Biomedical subjects

C M O'Hara

Publications and source records attributed to C M O'Hara.

At least 19 recordsLinked to original sources

PubMed2001

Performance of eight methods, including two new rapid methods, for detection of oxacillin resistance in a challenge set of Staphylococcus aureus organisms.

Using a set of 55 Staphylococcus aureus challenge organisms, we evaluated six routine methods (broth microdilution, disk diffusion, oxacillin agar screen, MicroScan conventional panels, MicroScan rapid panels, and Vitek cards) currently used in many clinical laboratories and two new rapid methods, Velogene and the MRSA-Screen, that require less than a day to determine the susceptibility of S. aureus to oxacillin. The methods were evaluated by using the presence of the mecA gene, as detected by PCR, as the "gold standard." The strains included 19 mecA-positive heterogeneously resistant strains of expression class 1 or 2 (demonstrating oxacillin MICs of 4 to >16 microg/ml) and 36 mecA-negative strains. The oxacillin MICs of the latter strains were 0.25 to 4 microg/ml when tested by broth microdilution with 2% NaCl-supplemented cation-adjusted Mueller-Hinton broth as specified by the NCCLS. However, when tested by agar dilution with 4% salt (the conditions used in the oxacillin agar screen method), the oxacillin MICs of 16 of the mecA-negative strains increased to 4 to 8 microg/ml. On initial testing, the percentages of correct results (% sensitivity/% specificity) were as follows: broth microdilution, 100/100; Velogene, 100/100; Vitek, 95/97; oxacillin agar screen, 90/92; disk diffusion, 100/89; MicroScan rapid panels, 90/86; MRSA-Screen, 90/100; and MicroScan conventional, 74/97. The MRSA-Screen sensitivity improved to 100% if agglutination reactions were read at 15 min. Repeat testing improved the performance of some but not all of the systems.

Humans↗

PubMed2000

Classification of Proteus vulgaris biogroup 3 with recognition of Proteus hauseri sp. nov., nom. rev. and unnamed Proteus genomospecies 4, 5 and 6.

Strains traditionally identified as Proteus vulgaris formed three biogroups. Biogroup 1, characterized by negative reactions for indole production, salicin fermentation and aesculin hydrolysis, is now known as Proteus penneri. Biogroup 2, characterized by positive reactions for indole, salicin and aesculin, was shown by DNA hybridization (hydroxyapatite method) to be a genetic species separate from biogroup 1 and from biogroup 3 which is positive for indole production and negative for salicin and aesculin. In this study, 52 strains were examined, of which 36 strains were Proteus vulgaris biogroup 3, which included the current type strain of the species P. vulgaris (ATCC 29905T), and compared to seven strains of Proteus vulgaris biogroup 2 and nine type strains of other species in the genera Proteus, Providencia and Morganella. By DNA hybridization, these 36 strains were separated into four distinct groups, designated as Proteus genomospecies 3, 4, 5 and 6. DNAs within each separate Proteus genomospecies were 74-99% related to each other in 60 degrees C hybridization reactions with < or = 4.5% divergence between related sequences. Proteus genomospecies 3 contained the former P. vulgaris type strain and one other strain and was negative in reactions for salicin fermentation, aesculin hydrolysis and deoxyribonuclease, unlike the reactions associated with strains considered as typical P. vulgaris which are positive in reactions for salicin, aesculin and DNase. Genomospecies 3 can be distinguished from Proteus genomospecies 4, 5 and 6 because it is negative for Jordan's tartrate. Proteus genomospecies 4, containing five strains, was differentiated from Proteus penneri, genomospecies 3 and 6 and most, but not all, strains of genomospecies 5, by its ability to ferment L-rhamnose. Proteus genomospecies 5 and 6, containing 18 and 11 strains, respectively, could not be separated from each other by traditional biochemical tests, by carbon source utilization tests or SDS-PAGE of whole-cell proteins. In an earlier publication, a request was made to the Judicial Commission that the former type strain of P. vulgaris (ATCC 13315) be replaced by P. vulgaris biogroup 2 strain ATCC 29905T, a strain considered more biochemically typical of P. vulgaris strains. This would have the effect of assigning the name P. vulgaris to P. vulgaris biogroup 2. Since this request has been acceded to, the name Proteus hauseri is herein proposed for Proteus vulgaris genomospecies 3. Its type strain is ATCC 700826T. Proteus genomospecies 4, 5 and 6 will remain unnamed until better phenotypic differentiation can be accomplished. All Proteus genomospecies were similar in their antimicrobial susceptibility patterns. Nineteen strains were isolated from urine, four from faeces, two from wounds, nine from other human sources and two from animals.

