . On Days 6, 7 and 8, Group A mares received intrauterine infusions of 6 g ticarcillin disodium and 0.2 g clavulanate potassium in 100 ml sterile saline . Group B mares received infusions of saline only . The incidence of swab specimens yielding no bacterial growth was significantly higher in Group A than Group B mares on Days 8 and 13 (4/6 vs 0/6) . Swab samples from 5 of the 6 mares in Group A yielded growth of fungi on Days 13 and 19 . Mares in Group B were then similarly treated with ticarcillin/clavulanate infusions, on Days 19, 20 and 21 . The incidence of swab specimens yielding no bacterial growth was 2/6 and 1/6 on Days 21 and 26, respectively; fungi were not recovered from these mares at any time . The incidence of no-growth swabs after antibiotic treatment tended to be higher in Group A and incidence of fungal recovery after antibiotic treatment was significantly higher in Group A.(ABSTRACT TRUNCATED AT 250 WORDS)

J Clin Microbiol, 1992 Nov, 30(11), 2903 - 10
Evaluation of the autoSCAN-W/A rapid system for identification and susceptibility testing of gram-negative fermentative bacilli; York MK et al.; The autoSCAN-Walk-Away (W/A) system for identification and susceptibility testing was evaluated for 400 gram-negative fermentative bacteria by using the API 20E (366 isolates) and/or tube biochemical tests as the reference identification system and a frozen microdilution MIC tray system for susceptibility testing . The W/A system performed well for identification of this group of organisms representing 14 genera and 30 species, showing a sensitivity of 96% and results available in 2 h . Of the 16 misidentifications, 6 were with Serratia liquefaciens . A total of 63 isolates (17%) required further tests to complete the identification, compared with 106 (29%) of the isolates which required additional tests for the API 20E identification . Approximately half (32) of the additional tests with the W/A system were required in order to separate Citrobacter diversus from C . amalonaticus . For susceptibility determinations, the W/A system demonstrated an overall agreement of 93% (4,102 determinations) with 40 major errors (0.98%) . However, of the 906 resistant organism-drug combinations in the study, there were 115 very major errors, for a false-susceptibility rate of 12.7% of the resistance determinations . Among these very major errors, 80% occurred with piperacillin and the cephalosporins . The W/A system completed the MIC determinations in 7 h; however, the difficulty in detecting resistance with some antimicrobial agents limited the advantages of the rapid susceptibility testing.

J Antimicrob Chemother, 1992 Oct, 30(4), 429 - 47
Survey of the prevalence of beta-lactamases amongst 1000 gram-negative bacilli isolated consecutively at the Royal London Hospital; Liu PY et al.; beta-Lactamase expression was examined in 1000 consecutive Gram-negative bacilli isolated from urine, wound swab, sputum or blood specimens received at the Microbiology Laboratory of the Royal London Hospital . This survey, performed between January and April, 1991, followed a similar study undertaken in early 1982 . The distribution of species was similar in the two surveys, except that the proportion of Pseudomonas aeruginosa isolates had increased from 11% in 1982 to 17.5% in the present study . This increase was balanced by a decreased proportion of enterobacteria . Amongst plasmid-mediated beta-lactamases, TEM-1 (especially), TEM-2, SHV-1 and OXA types continued to predominate in enterobacteria . Their frequency in Escherichia coli was unchanged (46% in 1991 compared with 43% in 1982), but had increased from 5 to 22% amongst Proteus mirabilis isolates . An apparent decrease in their frequency amongst Enterobacter cloacae isolates, from 48% in 1982 to 17% in 1991, probably reflected changes to strain prevalence rather than enzyme prevalence . Plasmid type beta-lactamases were present in fewer than 2% of P . aeruginosa isolates in both surveys . In the present study, chromosomal beta-lactamase derepression (constitutive hyperproduction) was detected in 10/76 isolates of E . cloacae, Enterobacter aerogenes, Citrobacter freundii, Serratia spp . and Morganella morganii, and in 2/170 P . aeruginosa isolates . These proportions were increased, compared with those seen the 1982 survey, though the significance was borderline (P approximately 0.05; chi 2 test) . Extended-spectrum plasmid mediated beta-lactamases, unknown in 1982, were found in 11/70 Klebsiellae pneumoniae isolates in the present study . Ten of these organisms, representing at least five distinct strains, produced TEM-10 enzyme, encoded by a plasmid of c . 90 kb; the remaining organism had an unidentified SHV-derived enzyme.

J Paediatr Child Health, 1992 Oct, 28(5), 402 - 3
Citrobacter freundii septic arthritis; Bruehl CL et al.; Septic arthritis in an 8 month old infant due to Citrobacter freundii was treated successfully with a third generation cephalosporin . Infections due to Citrobacter are uncommon in this age group and are almost unknown as a cause of septic arthritis.

J Bacteriol, 1992 Sep, 174(18), 5910 - 5
Expression of Vi antigen in Escherichia coli K-12: characterization of ViaB from Citrobacter freundii and identity of ViaA with RcsB; Houng HS et al.; The Vi antigen in Salmonella typhi is stably expressed and may act to protect the strain against the defensive system of the host . Citrobacter freundii, not usually a common human pathogen, also expresses the Vi antigen but expresses it unstably, exhibiting a reversible transition between the Vi+ and Vi- states . Two widely separated chromosomal regions, ViaA and ViaB, are needed for Vi synthesis . Escherichia coli K-12 harboring a functional ViaB plasmid can also express Vi antigen, but the cloned ViaB sequence can only be stably maintained and expressed in recA hosts . Vi- derivatives arise either through IS1-like insertional events occurring in ViaB sequences or by chromosomal mutations at the ViaA region . P1vir mapping indicates that the ViaA mutations are located at min 47.75 on the E . coli chromosome . All the spontaneous viaA mutants isolated from E . coli and S . typhi were identified as rcsB mutants by complementation tests using plasmid pJB100 . Introduction of rcsA::Tn10 into E . coli harboring functional ViaB sequences eliminates the expression of Vi antigen . These results indicate that Vi antigen synthesis is regulated by the same regulatory proteins involved in colanic acid synthesis in E . coli.

Jpn J Antibiot, 1992 Sep, 45(9), 1112 - 237
{Comparative studies on activities of antimicrobial agents against causative organisms isolated from urinary tract infections (1988) . III . Secular changes in susceptibility}; Kosakai N et al.; Sensitivities to various antibacterial and antibiotic agents of strains of Escherichia coli, Klebsiella spp., Proteus spp., Citrobacter spp., Enterobacter spp., Serratia marcescens and Pseudomonas aeruginosa isolated from patients with urinary tract infections (UTIs) in 9 hospitals during June to November 1988 were compared with those in the same period of previous year according to a classification, uncomplicated UTIs, complicated UTIs without indwelling catheter, and complicated UTIs with indwelling catheter . No remarkable changes were found in sensitivities of E . coli, Proteus spp., Citrobacter spp . and S . marcescens . The sensitivity of Klebsiella spp . to cephems decreased in complicated UTI without indwelling catheter and increased in complicated UTI with indwelling catheter . The sensitivity of Enterobacter spp . to third generation cephems decreased in complicated UTI with indwelling catheter . Sensitivities of P . aeruginosa to aspoxicillin and cefsulodin increased . The number of resistant strains to new quinolones increased slightly.

