bluuebunny/crossref_metadata_2025 huggingface dataset:
Interpretive standards and quality control guidelines for imipenem susceptibility tests with 10-micrograms disks
Authors: Barry A L , Thornsberry C , Gavan T L , Jones R N
Published: January 2020

Tests with 10-micrograms imipenem disks accurately categorized 98.5% of 551 bacterial isolates when interpretive breakpoints of less than or equal to 13 mm for resistant and greater than or equal to 16 mm for susceptible were used. Because a sufficient number of resistant or moderately susceptible strains were not available for testing, these interpretive standards must be considered tentative. Quality control limits for tests with Escherichia coli ATCC 25922 and Pseudomonas aeruginosa ATCC 27853 are 26 to 32 and 20 to 28 mm, respectively. Zones obtained with Staphylococcus aureus ATCC 25923 were too large and variable to be useful for quality control purposes.
Prosthetic valve endocarditis caused by group Ve-1 bacteria (Chromobacterium typhiflavum)
Authors: O'Leary T , Fong I W
Published: January 2020

A case of fatal prosthetic valve endocarditis was caused by group Ve-1 bacteria. Bacteriological characteristics and antibiotic susceptibilities are presented, as well as a brief discussion of the isolates of the Ve-1 organism in Ontario.
Identification of a species-specific antigen in Legionella pneumophila by a monoclonal antibody
Authors: Gosting L H , Cabrian K , Sturge J C , Goldstein L C
Published: January 2020

A species-specific antigen in Legionella pneumophila was identified by a monoclonal antibody in enzyme-linked immunosorbent and immunofluorescence assays of serogroups 1 through 8. The species-specific antigen was heat stable, and the molecular weight of the major band was 29,000 by immunoblot analysis. In direct immunofluorescence assays, the antigen was cryptic or only partly exposed on the surface of the cells but was effectively exposed by treating the cells with detergent and EDTA. The monoclonal antibody was utilized in direct immunofluorescence assays to specifically identify multiple cultures of L. pneumophila serogroups.
Serological cross-reactions between Mycoplasma genitalium and Mycoplasma pneumoniae
Authors: Lind K , Lindhardt B O , Schütten H J , Blom J , Christiansen C
Published: January 2020

The recently discovered mycoplasma species Mycoplasma genitalium was isolated from urethral specimens from men with nongonococcal urethritis (Tully et al., Lancet i:1288-1291, 1981). In a previous report (K. Lind, Lancet ii:1158-1159, 1982), prominent serological cross-reactions were demonstrated between this mycoplasma and M. pneumoniae. In the present study, the two mycoplasma species were compared more extensively. In classical mycoplasma medium without thallium acetate, M. genitalium grew more slowly than M. pneumoniae did but finally formed similar amounts of acetic acid and lactic acid from glucose. Although their colonies on solid medium were indistinguishable, transmission electron microscopy showed that the flask-formed cells of M. genitalium (especially their necks) were shorter than those of M. pneumoniae. The two species were distinct since DNA hybridization showed only 1.8% homology in base sequences, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed significantly different profiles of the two strains. However, considerable similarities were found in their antigenic reactions in various serological tests. The presence of common or closely related antigens was demonstrated in the two species with rabbit immune sera in complement fixation test with chloroform-methanol-extracted antigens by an indirect immunofluorescence test on microcolonies, and by metabolism inhibition and growth inhibition tests. Cross-reactions were also demonstrated by crossed immunoelectrophoresis. The role of M. genitalium as a human pathogen in the genital tract has not been assessed. If serological tests are to be used in this assessment, caution must be exercised due to the extensive cross-reactions demonstrated. Some of the species-specific antigens which we have demonstrated would be appropriate for use in such tests and would help to circumvent problems caused by cross-reactions.
Evaluation of the AutoMicrobic system for identification and susceptibility testing of gram-negative bacilli
Authors: Woolfrey B F , Lally R T , Ederer M N , Quall C O
Published: January 2020