Anti-Bacterial Agents↗

PubMed2000

Classification, identification, and clinical significance of Proteus, Providencia, and Morganella.

This review presents the current taxonomy of the genera Proteus, Providencia, and Morganella, along with the current methods for the identification of each species within the three genera, incorporating both conventional biochemical and commercial methods. While all of these organisms are ubiquitous in the environment, individual case reports and nosocomial outbreak reports that demonstrate their ability to cause major infectious disease problems are presented. Lastly, anticipated antimicrobial susceptibility patterns are reviewed. Many of these organisms are easily controlled, but the advent of newer and more powerful antimicrobial agents has led to some problems of which laboratorians need to be aware.

Anti-Bacterial Agents↗

PubMed2000

Evaluation of the MicroScan rapid neg ID3 panel for identification of Enterobacteriaceae and some common gram-negative nonfermenters.

The MicroScan Rapid Neg ID3 panel (Dade Behring, Inc., West Sacramento, Calif.) is designed for the identification of gram-negative bacilli. We evaluated its ability to accurately identify Enterobacteriaceae that are routinely encountered in a clinical laboratory and glucose nonfermenting gram-negative bacilli. Using 511 stock cultures that were maintained at -70 degrees C and passaged three times before use, we inoculated panels according to the manufacturer's instructions and processed them in a Walk/Away instrument using version 22.01 software. The time to identification was 2 h and 30 min. All panel identifications were compared to reference identifications previously determined by conventional tube biochemicals. At the end of the initial 2.5-h incubation period, 405 (79.3%) identifications were correct. An additional 49 (9.6%) isolates were correctly identified after required additional off-line biochemical tests were performed. Thus, at 24 h, 88.8% of the 511 strains tested were correctly identified. Twenty-two (4.3%) were identified to the genus level only. Twenty-six (5.1%) strains were misidentified. Because the system is based on fluorogenics, there are no conventional tests readily available with which to compare possibly incorrect reactions. Of the 28 Salmonella strains that were tested, 5 were incorrectly reported. The 21 remaining errors were scattered among the genera tested. Testing on nine strains gave a result of "no identification" (very rare biotype). The Rapid Neg ID3 panel in this study approached 89% accuracy for the identification of gram-negative organisms encountered in the hospital laboratory.

Bacterial Typing Techniques↗

PubMed2000

Incidence and identification of Klebsiella planticola in clinical isolates with emphasis on newborns.

Studies conducted in France and Germany suggest that up to 19% of clinically identified Klebsiella sp. are actually Klebsiella planticola, an environmental species that has been attributed to two cases of septicemia, with a rare isolate of Klebsiella terrigena (0. 4%) being identified. A 1-year survey of newborns on a neonatal ward, also conducted in Germany, reported that 72% of Klebsiella sp. were Klebsiella oxytoca and 8.7% were K. planticola. The tests necessary to identify these species are not found in most clinical identification schemes or in the database matrices of most commercial identification products. To determine the incidence of unrecognized K. planticola among the Klebsiella sp. isolates in our collection, we used the battery of seven supplemental tests amended from the work of Monnet and Freney to test 352 stock isolates and 84 fresh clinical isolates from four local hospitals. After testing 436 strains of Klebsiella, only one strain was identified as a possible K. planticola and none was identified as K. terrigena. We tested an additional 43 stock strains of K. oxytoca isolated from newborns by using eight biochemical tests and found one additional strain of K. planticola. The occurrence of K. planticola in our collection is far less frequent than that observed in other countries.