Science, 1992 Aug 7, 257(5071), 782 - 4
Uranium bioaccumulation by a Citrobacter sp . as a result of enzymically mediated growth of polycrystalline HUO2PO4; Macaskie LE et al.; A Citrobacter sp . accumulates heavy deposits of metal phosphate, derived from an enzymically liberated phosphate ligand . The cells are not subject to saturation constraints and can accumulate several times their own weight of precipitated metal . This high capacity is attributable to biomineralization; uranyl phosphate accumulates as polycrystalline HUO2PO4 at the cell surface . The precipitated metal is indistinguishable from crystalline HUO2PO4.4H2O grown by chemical methods.

Schweiz Med Wochenschr, 1992 Aug 4, 122(31-32), 1173 - 7
{Spondylitis due to Citrobacter diversus . Case report and review of Citrobacter diversus infections}; Mullner GF et al.; Citrobacter diversus (C.d.) is an unusual human pathogen . Its pathogenic potency is small or nil, and in most cases it is isolated in combination with other organisms . Most isolates are of indeterminate clinical significance . Occasionally C.d . may cause opportunistic infections, especially in subjects with primary or secondary conditions of poor host defense, immunosuppressed patients or neonates . We report a case of C.d . vertebral osteomyelitis following C.d . pneumonia . The osteomyelitis was cured by surgery and chemotherapy . This case is interesting since it involves an otherwise healthy subject with no underlying disease . C.d . was isolated in blood during the episode of pneumonia and in bone four weeks later . No other case of Citrobacter diversus vertebral osteomyelitis has hitherto been described.

FEMS Microbiol Immunol, 1992 Aug, 4(6), 323 - 8
Immunochemical studies on sialic acid-containing lipopolysaccharides from enterobacterial species; Gamian A et al.; Lipopolysaccharides with sialic acid as a component were isolated from six bacterial serovars of the family Enterobacteriaceae . SDS-polyacrylamide gel electrophoresis, immunoblotting and passive haemagglutination demonstrated that epitopes are localized in the O-specific polysaccharide of lipopolysaccharides . The strains were tested for a serological relationship; the only distinct cross-reactivity recorded was between Escherichia coli serovars O24 and O56 . The molecular mimicry phenomenon was found for Citrobacter freundii serovar O37 which shared epitopes with horse as well as human red blood cells.

Eur J Biochem, 1992 Aug 1, 207(3), 1123 - 7
The effect of amino acid substitution at position 219 of Citrobacter freundii cephalosporinase on extension of its substrate spectrum; Tsukamoto K et al.; The cephalosporinase of Citrobacter freundii GN346 is a class-C beta-lactamase comprising 361 amino acids . The substitution of the glutamic acid at position 219 in the enzyme by lysine was previously shown to broaden its substrate specificity to unfavorable substrates such as oxyimino cephalosporins {Tsukamoto, K., Ohno, R . & Sawai, T . (1990) J . Bacteriol . 172, 4348-4351} . To investigate the cause of this phenomenon, Glu219 was changed to glutamine, cysteine or tryptophan . All the resultant enzymes showed higher cefuroxime-hydrolytic activities than the wild type, the order of increasing cefuroxime-hydrolytic activity being as follows: Trp greater than Lys greater than Cys greater than Gln greater than Glu . The rate of hydrolysis of cefuroxime by the Trp219 enzyme was approximately 3 x 10(4) times that of the wild-type enzyme . The order of increasing cefuroxime hydrolysis was approximately proportional to the molecular volume of the amino acid substituted and independent of the ionic character of the amino acids . The cysteine residue at position 219 in the Cys219 enzyme allowed its complete reaction with an SH-blocking reagent, 4-chloromercuriphenylsulfonic acid . The modified enzyme with the bulkier residue showed a 45% higher cefuroxime-hydrolytic activity than the untreated enzyme . These results suggested that extension of the substrate spectrum may be attributed to alteration in the configuration of the enzyme around position 219.

Jpn J Antibiot, 1992 Aug, 45(8), 990 - 1002
{Antibacterial activities of sisomicin against fresh clinical isolates}; Deguchi K et al.; To investigate antibacterial activities of sisomicin (SISO), MICs of SISO as well as other aminoglycosides (AGs) were determined against many clinical isolates which were obtained in 1991 . Results are summarized below: 1 . No SISO-resistant strains were observed among isolates of Escherichia coli, Citrobacter diversus, Klebsiella pneumoniae, Klebsiella oxytoca, Enterobacter aerogenes, Proteus mirabilis and Morganella morganii . 2 . In comparison with the results of our previous study against isolates obtained in 1986, the rate of methicillin-resistant Staphylococcus aureus (MRSA) was higher, and SISO-resistant strains were observed at a high rate among the MRSA . Also, SISO-resistant strains of Serratia marcescens increased . However, the rate of SISO-resistant strains of Pseudomonas aeruginosa decreased, and among Citrobacter freundii, Enterobacter cloacae and Proteus vulgaris, SISO-resistant strains did not increase over the years . 3 . MICs of SISO against Providencia rettgeri and Providencia stuartii were high, suggesting that antibacterial activities of SISO was weak against genus Providencia . 4 . For comparison, according to MICs of ofloxacin and imipenem, new quinolone-resistant strains were observed at a high rate among various organisms, and carbapenem-resistant strains were observed at a high rate among S . marcescens and P . aeruginosa . 5 . SISO is still one of the useful AGs in the 1990's since it maintains its strong antibacterial activities against most clinical isolates obtained in recent years and its potential as a combination drug with beta-lactams is being reported.

Jpn J Antibiot, 1992 Aug, 45(8), 965 - 79
{Antimicrobial activities of major oral antibacterial agents against clinically isolated microbial strains from inpatients}; Deguchi K et al.; Antimicrobial activities were examined for major antibacterial agents against clinically isolated microbial strains which were isolated and identified from materials collected from inpatients with various infections in 1988, 1989 and 1990, and the following conclusions were obtained . 1 . Among strains isolated each year, methicillin-resistant Staphylococcus aureus (MRSA) were found frequently . 2 . CEPs-resistant Escherichia coli strains were observed among strains isolated each year . 3 . Increasing tendencies in resistances of Citrobacter freundii, Enterobacter spp., Serratia marcescens to cephems and new quinolones were observed . 4 . Increasing tendencies in resistances of Proteus vulgaris to ceftazidime (CAZ) and new quinolones appeared to exist . 5 . Among strains isolated each year, resistances of Pseudomonas aeruginosa to CAZ and quinolones were observed in high rates, but also their resistances to imipenem appeared to increase . 6 . Many of recently increasing multiple resistant bacteria seem to have different sites of drug action and/or to have non-hydrolytic modes of resistance.