The AutoMicrobic system (AMS) (Vitek Systems, Inc., Hazelwood, Mo.) was compared with the API-20E system for the identification of gram-negative bacilli by using 380 stock clinical isolates and 377 immediately encountered fresh clinical isolates. For the stock isolates, with Enterobacteriaceae-Plus Biochemical Cards and automated interpretation, 364 (95.8%) were in agreement to the species level. For the fresh clinical isolates, agreement at the genus and species levels was 89.7 and 85.9%, respectively, when Enterobacteriaceae-Plus Cards were interpreted by the AMS. Manual interpretation of Enterobacteriaceae-Plus Biochemical Cards improved species level agreement to 91.0%. Subsequent retesting of all discrepant isolates with the Gram-Negative Identification Card resulted in significant improvement of results, and for the stock and fresh clinical isolates, species level agreement was 98.7 and 97.3%, respectively. AMS susceptibility testing was evaluated by comparing ampicillin and cephalothin MICs determined in parallel by AMS and a reference broth microdilution test for stock isolates, and by comparison of AMS and standardized disk agar diffusion test results for fresh clinical isolates. For the stock isolates, AMS mean integer MICs approximated microdilution mean integer MICs with AMS, providing excellent MIC replicability. For ampicillin and cephalothin, 50 and 46.8%, respectively, of AMS integer MICs were within +/- 1 microgram/ml of the reference values, and 89.3 and 63.1% of AMS integer MICs were within +/- 2 micrograms/ml of the reference values. For the fresh clinical isolates, AMS and reference results were in disagreement for 4.5% of the antimicrobial agents tested, with 2.3% as a combination of "major" and "very major" errors.
Comparison of anaerobic susceptibility results obtained by two methods of inoculum preparation
Authors: Bourgault A M , Lamothe F
Published: January 2020

We evaluated the use of inocula prepared directly from blood agar plates in agar dilution susceptibility tests of anaerobic bacteria and compared the results with susceptibility results obtained from the National Committee for Clinical Laboratory Standards proposed thioglycolate broth cultures. The objectives were to evaluate the reproducibility of each of the two methods of inoculum preparation and to compare the MICs obtained by each method. The reproducibility studies were conducted on 14 stock strains. The mode MICs obtained by the direct agar method were identical to those obtained by the reference broth method 74% of the time and within +/- 1 log2 dilution 100% of the time. The degree of reproducibility of each of the two methods was identical (93% +/- 1 log2 dilution). MIC results obtained by the direct agar method agreed with the MICs obtained by the reference broth culture method in 92.9% of 1,125 MIC data pair determinations performed on stock cultures. The reproducibility of the direct agar method within +/- 1 log2 dilution step for 115 fresh clinical isolates was 93%, including 93.4% of the results with the Bacteroides fragilis group. Only two very major discrepancies (false-susceptible by the agar method) were identified among the 708 MIC data pairs on these clinical isolates. Preparation of inocula directly from growth on agar plates provides a rapid and reproducible method for agar dilution susceptibility testing of anaerobes.
Detection of methicillin-resistant Staphylococcus aureus by microdilution and disk elution susceptibility systems
Authors: Boyce J M , Lytle L S , Walsh D A
Published: January 2020

To determine whether methicillin-resistant (MR) Staphylococcus aureus from different geographic areas are detected reliably by various commercially available microdilution broth and disk elution systems, 73 such isolates obtained from hospitals in 13 cities were tested by a reference method (agar dilution) and by the Microscan, API 3600S, Autobac I, and MS-2 systems. Both Eugonic broth and Low Thymidine Eugonic broth were used in the evaluation of the Autobac I, and two versions of the MS-2 were used. The proportions of isolates categorized as MR by the various methods were: agar dilution method, 99%; Microscan, 100% (if the suggested cut-off of the manufacturer was used); API 3600S, 96%; Autobac I, 84 to 93%; and MS-2, 54 to 68%. With the MS-2 system, isolates from Jackson, Miss., were classified as susceptible to methicillin more often than were strains from other cities. With the Autobac I (Eugonic broth), only 55% of isolates from Houston, Tex., were classified as MR, whereas 89% of isolates from all other cities were correctly classified as MR. With the API 3600S, strains from some cities were categorized as nafcillin susceptible, whereas strains from other cities were classified as resistant to nafcillin. The results of this study suggest that future evaluations of antimicrobial susceptibility testing systems should include MR strains of S. aureus from several geographic areas.
Analytical accuracy of determinations of aminoglycoside concentrations by enzyme multiplied immunoassay, fluorescence polarization immunoassay, and radioimmunoassay in the presence of heparin
Authors: O'Connell M E , Heim K L , Halstenson C E , Matzke G R
Published: January 2020