Bacterial Typing Techniques↗

PubMed1999

Comparison of agar dilution, disk diffusion, MicroScan, and Vitek antimicrobial susceptibility testing methods to broth microdilution for detection of fluoroquinolone-resistant isolates of the family Enterobacteriaceae.

Fluoroquinolone resistance appears to be increasing in many species of bacteria, particularly in those causing nosocomial infections. However, the accuracy of some antimicrobial susceptibility testing methods for detecting fluoroquinolone resistance remains uncertain. Therefore, we compared the accuracy of the results of agar dilution, disk diffusion, MicroScan Walk Away Neg Combo 15 conventional panels, and Vitek GNS-F7 cards to the accuracy of the results of the broth microdilution reference method for detection of ciprofloxacin and ofloxacin resistance in 195 clinical isolates of the family Enterobacteriaceae collected from six U.S. hospitals for a national surveillance project (Project ICARE [Intensive Care Antimicrobial Resistance Epidemiology]). For ciprofloxacin, very major error rates were 0% (disk diffusion and MicroScan), 0.9% (agar dilution), and 2.7% (Vitek), while major error rates ranged from 0% (agar dilution) to 3.7% (MicroScan and Vitek). Minor error rates ranged from 12.3% (agar dilution) to 20.5% (MicroScan). For ofloxacin, no very major errors were observed, and major errors were noted only with MicroScan (3.7% major error rate). Minor error rates ranged from 8.2% (agar dilution) to 18.5% (Vitek). Minor errors for all methods were substantially reduced when results with MICs within +/-1 dilution of the broth microdilution reference MIC were excluded from analysis. However, the high number of minor errors by all test systems remains a concern.

Anti-Infective Agents↗

PubMed1999

Biochemical identification of Citrobacter species defined by DNA hybridization and description of Citrobacter gillenii sp. nov. (formerly Citrobacter genomospecies 10) and Citrobacter murliniae sp. nov. (formerly Citrobacter genomospecies 11).

Recent work describing six named species and two unnamed genomospecies within Citrobacter has enlarged the genus to 11 species. DNA relatedness and phenotypic tests were used to determine how well these species can be identified. One hundred thirty-six strains were identified to species level by DNA relatedness and then identified phenotypically in a blinded fashion. By using conventional tests, 119 of the 136 strains (88%) were correctly identified to species level. Three additional strains (2%) were identified as citrobacteria but were not identified to species level, and 14 strains (10%) were misidentified as other Citrobacter species. Carbon source utilization tests were used to identify 86 of the strains. Eighty-four strains (98%) were correctly identified, and two strains (2%) were misidentified as other Citrobacter species. Additional strains of Citrobacter genomospecies 10 and Citrobacter genomospecies 11 were identified, allowing these species to be formally named as Citrobacter gillenii sp. nov. and Citrobacter murliniae sp. nov., respectively.

Citrobacter↗

PubMed1998

Two new Rahnella genomospecies that cannot be phenotypically differentiated from Rahnella aquatilis.

Fifty-one Rahnella aquatilis and R. aquatilis-like strains from water, snails and human sources were characterized by routine biochemical tests, carbon source utilization tests, DNA relatedness (hydroxyapatite method) and 16S rRNA sequencing. The results of the genetic methods indicated that the strains comprised three closely related species within the genus Rahnella. It was not possible to differentiate R. aquatilis from the two newly recognized species. The new species were therefore given the vernacular names Rahnella genomospecies 2 and Rahnella genomospecies 3.

Animals↗

PubMed1998

First report of a human isolate of Erwinia persicinus.