Biotechnol Appl Biochem, 1992 Aug, 16(1), 77 - 85
Cloning and expression of the Erwinia herbicola tyrosine phenol-lyase gene in Escherichia coli; Iwamori S et al.; The tyrosine phenol-lyase (TPL) gene of Erwinia herbicola was cloned and expressed in Escherichia coli, and the complete nucleotide sequence of the gene determined . The TPL gene comprises 1368 bp, encoding 456 amino acids which have 90% amino acid identity with TPL from Citrobacter freundii . After replacing the 5'-flanking region of the TPL gene with the E . coli lac promoter, TPL protein could be hyperproduced constitutively in E . coli without induction by L-tyrosine.

Antimicrob Agents Chemother, 1992 Aug, 36(8), 1756 - 63
Antibacterial activity of RU44790, a new N-tetrazolyl monocyclic beta-lactam; Chantot JF et al.; RU44790 belongs to a new class of synthetic monocyclic beta-lactam antibiotics which feature a bioisosteric tetrazole moiety instead of the more classical acidic functions at the N-1 position of the beta-lactam ring . Its antibacterial activity was evaluated against some 900 strains and was compared with those of other recent beta-lactam derivatives, especially aztreonam . RU44790 is endowed with potent activity against gram-negative bacteria . At less than or equal to 0.6 micrograms/ml, RU44790 inhibited 90% of all strains of the family Enterobacteriaceae with the exception of Citrobacter spp . (MIC for 90% of strains tested, 1.2 micrograms/ml) . The activity was similar to that of aztreonam against strains that are normally susceptible to expanded-spectrum cephalosporins . On the other hand, the new compound was 10 to 100 times more potent than aztreonam and most of the other antibiotics tested against enterobacteria that produce chromosome-encoded or plasmid-mediated extended-spectrum beta-lactamases . Pseudomonas aeruginosa isolates were equally susceptible to both monobactams . RU44790 was inactive against staphylococci and had only marginal activity against streptococci (MIC for 50% of strains tested, 2.5 micrograms/ml) . RU44790 was highly resistant to hydrolysis by various beta-lactamases, particularly cephalosporinases such as P99 . The latter enzyme was also inhibited by the compound . RU44790 showed a high affinity for penicillin-binding protein 3 of Escherichia coli . The results suggest that RU44790 has good potential in the treatment of infections caused by gram-negative microorganisms.

J Antibiot (Tokyo), 1992 Aug, 45(8), 1335 - 45
Cefclidin (E1040), a novel cephalosporin: lack of selection of beta-lactamase overproducing mutants in an in vitro pharmacokinetic model system; Watanabe NA et al.; The bactericidal activity of cefclidin (E1040), a new cephalosporin, against a clinical strain of Citrobacter freundii was compared with that of ceftazidime in a two-compartment in vitro pharmacokinetic model system designed to simulate plasma concentrations in humans for 12 hours after intravenous administration of a 1 g dose . Both cefclidin and ceftazidime showed rapid bactericidal activity against C . freundii . However, during the simulation of ceftazidime treatment, regrowth was observed after two hours and a subpopulation emerged which was resistant to ceftazidime . Neither regrowth nor the emergence of resistant mutants was observed with cefclidin during the 12-hour simulation . The ceftazidime-resistant mutants constitutively overproduced beta-lactamase at levels which were about 500-fold higher than that of the parent wild-type strain . Against this beta-lactamase overproducing mutant, no bactericidal activity of ceftazidime was observed in the in vitro model system, whereas the bactericidal activity of cefclidin was observed during the 12-hour period . The emergence of Enterobacter cloacae mutants derepressed for beta-lactamase production was also observed with ceftazidime but not cefclidin . The affinity of cefclidin for the beta-lactamase isolated from these mutants was lower than that of ceftazidime, and the kinetic parameters of enzymatic hydrolysis showed that cefclidin was hydrolyzed more slowly at a low concentration (0.2 microM) than was ceftazidime . It is suggested that the high activity of cefclidin against strains derepressed for beta-lactamase plays a major role in the absence of emergence of resistant mutants.

J Chemother, 1992 Aug, 4(4), 203 - 10
Epidemiological survey of genes encoding aminoglycoside phosphotransferases APH (3') I and APH (3') II using DNA probes; Alvarez M et al.; The epidemiological survey of APH (3') I and APH (3') II genes, at a time when the specific antibiotic pressure was very low, was carried out by DNA-DNA hybridization . The sample included 334 aminoglycoside resistant Gram-negative bacteria collected from patients of a General Hospital . Of these, 251 hybridized with the APH (3') I-probe and 19 with the APH (3') II-probe but only 190 strains showed high resistance levels (CIM greater than 64 micrograms/ml) for kanamycin, neomycin and paromomycin . These strains were isolated both from inpatients and outpatients with different infectious diseases . The APH (3') I-gene was dispersed among all the bacterial species and clinical specimens tested but the APH (3') II-gene was not found in Pseudomonas spp, Escherichia coli, Citrobacter freundii and Enterobacter cloacae, nor in infected catheters . Several plasmids of different sizes carrying APH (3') genes were detected among different bacteria . Plasmids along with transposable elements (the probes used in this work were developed from Tn906 and Tn5) and the high consumption of other antibiotics whose resistance is carried by these bacteria might be playing an important role in the maintenance and dispersion of APH (3') genes.

Mol Gen Genet, 1992 Aug, 234(2), 228 - 32
Role of IS1 in the conversion of virulence (Vi) antigen expression in Enterobacteriaceae; Ou JT et al.; When Escherichia coli HB101 harbors pWR127, a plasmid comprising the viaB gene from Citrobacter freundii WR7004 and the ColE1-derived pACKC1, the strain produces the virulence (Vi) antigen . Vi antigen expression is abolished (Vi- phenotype), however, when an IS1 or IS1-like DNA element inserts into the viaB region . To determine the sites of IS1 insertion, pWR127 DNAs extracted from 95 independently isolated Vi- strains were analyzed by digestion with the restriction endonuclease PstI and agarose gel electrophoresis . Ten insertion sites were found distributed non-randomly in an area of about 1.3 kb . Nine Vi+ strains (two Citrobacter, two E . coli, and five Salmonella strains), four of which contain pWR127, were then tested for the presence of IS1 by DNA-DNA hybridization . Of the nine strains, five were stable Vi+ and did not contain IS1 . The other four which generated Vi- strains, contained IS1 . When pRR134, a plasmid that contains IS1 was transferred into a stable Vi+ Salmonella typhimurium strain carrying pWR127 (OU5140), Vi- strains were produced from which pWR127 derivatives carrying IS1 inserts could be isolated . It appears, therefore, that the presence of an IS1 or IS1-like element in a strain is required for conversion of the Vi+ expression state to the Vi- expression state.