The accuracy of gentamicin, netilmicin, and tobramycin concentration determinations by enzyme multiplied immunoassay technique (EMIT; Syva Corp., Palo Alto, Calif.), fluorescence polarization immunoassay (TDx; Abbott Diagnostics, Irving, Tex.), and radioimmunoassay were compared in the presence of 0 to 3,000 USP units of porcine heparin per ml. Gentamicin, netilmicin, and tobramycin concentrations determined by EMIT decreased by 10 and 50% in the presence of 75 and 1,000 USP units/ml, 2 and 5 USP units/ml, and 2 and 7.5 USP units/ml, respectively. Accuracy of the TDx and radioimmunoassay determinations, however, were not affected by the presence of heparin. Blood samples for the determination of gentamicin, netilmicin, and tobramycin by EMIT should not be collected in evacuated heparinized tubes.
Composition of the antigenic material removed from Campylobacter jejuni by heat
Authors: Buck G E , Smith J S , Parshall K A
Published: January 2020

The antigenic material removed form Campylobacter jejuni by the boiling of whole cells in saline was examined biochemically. Analyses showed that the extracted material contained 3 micrograms of protein per ml per mg of wet cells and ca. 2.6 micrograms of carbohydrate per ml per mg of wet cells. Further extraction of the material with chloroform-methanol produced about 0.5 microgram of water-insoluble residue per ml per mg of wet cells, suggesting the presence of lipid as well. Additional analyses revealed the presence of hexose, pentose, heptose, hexosamine, and 2-keto-3-deoxyoctonic acid, and the extract was also positive by the Limulus amoebocyte lysate assay for lipopolysaccharide. An examination by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that at least 10 different protein bands could be detected. One of the major bands corresponded to the major outer membrane protein, as determined by comparison with an outer membrane protein preparation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Another major protein in the heated extract corresponded to a band previously shown to be flagellin. An analysis of the time course of the release of material showed that a significant amount was removed after 3 to 10 min at 100 degrees C, but the release of material seemed to be delayed at lower temperatures. These results show that the treatment of C. jejuni with heat produces a complex mixture of components, including cell wall lipopolysaccharide, the major outer membrane protein, and flagellin. It is likely that some cytoplasmic components are present as well. Blebs of outer membrane have been observed with this organism by electron microscopy. Our results confirm this and suggest that the heating of cells accelerates this blebbing process.
Double-conjugate enzyme-linked immunosorbent assay for immunoglobulins G and M against Treponema pallidum
Authors: Farshy C E , Hunter E F , Larsen S A , Cerny E H
Published: January 2020

An enzyme-linked immunosorbent assay (ELISA) for the simultaneous measurement of immunoglobulin G (IgG) and IgM was developed to detect antibodies to Treponema pallidum. Wells of polystyrene microtiter plates were coated with T. pallidum antigen, diluted patient serum was added, and IgG and IgM which bound to the T. pallidum antigen were measured by the simultaneous addition of alkaline phosphatase-labeled anti-human IgG and horseradish peroxidase-labeled anti-human IgM. Bound IgG was detected first, followed by bound IgM. After development of the procedure, 145 categorized sera were evaluated: 60 from individuals without syphilis; 62 from patients with syphilis, including 22 with primary, 20 with secondary, and 20 with latent phases of syphilis; and 23 from patients with rheumatoid arthritis. Of the 60 sera from individuals without syphilis, 100% were nonreactive for IgG antibody and 16% were reactive for IgM. Of the 23 sera from patients with rheumatoid arthritis, 3 were reactive for IgG and 3 were nonreactive for IgM. Of the 62 sera from patients with syphilis, 61 (98%) were reactive for IgG antibody with increased titers as the stage of syphilis increased, whereas IgM reactivity decreased. This enzyme-linked immunosorbent assay appears to be a simple method for the simultaneous measurement of antibodies under equal assay conditions.
Nutritionally variant streptococci from corneal ulcers in horses
Authors: Higgins R , Biberstein E L , Jang S S
Published: January 2020