Erwinia persicinus was first described in 1990 after being isolated from a variety of fruits and vegetables, including bananas, cucumbers, and tomatoes. In 1994, it was shown to be the causative agent of necrosis of bean pods. We now report the first human isolate of E. persicinus. The strain was isolated from the urine of an 88-year-old woman who presented with a urinary tract infection. By the hydroxyapatite method, DNA from this strain was shown to be 94.5% related at 60 degrees C and 86% related at 75 degrees C to the type strain of E. persicinus. The biochemical profile of E. persicinus is most similar to those of Erwinia rhapontici, Pantoea agglomerans, and Enterobacter species. It is negative in tests for lysine, arginine, ornithine, dulcitol, and urea. It is motile and positive in tests for D-sorbitol and sucrose. It is susceptible to the expanded-spectrum cephalosporins, aminoglycosides, and fluoroquinolones, but it is resistant to ampicillin, ticarcillin, and cefazolin.

Aged↗

PubMed1998

Isolation of Enterobacter intermedium from the gallbladder of a patient with cholecystitis.

We describe the isolation and identification of Enterobacter intermedium from the gallbladder of a patient with cholecystitis. There have been only four documented isolations of this organism from humans; it normally occurs in surface water and unpolluted soils. The identification was initially made by a MicroScan Walk/Away system with a Neg Combo 18 conventional identification-susceptibility panel. The organism is susceptible to the aminoglycosides and imipenem but resistant to the cephalosporins and ciprofloxacin.

Anti-Bacterial Agents↗

PubMed1998

Characterization of staphylococci with reduced susceptibilities to vancomycin and other glycopeptides.

During the last several years a series of staphylococcal isolates that demonstrated reduced susceptibility to vancomycin or other glycopeptides have been reported. We selected 12 isolates of staphylococci for which the vancomycin MICs were > or =4 microg/ml or for which the teicoplanin MICs were > or =8 microg/ml and 24 control strains for which the vancomycin MICs were < or =2 microg/ml or for which the teicoplanin MICs were < or =4 microg/ml to determine the ability of commercial susceptibility testing procedures and vancomycin agar screening methods to detect isolates with reduced glycopeptide susceptibility. By PCR analysis, none of the isolates with decreased glycopeptide susceptibility contained known vancomycin resistance genes. Broth microdilution tests held a full 24 h were best at detecting strains with reduced glycopeptide susceptibility. Disk diffusion did not differentiate the strains inhibited by 8 microg of vancomycin per ml from more susceptible isolates. Most of the isolates with reduced glycopeptide susceptibility were recognized by MicroScan conventional panels and Etest vancomycin strips. Sensititre panels read visually were more variable, although with some of the panels MICs of 8 microg/ml were noted for these isolates. Vitek results were 4 microg/ml for all strains for which the vancomycin MICs were > or =4 microg/ml. Vancomycin MICs on Rapid MicroScan panels were not predictive, giving MICs of either < or =2 or > or =16 microg/ml for these isolates. Commercial brain heart infusion vancomycin agar screening plates containing 6 microg of vancomycin per ml consistently differentiated those strains inhibited by 8 microg/ml from more susceptible strains. Vancomycin-containing media prepared in-house showed occasional growth of susceptible strains, Staphylococcus aureus ATCC 29213, and on occasion, Enterococcus faecalis ATCC 29212. Thus, strains of staphylococci with reduced susceptibility to glycopeptides, such as vancomycin, are best detected in the laboratory by nonautomated quantitative tests incubated for a full 24 h. Furthermore, it appears that commercial vancomycin agar screening plates can be used to detect these isolates.

Anti-Bacterial Agents↗

PubMed1997

Evaluation of Vitek GNI+ and Becton Dickinson Microbiology Systems Crystal E/NF identification systems for identification of members of the family Enterobacteriaceae and other gram-negative, glucose-fermenting and non-glucose-fermenting bacilli.