Diagn Microbiol Infect Dis, 1992 Jul, 15(5), 459 - 63
The E-Test applied to susceptibility tests of gonococci, multiply-resistant enterococci, and Enterobacteriaceae producing potent beta-lactamases; Sanchez ML et al.; 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.

Jpn J Antibiot, 1992 Jul, 45(7), 774 - 98
{Antimicrobial activities of ceftriaxone against clinically isolated strains}; Deguchi K et al.; Antibiotic activities (MICs) of ceftriaxone (CTRX) against 1,210 strains of bacteria including 28 spp . isolated in 1987 and 1990 were compared with those of other cephems . 1 . When compared to data on clinically isolated strains reported in the early 1980s, strains of the following species isolated in 1990 showed extremely elevated MIC90s of CTRX: Staphylococcus spp., Streptococcus pneumoniae, Escherichia coli, Citrobacter spp., Enterobacter spp., Serratia spp., Proteus vulgaris, Morganella morganii and Providencia spp . No changes were observed in MIC90s between the 2 periods for microorganisms such as Streptococcus pyogenes, Haemophilus influenzae, Klebsiella pneumoniae, Proteus mirabilis and Peptostreptococcus spp . 2 . The MIC90 of CTRX to S . pneumoniae was high because a large number of benzylpenicillin (PCG)-insensitive S . pneumoniae (PISP) was present among this species . The MIC80 to Bacteroides fragilis group was also high because highly resistant B . fragilis and B . thetaiotaomicron were isolated in large proportions among the bacteria of this group . Other oxime-type cephems also had high MICs against the above mentioned bacteria . Therefore, a further evaluation has to be made with regard to activities of oxime-type cephems such as CTRX against PISP and B . fragilis group . 3 . Sample strains included, in high ratios, methicillin-resistant Staphylococcus aureus (MRSA), cephamycin-resistant as well as oxime-type cephem-resistant intestinal bacteria, Gram-negative bacteria, and new-quinolone-resistant bacteria . Some of there resistant bacteria are also CTRX-resistant, and CTRX had insufficient activities against them . 4 . With regard to the assessment of changes of frequencies of specific drug-resistant bacteria, including those with CTRX-resistance from year to year, the authors would like to point out the following comment of theirs made in 1989 and 1991, which appears to be increasing its significance, "Subjects of future studies should include dose on the mechanisms for the acquisition of bacterial resistance to entire beta-lactam antibiotics and the social circumstances in which resistant bacteria appear" . 5 . It appears that those strains resistant to cephems including CTRX are increasingly found among clinically isolated strains in recent years . CTRX, however, was found still effective against most clinical pathogens . Furthermore, considering that CTRX is one of the few drugs which sustain high blood concentrations of active forms we concluded that CTRX is a useful cephem-group antibiotic.

Antimicrob Agents Chemother, 1992 Jul, 36(7), 1418 - 23
In vitro activity and beta-lactamase stability of LJC 10,627; Neu HC et al.; The in vitro activity of LJC 10,627, a new carbapenem, was compared with those of imipenem, cefotaxime, ceftazidime, and gentamicin . LJC 10,627 inhibited 90% of Escherichia coli, Klebsiella pneumoniae, Klebsiella oxytoca, Enterobacter aerogenes, Enterobacter agglomerans, Enterobacter cloacae, Hafnia alvei, Citrobacter freundii, Citrobacter diversus, Proteus mirabilis, Morganella morganii, Proteus rettgeri, Serratia marcescens, Pseudomonas cepacia, salmonellae, shigellae, aeromonas, and yersiniae at less than or equal to 2 micrograms/ml . Haemophilus influenzae was inhibited by 0.5 microgram/ml, and moraxellae were inhibited by 0.12 microgram/ml . LJC 10,627 was twofold more active than imipenem against aerobic gram-negative organisms and inhibited ceftazidime-, cefotaxime-, and gentamicin-resistant members of the genera Klebsiella, Enterobacter, Citrobacter, and Serratia at less than or equal to 2 micrograms/ml . Xanthomonas maltophilia strains were resistant to the drug . Imipenem was two- to fourfold more active than LJC 10,627 against Staphylococcus aureus and Staphylococcus epidermidis . LJC 10,627 did not inhibit most methicillin-resistant Staphylococcus aureus or methicillin-resistant Staphylococcus epidermidis strains . LJC 10,627 inhibited Streptococcus pyogenes and Streptococcus pneumoniae at 0.06 and 0.12 microgram/ml, respectively . Bacteroides fragilis and other Bacteroides spp . were inhibited by 0.5 microgram of LJC 10,627 per ml . Serum (50%) did not affect the MICs . LJC 10,627 was not hydrolyzed by plasmid-mediated beta-lactamases of Bush types 2b, 2b', TEM-1, TEM-2, TEM-3, TEM-5, TEM-7, TEM-9, and SHV-1; the chromosomal beta-lactamases of Bush type 1; P-99; a Morganella enzyme; or a Citrobacter freundii enzyme . The Bush type 2c and 2d enzymes OXA-1, OXA-2, PSE-1, PSE-2, and PSE-4 did not hydrolyze LJC 10,627, nor did the beta-lactamases of Staphylococcus aureus, Moraxella spp., Bacteroides fragilis, and Proteus vulgaris . The beta-lactamase of Xanthomonas hydrolyzed LJC 10,627, albeit at approximately one-third the rate that imipenem was hydrolyzed.

Eur J Clin Microbiol Infect Dis, 1992 Jul, 11(7), 652 - 9
Comparative in vitro activity and beta-lactamase stability of RU29246, the active metabolite of HR916B; Yu KW et al.; HR 916B is a new orally absorbed cephalosporin . In tests its active metabolite, RU29246, inhibited Streptococcus pyogenes, Streptococcus agalactiae and Streptococcus pneumoniae at less than or equal to 0.12 micrograms/ml, which is similar to the antibacterial activity of cefuroxime, and was more active than cefaclor . It was also more active (MIC 2 micrograms/ml) than cefixime, cefuroxime, cefaclor and cefotaxime against staphylococci . RU29246 inhibited Escherichia coli, Klebsiella pneumoniae, Citrobacter diversus, Klebsiella oxytoca, Proteus mirabilis, Providencia stuartii and Salmonella spp . at less than or equal to 1 microgram/ml, thus being more active than cefuroxime and cefaclor, but was less active than cefixime and cefotaxime . It did not inhibit Pseudomonas aeruginosa and other Pseudomonas spp., Enterobacter spp., Serratia marcescens or Bacteroides fragilis . RU29246 was not hydrolyzed by TEM-1, Staphylococcus aureus TEM-2 or Moraxella catarrhalis beta-lactamases, but was hydrolyzed by TEM-3 and the chromosomal beta-lactamases of Proteus vulgaris and Morganella morganii . Plasmid and chromosomal beta-lactamases were inhibited by RU29246.