Of 24 isolates of nutritionally variant streptococci recovered from equine corneal ulcers, 22 were tested for growth requirements, physiological and biochemical reactions, and susceptibility to different antimicrobial agents. Satisfactory growth was obtained by supplementing blood agar and Todd-Hewitt broth with pyridoxal hydrochloride, and all of the media for the culture and the biochemical testing were supplemented with 0.002% of this substance. Biochemical patterns of 12 of the isolates resembled those of two viridans streptococcal species, Streptococcus intermedius and Streptococcus constellatus. Patterns of 10 isolates did not resemble those of any recognized viridans species. All of the isolates were inhibited by less than or equal to 0.25 microgram of erythromycin per ml, less than or equal to 4 micrograms of chloramphenicol per ml, and less than or equal to 4 micrograms of gentamicin per ml, and all but two were inhibited by less than or equal to 0.1 microgram of penicillin per ml.
Multicenter evaluation of a 1-h enzyme-linked immunosorbent assay for rubella serology
Authors: Boteler W L , Barnes K J , Buimovici-Klein E , O'Beirne A J
Published: January 2020

A 1-h enzyme-linked immunosorbent assay (Rubestat) was developed for rubella virus immunoglobulin G detection. The assay used phenolphthalein monophosphate as the substrate, which, when developed, can easily be read visually. Rubestat compared very favorably to hemagglutination inhibition and commercial enzyme-linked immunosorbent assays in its ability to determine immune status. Rubestat demonstrated greater than 97% specificity, sensitivity, and accuracy as compared with other methodologies at 10 different laboratories. The Rubestat index values were precise, with coefficients of variation for intra- and interassay variation of less than 10%. Mean index values had a linear correlation with hemagglutination inhibition titers (r2 greater than 0.97). A population distribution of index values illustrated two distinct bell-shaped curves representing the positive and negative populations. Studies of acute and convalescent serum pairs showed Rubestat to be as accurate as hemagglutination inhibition in determining seroconversion.
Feasible method for routine surveillance culturing of stools from neutropenic patients
Authors: Smith J A , Sherlock C H , Burdge D R
Published: January 2020

This study was undertaken to develop an accurate, yet inexpensive, method for determining whether the bowel of a neutropenic patient is colonized with bacteria resistant to the antimicrobial agents used in empiric therapy. Selective agar media were prepared in which Mueller-Hinton agar or MacConkey agar were supplemented with one of the following antimicrobial agents: carbenicillin (16 micrograms/ml), gentamicin (4 micrograms/ml), or tobramycin (4 micrograms/ml). Moxalactam was incorporated initially at 16 micrograms/ml and subsequently at 8 micrograms/ml. Stools from neutropenic patients and bone marrow transplant recipients were inoculated on these media and on unsupplemented MacConkey agar. All bacteria that grew on the antibiotic-containing media were categorized as resistant to the supplementing drug; failure to detect an organism that did grow on the antibiotic-free MacConkey agar indicated susceptibility. These results were compared with those obtained for all isolates on all media by agar disk diffusion. There were 512 gram-negative enteric isolates from 320 stools obtained from 98 patients. The antibiotic-containing media suppressed the growth of 95% of bacteria that were identified as susceptible by agar disk diffusion. In detecting resistant organisms, the correlation between agar disk diffusion and direct stool screening with Mueller-Hinton agar ranged from 73 to 83%, and on MacConkey agar it ranged from 87 to 97%. The predictive value of a resistant result was 80 to 97% for the four antimicrobial agents when MacConkey agar was used. MacConkey agar performed better than Mueller-Hinton agar because of the greater ease of detecting different bacterial morphotypes. The cost of direct stool screening with antibiotic-supplemented MacConkey agar is approximately half the cost of routine methods of surveillance. Its cost and accuracy make the method a useful adjunct to the routine management of neutropenic patients.
Detection of Cryptosporidium oocysts in sputum during screening for mycobacteria
Authors: Miller R A , Wasserheit J N , Kirihara J , Coyle M B
Published: January 2020

We report a case in which Cryptosporidium oocysts were detected during routine auramine screening of a sputum specimen for acid-fast bacilli. The patient was a 28-year-old male with acquired immune deficiency syndrome and intestinal cryptosporidiosis. Microbiology laboratory workers should be alert to the possibility that these oocysts may be present in sputum specimens from patients with acquired immune deficiency syndrome.
Direct identification and susceptibility testing of gram-negative bacilli from BACTEC bottles by use of the MS-2 system with updated bacterial identification software
Authors: Dipersio J R , Ficorilli S M , Varga F J
Published: January 2020