We evaluated the Vitek GNI+ and Becton Dickinson Crystal E/NF identification systems for their ability to accurately identify 619 and 626 strains, respectively, of members of the family Enterobacteriaceae and other glucose-fermenting and non-glucose-fermenting gram-negative rods. All strains tested were taken from a stock collection and passed three times on 5% sheep blood agar prior to testing. These strains represented a more rigorous challenge to both systems than one resulting from the testing of consecutive clinical isolates. Testing with both systems was done according to the manufacturers' instructions, and tests were repeated in duplicate when errors occurred. Vitek version 5.01 and Crystal version 3.0 softwares were used for identifications. The identification results from each system were compared with identifications previously determined with reference biochemicals. At the completion of the appropriate incubation period, the GNI+ and Crystal systems correctly identified 80.1 and 71.1% of the total isolates, respectively. After additional tests suggested by the software programs were completed, the GNI+ had an accuracy of 87.6% and the Crystal system's accuracy had improved to 87.9%. The error rates for the GNI+ and Crystal systems were 6.5 and 5.3%, respectively. A report of "no identification" was given for 6.0 and 6.9% of the isolates, respectively, and was associated with no particular organism group. One isolate each of Acinetobacter lwoffii and Vibrio alginolyticus would not grow in the Vitek card. The average times to detection for correct enteric identifications in the GNI+ system were 4.1 and 6.8 h for nonenteric identifications, while the Crystal results were routinely read at 18 h. We conclude that there was no significant difference (P > 0.05) between the results of the GNI+ card and those of the Crystal E/NF system after additional testing was performed with the group of organisms tested, but the overall accuracy for both systems in this study was below 90%.

Bacterial Typing Techniques↗

PubMed1996

Inhibition of dopamine synthesis by dopamine D2 and D3 but not D4 receptors.

The goal of the current study was to determine which of the D2-like receptors (D2, D3 or D4) are involved in autoreceptor regulation of dopamine synthesis. We have derived a model system utilizing a mouse mesencephalic cell line, MN9D, which both synthesizes and releases dopamine, to characterize the modulation of tyrosine hydroxylase activity, the rate limiting enzyme in the conversion of tyrosine to dopamine, by the D2-like receptors. Previously, we have shown that stimulation of D2 and D3, but not D4, dopamine receptors transfected into MN9D cells inhibited the release of dopamine. In the current study, we show that quinpirole stimulation of transfected D2 and D3, but not D4, dopamine receptors inhibited K+-stimulated tyrosine hydroxylase activity in a pertussis toxin-sensitive manner, strongly suggesting G-protein coupling as a mechanistic pathway. The D2 receptor effect could be maintained for at least 60 min, whereas the D3 receptor effect desensitized. Treatment with 10 microM forskolin, which raises cyclic AMP levels or with 100 nM okadaic acid, a potent phosphatase inhibitor, had no effect on the D2-or D3-mediated inhibition, suggesting that these effects may be independent of both cyclic AMP- and okadaic acid-sensitive phosphatase activity. Taken together, these data confirm the hypothesis that dopamine D2 and D3 receptors can perform dual roles in autoreceptor regulation.

Animals↗

PubMed1996

Dopamine D2L receptor couples to G alpha i2 and G alpha i3 but not G alpha i1, leading to the inhibition of adenylate cyclase in transfected cell lines.

Previously, we showed that both D2 and D4 dopamine receptors inhibited adenylate cyclase in a pertussis toxin (Ptx)-sensitive manner in the dopamine-producing MN9D cell line, whereas only D2 receptors did so in a fibroblast cell line, CCL1.3. Of the known Ptx-sensitive G proteins, MN9D cells expressed G alpha i2, G alpha oA and G alpha oB, whereas CCL1.3 cells expressed only G alpha i2. Here we cotransfected MN9D and CCL1.3 cells with either the long form of the D2 receptor (D2L) or the D4 receptor and a mutant Ptx-resistant G protein alpha-subunit. When cotransfected CCL1.3 cell lines were tested for the ability of Ptx to block receptor-mediated inhibition of cyclic AMP accumulation, D2 receptors were found to couple to mutant G alpha i2 and G alpha i3 but not G alpha i1 or G alpha oA. D2 also coupled to mutant G alpha i2 but not G alpha oA in MN9D cells. In contrast, D4 receptors did not couple to either mutant G alpha i2 or G alpha oA subunits in MN9D cells. These data suggest that D4 receptor-mediated inhibition of adenylate cyclase is not coupled via the same mechanisms used by D2 receptors. D2L receptors are capable of coupling to more than one G protein in the modulation of cyclic AMP.