Mol Gen Mikrobiol Virusol, 1992 Jul-Aug, (7-8), 4 - 7
{Plasmid localization and cloning of restriction modification genes from Citrobacter freundii 4111 strain}; Kravets AN et al.; Over 60 producing strains of restriction endonucleases type II have been found among 500 different strains, mostly Enterobacteriaceae . The strain Citrobacter freundii 4111 produces restriction endonuclease CfrBI, a new isoschisomer of StyI . The genes of the restriction-modification system CfrBI were located on the multicopy plasmid pZE8 containing the Co1E1-type replicon and cloned to E . coli K802 . The deletion variant of 3.2-kb pZE8 which contains intact restriction-modification and a DNA fragment responsible for autonomous plasmid replication was selected among the recombinant plasmids . The strain with higher R . CfrBI production (at least 10,000,000 U/g cells, which is 500-fold higher than the wild strain) was constructed.

Clin Infect Dis, 1992 Jun, 14 Suppl 2, S189 - 94; discussion S195-6
Review of the in vitro antibacterial activity of cefprozil, a new oral cephalosporin; Thornsberry C; Cefprozil is a newer oral cephalosporin with a spectrum of activity against organisms that include gram-positive and gram-negative pathogens . A review of published data shows that cefprozil is active (susceptibility, less than or equal to 8 micrograms/mL; moderate susceptibility, 16 micrograms/mL; resistance, greater than or equal to 32 micrograms/mL) against gram-positive species such as streptococci, methicillin-susceptible staphylococci, and Listeria monocytogenes; it may have marginal activity against some enterococci . Among the gram-negative species, cefprozil has activity against Escherichia coli, Proteus mirabilis, Citrobacter diversus, Klebsiella pneumoniae, Klebsiella oxytoca, Haemophilus influenzae, and Moraxella catarrhalis . For anaerobic species, cefprozil has activity against clostridial species, including Clostridium difficile, peptostreptococci, and possibly Bacteroides melaninogenicus and Eubacterium . The activity of cefprozil is generally greater than that of cephalexin and generally similar to that of cefaclor . In these reports, cefprozil showed more in vitro activity than cephalexin and cefaclor against penicillin-resistant pneumococci, penicillin-resistant viridans streptococci, beta-lactamase-positive methicillin-susceptible Staphylococcus aureus, and C . difficile, although the clinical significance of some of these differences has yet to be studied.

Antimicrob Agents Chemother, 1992 Jun, 36(6), 1336 - 41
In vitro activity and susceptibility to hydrolysis of S-1006; Neu HC et al.; The in vitro activity of S-1006, the active component of a new orally absorbed cephalosporin, S-1108, inhibited 90% of Staphylococcus aureus isolates at less than or equal to 2 micrograms/ml, 90% of group A, B, C, F, and G streptococci and Streptococcus pneumoniae isolates at less than or equal to 0.12 microgram/ml, and all Haemophilus influenzae isolates at less than or equal to 0.06 microgram/ml . Although 50% of the members of the family Enterobacteriaceae were inhibited by less than or equal to 2 micrograms of S-1006 per ml, Enterobacter spp . and Citrobacter freundii resistant to ceftriaxone were resistant to S-1006 . The MICs of S-1006 for approximately 20% of Providencia, Proteus vulgaris, and Serratia isolates were 4 micrograms/ml . S-1006 was hydrolyzed by the plasmid TEM-3, TEM-5, PSE-1, and PSE-4 beta-lactamases and by the chromosomal beta-lactamase of Enterobacter and Morganella spp . and P . vulgaris.

Microb Releases, 1992 Jun, 1(1), 17 - 22
Mobilization of nonconjugative pBR322-derivative plasmids from laboratory strains of Escherichia coli to bacteria isolated from seawater; Sorensen SJ; Mobilization of derivatives of plasmids pBR322 and pBR325 was shown to occur between Escherichia coli K12 strains in LB-broth at 37 degrees C, provided a mobilizer plasmid (F') was present either together with the nonconjugative plasmid or in a second donor strain . Evidence from restriction endonuclease analysis suggested that the mobilization was facilitated by a transposition phenomenon involving the "gamma-delta" sequence of F' . It was shown that mobilization of a derivative of pBR325 from E . coli K12 to bacteria isolated from seawater occurred in incubations in both LB-broth and filtered seawater and that Pseudomonas sp., Enterobacter aerogenes, Klebsiella oxytoca, E . coli, and Citrobacter amalonaticus isolates were recipient-active.

FEBS Lett, 1992 May 18, 302(3), 256 - 60
The polypeptide chain fold in tyrosine phenol-lyase, a pyridoxal-5'-phosphate-dependent enzyme; Antson AA et al.; The tyrosine phenol lyase (EC 4.1.99.2) from Citrobacter intermedius has been crystallised in the apo form by vapour diffusion . The space group is P2(1)2(1)2 . The unit cell has dimensions a = 76.0 A, b = 138.3 A, c = 93.5 A and it contains two subunits of the tetrameric molecule in the asymmetric unit . Diffraction data for the native enzyme and two heavy atom derivatives have been collected with synchrotron radiation and an image plate scanner . The structure has been solved at 2.7 A resolution by isomorphous replacement with subsequent modification of the phases by averaging the density around the non-crystallographic symmetry axis . The electron density maps clearly show the relative orientation of the subunits and most of the trace of the polypeptide chain . Each subunit consists of two domains . The topology of the large domain appears to be similar to that of the aminotransferases.

Nihon Kyobu Shikkan Gakkai Zasshi, 1992 May, 30(5), 898 - 902
{A case of alveolar hydatid disease of the lung in the Kansai district of Japan}; Nishioka Y et al.; A 78-year-old man was admitted to hospital because of episodes of high grade fever and multiple nodular shadows on chest roentgenogram . He had a past history of percutaneous drainage and partial resection of the left lobe of the liver for liver abscess of unknown origin in 1987 . The high grade fever was secondary to sepsis due to Citrobacter freundii . The sepsis improved with antibiotic therapy, but the abnormal shadows on chest roentgenogram did not improved . Immunoserological tests indicated a probable diagnosis of alveolar hydatid disease of the lung, which is very rare in the Kansai district of Japan . Open lung biopsy was performed and the diagnosis of alveolar hydatid disease of the lung was confirmed.