The Abbott MS-2 system (Abbott Laboratories, Diagnostic Division, Irving, Tex.), equipped with updated bacterial identification software (version 03.02), was used to perform both direct identification and susceptibility tests on gram-negative bacilli from positive BACTEC blood culture bottles. Ninety-eight of 101 Enterobacteriaceae strains, one strain of Acinetobacter calcoaceticus, and two strains of Pseudomonas aeruginosa were correctly identified by following a direct inoculation procedure. One Enterobacter sakazakii isolate was misidentified as Enterobacter cloacae. Ninety-seven percent of 936 direct susceptibility results were identical to the results obtained by using the standard MS-2 susceptibility procedure. Only five tests yielded a sensitive-direct and a resistant-standard MS-2 susceptibility result.
Rapid microimmunodiffusion method with species-specific antiserum raised to purified antigen for identification of Vibrio vulnificus
Authors: Nishibuchi M , Seidler R J
Published: January 2020

An antigen common to Vibrio vulnificus strains, designated VVA, was purified by ammonium sulfate precipitation, gel filtration, ion-exchange column chromatography, and preparative gel electrophoresis. The molecular weight of VVA was 64,000 when estimated by gel filtration and 40,000 when measured by denaturing polyacrylamide gel electrophoresis. Antiserum prepared against purified VVA (anti-VVA serum) did not agglutinate whole cells of V. vulnificus. Therefore, VVA was considered a possible internal antigen. By using anti-VVA serum, a microimmunodiffusion method was designed to detect the antigen VVA in bacterial cell lysates prepared from a single colony. This simple method allowed the specific identification of V. vulnificus as soon as 10 h after antigen preparation and therefore can be a useful tool in the identification of V. vulnificus from environmental or clinical specimens. VVA was not detected as a line of complete identity in some 20 other Vibrio species or in 7 other bacterial genera. VVA was present in all 63 isolates of V. vulnificus obtained from clinical and nonclinical sources.
Detection of Toxoplasma gondii antigens by a dot-immunobinding technique
Authors: Brooks R G , Sharma S D , Remington J S
Published: January 2020

A sensitive assay for the detection of antigens of Toxoplasma gondii by spotting samples directly onto nitrocellulose paper was developed. The sensitivity ranged from 10 to 40 pg of antigen diluted in phosphate-buffered saline and 40 to 130 pg of antigen diluted in normal mouse serum, normal human serum, or human cerebrospinal fluid. T. gondii antigen in serum samples taken from mice infected with T. gondii was detectable by day 2 of infection. Antigen was also detectable in cerebrospinal fluid samples taken from four of six infants congenitally infected with T. gondii and in serum samples from two of these infants.
Subclass distribution of rubella virus-specific immunoglobulin G
Authors: Linde G A
Published: January 2020

An enzyme-linked immunosorbent assay was used to study the subclass distribution of rubella virus-specific immunoglobulin G (IgG) in 97 serum samples from healthy donors and from patients with recent or remote rubella infections. Plastic beads coated with rubella antigen were incubated with test serum and then with monoclonal antibodies to the four human subclass of IgG. Rubella virus-specific IgG1 was present in all serum samples containing rubella virus-specific IgG antibodies. Rubella virus-specific IgG2 was present in 1 of 35 samples from healthy donors that also contained specific IgG1. Rubella virus-specific IgG3 was found in serum samples from patients with recent rubella infections but had disappeared by 6 months after the onset of symptoms. Rubella virus-specific IgG4 was found in low amounts in 7 of 35 samples from healthy immune donors. Of 20 serum samples that were negative by other serological techniques, 8 gave absorbances above cutoff levels in the assays for rubella virus-specific total IgG and IgG1. In 1 of 20 serum samples, the assays for total IgG and IgG2 were positive. High absorbance in the assay for rubella virus-specific IgG4 was found in one serum. This serum was negative in all other assays for rubella virus-specific antibodies.
Monoclonal and specific polyclonal antibodies for immunoassay of Clostridium difficile toxin A
Authors: Lyerly D M , Phelps C J , Wilkins T D
Published: January 2020