Adenylyl Cyclases↗

PubMed1995

Replacement of NCTC 4175, the current type strain of Proteus vulgaris, with ATCC 29905. Request for an opinion.

The current type strain of Proteus vulgaris, NCTC 4175 (= ATCC 13315), differs substantially from typical strains of this species both biochemically and chemotaxonomically. DNA relatedness studies revealed that strains previously classified as P. vulgaris belong to six genomospecies. One of these genomospecies contains strains that are negative in indole, salicin, and esculin reactions (biogroup 1) and has been named Proteus penneri. A second genomospecies, which is most frequently isolated from human urine, contains typical P. vulgaris strains that are positive in indole, salicin, and esculin reactions (biogroup 2). The members of the remaining four genomospecies are indole positive and negative in salicin and esculin reactions (biogroup 3). Of 36 biogroup 3 strains studied, only strain NCTC 4175T (T = type strain) and one other strain, CDC 1732-80, belong to genomospecies 3. To retain NCTC 4175 as the type strain of P. vulgaris would restrict this species to these two strains, whose origins are unknown. This would mean that hundreds of strains for which the description of P. vulgaris was written and which have been representatives of this species for the past 50 years would have to be renamed as members of a new species. To prevent this confusion, we request that biogroup 2 reference strain ATCC 29905 (= CDC PR1) replace NCTC 4175 as the type strain of P. vulgaris.

DNA, Bacterial↗

PubMed1995

Ability of commercial identification systems to identify newly recognized species of Citrobacter.

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.

Citrobacter↗

PubMed1995

Ability of commercial and reference antimicrobial susceptibility testing methods to detect vancomycin resistance in enterococci.

We evaluated the abilities of 10 commercially available antimicrobial susceptibility testing methods and four reference methods (agar dilution, broth microdilution, disk diffusion, and the agar screen plate) to classify enterococci correctly as vancomycin susceptible or resistant using 50 well-characterized strains of enterococci. There was a high level of agreement of category classification data obtained with broth-based systems (Sceptor, MicroMedia, Pasco, and Sensititre), agar dilution, and an antibiotic gradient method (E test) with data obtained by reference broth microdilution; no very major or major errors were seen, and minor errors were < or = 6%. Increased minor error rates were observed with disk diffusion (12%), Alamar (16%), Uniscept (16%), and conventional (overnight) MicroScan panels (16%). The errors were primarily with Enterococcus casseliflavus strains and organisms containing the vanB vancomycin resistance gene. Very major error rates of 10.3 and 20.7% were observed with Vitek and MicroScan Rapid (MS/Rapid) systems, respectively; however, only the MS/Rapid system produced major errors (13.3%). On repeat testing of discrepant isolates, the very major error rate with the Vitek system dropped to 3.4%, while the very major error rate with the MS/Rapid system increased to 27.6%; major errors with the MS/Rapid system were not resolved. Many of the commercial systems had only 4 dilutions of vancomycin, which resulted in up to 84% of values being off scale (e.g., Uniscept). Of the methods tested, most conventional broth- and agar-based methods proved to be highly accurate when incubation was done for a full 24 h, although several of the tests had high minor error rates. Automated systems continued to demonstrate problems in detecting low-level resistance.

Automation↗

PubMed1995

Genetic and biochemical characterization of Citrobacter rodentium sp. nov.

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.

Animals↗