J Antimicrob Chemother, 1992 May, 29(5), 563 - 73
Development of resistance during ceftazidime and cefepime therapy in a murine peritonitis model; Pechere JC et al.; Resistance emerging after ceftazidime or cefepime therapy was investigated in a peritonitis model . Mice were given a peritoneal challenge (10(8) cfu plus talcum) and treated by either antibiotic (50 mg/kg/dose, which produced similar antibiotic concentrations in peritoneal fluid in both cases) . After one or three doses, resistance never developed in Serratia marcescens or Citrobacter freundii infections . After Enterobacter cloacae and Pseudomonas aeruginosa challenge, ceftazidime selected more resistance (21/36 cases) than did cefepime (1/36 cases) . In mice challenged with resistant strains selected by ceftazidime therapy, cefepime (six doses) successfully treated 7/18 E . cloacae infections but 0/18 P . aeruginosa infections; ceftazidime was never effective . Neither cefepime nor ceftazidime cured mice infected with the resistant strain selected by cefepime . MICs were poor predictors of further emergence of resistance in mice inoculated with strains classified as susceptible, but antibiotic-containing agar gradients plated with a high inoculum (10(8) cfu) allowed better prediction . In selected clinical situations, cefepime may be preferable because it may be associated with less frequent emergence of resistance.

Diagn Microbiol Infect Dis, 1992 May-Jun, 15(4), 331 - 7
In vitro activity of cefquinome, a new cephalosporin, compared with other cephalosporin antibiotics; Chin NX et al.; The in vitro activity of cefquinome, a new aminothiazolyl cephalosporin with a C-3 bicyclic pyridinium group, was compared with ceftazidime, cefpirome, and cefepime . Cefquinome inhibited members of the Enterobacteriaceae at less than or equal to 0.5 microgram/ml for Escherichia coli, Klebsiella pneumoniae, K . oxytoca, Citrobacter diversus, Salmonella Shigella, Proteus mirabilis, Morganella, and Providencia . Although most Citrobacter freundii and Enterobacter cloacae were inhibited by less than 2 micrograms/ml, some strains resistant to ceftazidime were resistant, {minimum inhibitory concentration (MIC) greater than 16 micrograms/ml} . Serratia marcescens were inhibited by less than 1 microgram/ml and Pseudomonas aeruginosa by 8 micrograms/ml similar to the activity of cefepime . The majority of Haemophilus influenzae and Neisseria gonorrhoeae were inhibited by less than 0.25 microgram/ml . Most enterococci had cefquinome MICs of 4-8 micrograms/ml . Cefquinome was extremely active against group-A streptococci and Streptococcus pneumoniae with MICs less than 0.12 microgram/ml . 90% of methicillin-susceptible Staphylococcus aureus 90% were inhibited by 2 micrograms/ml . Overall, the in vitro activity of cefquinome was comparable with aminothiazolyl cephalosporins . It inhibited some Enterobacter and Citrobacter freundii resistant to ceftazidime as did cefpirome and cefepime . Cefquinome was not destroyed by the common plasmid beta-lactamases TEM-1, TEM-2, SHV-1, or by the chromosomal beta-lactamases of Klebsiella, Branhamella, and Pseudomonas, but it was hydrolyzed by TEM-3, TEM-5, and TEM-9 . Its activity was not adversely decreased in different medium or protein, and minimum bactericidal concentrations (MBCs) for most species except for Enterobacter were within a dilution of MICs.

Diagn Microbiol Infect Dis, 1992 May-Jun, 15(4), 291 - 4
Use of a commercial double-test tablet (Rosco PGUA/indole) for screening of Escherichia coli; Dominguez A et al.; A commercial double-test tablet (Rosco PGUA/indole) for detection of beta-glucuronidase (beta-GUR) activity and indole production was evaluated on a collection of 393 isolates of Enterobacteria . Both beta-GUR and indole were positive on 96.6% of Escherichia coli strains . beta-GUR, only, was also detected in 25 Shigella spp., four Enterobacter cloacae, eight Citrobacter freundii, and five Salmonella enteritidis strains, none of which were indole producers . An additional 261 consecutive clinical isolates of oxidase-negative nonswarming Gram-negative bacilli were studied in a parallel comparative field trial against conventional identification methods . For 200 strains, the standard method and PGUA/indole test were performed from the primary culture plate . The remaining 61 (23.4%) required subculture before testing . Sensitivity, specificity, positive predictive value, and negative predictive value of PGUA/indole test in the screening for E . coli were, respectively, 94.1%, 100%, 100%, and 87.1% . In our experience, PGUA/indole test is a rapid, precise, simple-to-perform, and economical method for screening E . coli . However, the need for a large inoculum may limit its application on primary cultures.

Jpn J Antibiot, 1992 May, 45(5), 489 - 501
{Clinical laboratory approach to evaluate efficacy of amikacin . Reevaluation of in vitro MIC break points in disc susceptibility test}; Uete G et al.; Antimicrobial activities of amikacin (AMK) against 269 strains of various clinical isolates obtained in 1989 were determined and the reliability of the AMK disc susceptibility test in estimating approximate values of MICs was studied . In addition, clinical significance of various systems for the interpretation of the disc tests was evaluated to determine more useful method in the evaluation of clinical efficacy of AMK . Included in the study were a 3 category system of NCCLS, a 4 category system used in Japan, and the system proposed by the British Society for Antimicrobial chemotherapy . In this study, MICs were determined using the Mueller-Hinton agar containing 50 mg/L of Ca and 25 mg/L of Mg at an inoculum level of 10(3-4) CFU/ml . MIC80 values of AMK against Staphylococcus aureus and Staphylococcus epidermidis were 6.25 and 25 micrograms/ml, respectively . Those against Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Proteus vulgaris, Enterobactor aerogenes, and Citrobacter spp . were less than 3.13 micrograms/ml . MIC values against Pseudomonas aeruginosa and Serratia marcescens were both less than or equal to 12.5 micrograms/ml . The disc susceptibility test was carried out according to the instruction in the Showa disc manual . Inhibition zones obtained with the disc method were compared with MIC values . The results of AMK disc susceptibility test obtained either with 30 micrograms discs or 10 micrograms discs were well correlated with MICs, indicating that the disc method was reliable in estimating approximate values of MICs (r = -0.807 to -0.897, P less than 0.01 in both instances) . In the 4 category classification system currently used in Japan, the break points in MIC values of AMK proposed are (+3) MIC less than or equal to 3 micrograms/ml, (+2) MIC greater than 3-15 micrograms/ml, and (+) MIC greater than 15-60 micrograms/ml . The results obtained with Showa 30 micrograms discs and 30 micrograms discs prepared in this laboratory showed false positive results in 13% and 10.8% of the samples, and false negative results in 1.5% and 3.3%, respectively . In the 3 category classification system of NCCLS, the MIC break points proposed are defined sensitive (S) for MIC less than or equal to 16 micrograms/ml and resistant (R) for MIC greater than or equal to 32 micrograms/ml . In this study, the 30 micrograms disc tests using the above-mentioned 2 different types of discs resulted in false positive responses in 6.3% and 5.2% of the samples tested and false negative results in 1.1% and 1.9% of the samples, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)

Antimicrob Agents Chemother, 1992 May, 36(5), 1049 - 52
Nucleotide sequence of the ampC-ampR region from the chromosome of Yersinia enterocolitica; Seoane A et al.; The nucleotide sequence of a 3.1-kb region from the chromosome of the Yersinia enterocolitica O:5b strain IP97 containing the gene for an inducible chromosomal cephalosporinase has been determined . The cephalosporinase gene was homologous to other enterobacterial chromosomal cephalosporinase genes, and it was accompanied by a gene homologous to the regulatory ampR gene . The arrangement of genes in the Y . enterocolitica ampCR unit was identical to that in the Enterobacter cloacae and Citrobacter freundii ampCR units.

Kinderarztl Prax, 1992 Apr, 60(2), 46 - 8
{The pathogen spectrum and its resistance behavior in children with urinary tract infections in Angola}; Eggert W et al.; In 225 children (135 boys, 90 girls) suffering from clinical relevant urinary tract infection the bacterial spectrum and resistance behaviour to routinely used antibiotics were evaluated . In 65.4% of the patients a significant bacteriuria was found: E . coli (34.6%), proteus (22.3%), klebsiella (14.6%), citrobacter (9.2%), enterobacter (5.4%) and pseudomonas (5.4%) . In testing a high resistance of bacterias to trimethoprim/sulfamethoxazol, ampicillin, and gentamycin was found, whereas good susceptibility was found to nitrofurantoin and nalidixin acid.

Ethiop Med J, 1992 Apr, 30(2), 79 - 88
Bacteriological quality of infant feeding bottle-contents and teats in Addis Abeba, Ethiopia; W/Tenssay Z et al.; Bacteriological quality of 244 infant feeding-bottles and 61 teats were studied from January 1989 to November 1989 . Samples were collected from feeding-bottles of babies who were brought by nursing mothers to clinics and hospitals for varying complaints . Of the 244 bottle-content samples, 144 (59%) harboured more than 2 x 10(6) colonies of bacteria per ml . of samples . The predominant bacterial isolates both from bottle-contents and teat-swabs were the coliform group of bacteria, such as E . coli, Enterobacter spp., Klebsiella spp., and Citrobacter spp . Enteric pathogens such as enteropathogenic E . coli, Shigella spp., S . aureus, and Bacillus cereus, were also isolated from some samples . In some cases enteropathogenic E . coli (EPEC) serotype were isolated both from feeding utensils and from the stools of the bottle-fed babies . Neither the different handling care of bottles used in the studied population, nor the educational status of the nursing mothers made a significant difference in the bacteriological quality of the infant feeding bottles in terms of the total bacterial counts per ml . of the bottle-contents . Preparation of food in advance of need combined with improper storage and inadequate cooking, and poor hygiene could be factors that resulted in a highly contaminated bottle contents . The superior quality of breast milk, and continual teaching of bottle feeding mothers on better handling of feeding bottles and feeds should be constantly emphasized . Teaching mothers on proper handling of feeding utensils accompanied by regular home visiting of health workers to observe hygienic condition of feeding utensils and home environment may improve the prevailing condition . Further and well controlled community based, longitudinal studies are also needed if we are to reduce gastroenteritis in babies as a result of using feeding bottles.

Epidemiol Infect, 1992 Apr, 108(2), 231 - 41
Further observations on the serological response to experimental Salmonella typhimurium in chickens measured by ELISA; Barrow PA; An indirect ELISA developed for the serological detection of Salmonella typhimurium in chickens using lipopolysaccharide as detecting antigen has been evaluated further in experimental infections . Following oral infection of 24-week-old laying hens with an invasive strain of S . typhimurium, high titres of specific circulating IgG were induced which were maintained for 20 weeks . Similar IgG titres were found in egg yolk . When 4-day-old chickens were infected high antibody titres persisted for 45 weeks . Chickens inoculated orally or intramuscularly with different numbers of S . typhimurium organisms showed graded serum IgG responses to LPS . The IgG titres in experimentally infected in-bred lines of chickens which showed greater genetic resistance to salmonella infection were significantly lower than those found in more susceptible lines . Oral and intramuscular infection with 18 different types of enterobacteria, including avian pathogenic E . coli, Citrobacter spp., Klebsiella spp., Proteus spp . and citrobacter-like organisms possessing some salmonella LPS (none possessed the O-4 antigen) and flagella antigens, did not induce S . typhimurium LPS-specific IgG responses . Chickens infected orally with rough or non-flagellate mutants of S . typhimurium did not induce high titres of LPS or flagella-specific IgG respectively . Sera obtained from S . typhimurium-infected chickens showed much higher titres against S . typhimurium LPS than with those antigens from other serotypes, including S . enteritidis.

J Med Microbiol, 1992 Apr, 36(4), 273 - 8
Citrobacter diversus brain abscess: case reports and molecular epidemiology; Morgan MG et al.; Citrobacter diversus brain abscess occurred in two infants in Aberdeen, 5 months apart . These are the first reported cases of this condition in the UK since 1976 . Restriction endonuclease analysis with SacI enzyme showed blood and CSF isolates from both patients to be identical and different from 10 other clinical isolates of C . diversus and one C . amalonaticus strain . Furthermore, isolates of C . diversus from patients belonged to biotype "d" whereas control isolates were of biotypes "a" or "e" . Because both infants attended the same central and peripheral maternity units, this raised suspicions of long term contamination of the hospital environment by this organism . Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns of whole-cell proteins and immunoblotting with normal human serum were remarkably homogeneous for all 13 C . diversus strains and thus were not useful for typing . However, the only C . amalonaticus strain was clearly differentiated from C . diversus strains by SDS-PAGE . Management of the infants included multiple intravenous antibiotic therapy for 4-6 weeks and repeated computerised tomography (CT) scanning and drainage of the abscess cavity . Both children survived albeit with some minor degree of brain damage.

FEMS Microbiol Immunol, 1992 Apr, 4(4), 201 - 7
Immunochemical characterization of Citrobacter strain PCM 1487 O-specific polysaccharide- and core oligosaccharide-protein conjugates; Lugowski C et al.; O-specific-polysaccharide and core oligosaccharide were isolated from Citrobacter strain PCM 1487 lipopolysaccharide and purified . The polysaccharide was selectively devoid of 4-deoxy-D-arabinohexose residues . Covalent conjugates of the modified O-specific polysaccharide and of the core oligosaccharide with tetanus toxoid were prepared . Immunochemical characterization of these conjugates proved that they are strong immunogens . Using monospecific rabbit antisera raised against the conjugates, the antigenic relationships between lipopolysaccharides of various strains of Citrobacter, Shigella sonnei, and Shigella flexneri were studied by quantitative microprecipitin and quantitative microprecipitin inhibition and immunoblotting tests.

Eur J Clin Microbiol Infect Dis, 1992 Apr, 11(4), 372 - 81
In vitro activity of OPC-17116 compared to other broad-spectrum fluoroquinolones; Sader HS et al.; The in vitro activity of OPC-17116 was compared to that of five similar fluoroquinolones (ciprofloxacin, enoxacin, norfloxacin, ofloxacin and temafloxacin) . A total of 700 isolates from recent cases of clinical bacteremia were tested . Fifty additional stock strains with well-characterized resistance mechanisms were also processed . The minimal concentrations inhibiting 90% of strains (MIC90) of Enterobacteriaceae species were for OPC-17116 0.015-0.5 micrograms/ml and for ciprofloxacin 0.015-0.25 micrograms/ml . Moraxella catarrhalis, Haemophilus influenzae and Neisseria gonorrhoeae were very susceptible to OPC-17116 (MIC90 0.015 micrograms/ml) thus being fourfold more active than ciprofloxacin . For all beta-hemolytic streptococci and pneumococci OPC-17116 MICs were less than or equal to 0.5 micrograms/ml . The most resistant enteric bacilli were among the Citrobacter freundii and Providencia rettgeri strains (MIC90 0.5 micrograms/ml) . Pseudomonas aeruginosa strains were comparably susceptible to OPC-17116 (MIC90 0.5 micrograms/ml) . Low pH and CO2 incubation had an adverse effect on OPC-17116 MICs, and resistance development was documented among current clinical isolates of staphylococci, pseudomonas and some Enterobacteriaceae.

J Clin Microbiol, 1992 Mar, 30(3), 590 - 4
Bacteriophage lytic patterns for identification of salmonellae, shigellae, Escherichia coli, Citrobacter freundii, and Enterobacter cloacae; He XQ et al.; A series of bacteriophages specific for Escherichia coli (E-1, E-2, E-3, and E-4), Citrobacter freundii (phi I, phi II, and phi III), Enterobacter cloacae (Ent), and Shigella spp . (Sh) have been isolated from hospital sewage . These bacteriophages, in combination with Felix Salmonella phage O-I, were used as a diagnostic phage typing set which included seven phage preparations: O-I, C (phi I and phi III), Sh, E (E-1 and E-2), CE (phi II and E-3), E-4, and Ent . After 20,280 cultures of 27 species and 9 biogroups of 15 genera of the family Enterobacteriaceae and 276 cultures of 8 species of 6 genera outside the Enterobacteriaceae were tested, it was shown that most strains of salmonellae, E . coli, C . freundii, and E . cloacae can be identified accurately . The sensitivities of identification were 83.6% for E . cloacae, 88.8% for C . freundii, 90.3% for E . coli, and 95.76% for salmonellae . The specificities were 99.78% for salmonellae, 99.84% for E . cloacae, 99.89% for E . coli, and 99.97% for C . freundii . The results of bacteriophage lytic patterns were highly correlated with Shigella serotypes . Therefore, such a phage typing set may be used routinely in public hygiene and clinical laboratories.

Pediatr Infect Dis J, 1992 Feb, 11(2), 99 - 104
Long term epidemiological analysis of Citrobacter diversus in a neonatal intensive care unit; Goering RV et al.; A prolonged outbreak of Citrobacter diversus central nervous system infection among hospitalized term infants, peaking in 1979, ceased with establishment of nurse-patient cohorting . The outbreak was attributed to dissemination of an epidemic strain among infants in an antiquated neonatal intensive care unit . When C . diversus colonization recurred within the new neonatal intensive care unit in 1984, cohorting and bacteriologic surveillance were reinstituted . By utilizing biotypes, plasmid profiles and antibiograms, four different C . diversus strains were identified circulating during 1979 . Strains recovered between 1984 and 1988 from neonatal intensive care unit infants were similar to those from community-acquired sources . A strain considered avirulent in 1979 was found causing bacteremia in two infants (one with central nervous system disease) in 1984 to 1988 . During cohorting C . diversus acquisition was 0.019/patient-month; after cohorting ceased it was 0.017/patient-month . Multiple source introductions appeared to occur with different C . diversus strains, some causing infant disease . No efficacy of cohorting was evident.

J Gen Microbiol, 1992 Feb, 138 ( Pt 2), 297 - 304
The Vi antigen of Salmonella typhi: molecular analysis of the viaB locus; Kolyva S et al.; Strains of Salmonella typhi isolated from the blood of patients with typhoid fever invariably express a capsular polysaccharide, termed the Vi antigen . Vi antigen expression is controlled by two separate chromosomal loci, viaA and viaB . The viaA locus is commonly found in enteric bacteria . In contrast, the viaB locus appears to be specific to Vi-expressing strains of Salmonella and Citrobacter . Here the cloning, expression and analysis of viaB determinants from S . typhi Ty2 is described . Whole-cell DNA from strain Ty2 was size-fractionated and cloned into the pLA2917 cosmid vector . A recombinant cosmid, pVT1, conferring a Vi-positive phenotype upon Escherichia coli and upon the Vi-non-expressing strain Ty21a of S . typhi, was characterized and used for further studies . Transposon Tn5 insertion mutagenesis demonstrated that the Vi-antigen-encoding region on pVT1 consisted of a 15 kb fragment . A subclone, designated pVT3, which contained an 18 kb insert, was sufficient to confer Vi antigen expression upon E . coli and S . typhi Ty21a . Results of recombination experiments indicated that this DNA sequence was the viaB locus of S . typhi Ty2 . In E . coli SE5000 maxicells, the viaB determinants encoded at least eight polypeptides, with molecular masses of 80, 65, 59, 48, 44, 39, 35 and 28 kDa . Functional characterization of viaB mutations in S . typhi Ty2 suggested that the 80 and 65 kDa proteins were required for cell-surface localization of the Vi antigen.

Res Microbiol, 1992 Feb, 143(2), 211 - 6
Novobiocin, brilliant green, glycerol, lactose agar: a new medium for the isolation of Salmonella strains; Poisson DM; 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.

Nippon Koshu Eisei Zasshi, 1992 Jan, 39(1), 45 - 9
{Monthly variation of viable count and coliforms count of sandpits in parks}; Hayashi M et al.; Field and experimental microbiological investigations of samples from sandpits from 5 parks were performed 2 times a month for 1 year, January-December 1990 . Total viable count from the sandpits was highest in the surface layer sample than the inner layer throughout the year . However, there was no such tendency in the number of Coliforms and Enterobacter, Escherichia coli, Citrobacter . The number of bacteria on the surface and inner layer was high from April to June, and in September and October, while it was low in July and August . The proportion of Coliform bacteria represented in the total viable count ranged from 2.1 to 8.9% in the surface layer and from 1.6 to 19.6% in the inner layer . There was a significant correlation between the numbers of bacteria on the surface and inner layer . Relationships between the total viable count and Coliform count and between Coliform count and E . coli, Enterobacter and Citrobacter were also observed . In view of the results it can be concluded that park sandpits are not, from a hygienic point of view, to be an ideal environment for children to play in.

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