Monoclonal antibody, affinity-purified antibody, and monospecific antiserum against toxin A were produced. The monoclonal antibody was an immunoglobulin G2a kappa chain isotype that immunoprecipitated toxin A, as shown by crossed immunoelectrophoresis. These antibodies were compared by counterimmunoelectrophoresis, latex agglutination, and indirect enzyme-linked immunosorbent assay for their sensitivity in detecting toxin A. Our findings indicate that these antibodies may be useful as immunodiagnostic reagents for Clostridium difficile disease.
Comparison of enzyme-linked immunosorbent assay, hemagglutination inhibition, and passive latex agglutination for determination of rubella immune status
Authors: Steece R S , Talley M S , Skeels M R , Lanier G A
Published: January 2020

Hemagglutination inhibition, enzyme-linked immunosorbent assay (ELISA), and passive latex agglutination were used to test 495 human serum samples for determination of rubella immunity. Overall agreements for immune status were as follows: hemagglutination inhibition versus ELISA, 94.7% (469 of 495); hemagglutination inhibition versus passive latex agglutination (1:10 dilution), 99.2% (491 of 495); and passive latex agglutination (1:10 dilution) versus ELISA, 94.7% (469 of 495). Both ELISA and passive latex agglutination are satisfactory for rubella immunity screening, with the reservation that the ELISA method examined yielded a large number of false-negative results.
Western blot analysis of the human antibody response to Campylobacter jejuni cellular antigens during gastrointestinal infection
Authors: Nachamkin I , Hart A M
Published: January 2020

Western blot analysis was used to identify antigenic components of Campylobacter jejuni whole cells and outer membranes that elicit antibody responses in patients with campylobacter enteritis. Acute- and convalescent-phase sera from eight patients were analyzed for antibody activity against their homologous infecting strains and heterologous clinical isolates. Whole-cell and Sarkosyl-insoluble membrane components were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred electrophoretically to nitrocellulose paper for immunoblotting experiments. After the separated components were probed with patient sera, antibody binding was detected by autoradiography with 125I-protein A. Using this method, we detected several immunogenic components in whole cells and outer membranes. In the acute-phase response of some patients to infection, two to three components with approximate molecular weights of 66,000 (p66), 43,000 to 46,000 (major outer membrane protein), and 12,000 (p12) were detected in immunoblots. Convalescent-phase sera showed a more broad array of antibody binding to cell components. p66, shown to be campylobacter flagellin, was the major immunodominant component in almost all sera tested, however, p66 was not a major protein in Coomassie blue-stained gels. The major outer-membrane protein also bound to antibody, but with less intensity than p66. In general, the antibody specificity of patient sera was not limited to the homologous infecting strain, and antibodies cross-reacted with most components in heterologous strains. A low-molecular-weight component, identified as lipopolysaccharide with a modified silver stain, showed serological specificity for some patient sera. The results of this study showed that the antibody response of patients with campylobacter enteritis to C. jejuni antigens is variable. Flagellin appeared to be the major immunodominant component during infection.
Koserella trabulsii, a new genus and species of Enterobacteriaceae formerly known as Enteric Group 45
Authors: Hickman-Brenner F W , Huntley-Carter G P , Fanning G R , Brenner D J , Farmer J J
Published: January 2020

The name Koserella trabulsii is proposed for a group of Enterobacteriaceae formerly called Enteric Group 45. This group consists of 12 strains that were originally identified as atypical Hafnia alvei. K. trabulsii strains were negative for indole production, Voges-Proskauer, H2S production, urea hydrolysis, phenylalanine deaminase, and acid production from glycerol, lactose, sucrose, and D-sorbitol; they were positive for methyl red, citrate (Simmons), lysine and ornithine decarboxylases, arginine dihydrolase (negative in 1 to 2 days and positive in 3 to 7 days), and acid production from cellobiose and melibiose; and they were resistant to the Hafnia-specific bacteriophage of Guinée and Valkenburg. They were tested for DNA relatedness by the hydroxyapatite method with 32PO4-labeled DNA from the designated type strain (CDC 3349-72, ATCC 35313). The 12 strains were 87 to 99% related in 60 degrees C reactions. Relatedness of K. trabulsii to 71 DNA hybridization reference strains of representative species of Enterobacteriaceae was 4 to 37%. It was 15 to 16% related to H. alvei. All strains were susceptible to nalidixic acid, sulfadiazine, gentamicin, kanamycin, and chloramphenicol, and 83% were susceptible to nalidixic acid, sulfadiazine, gentamicin, kanamycin, and chloramphenicol, and 83% were susceptible to tetracycline. Most of the strains were resistant or intermediate to penicillin, ampicillin, carbenicillin, colistin, and cephalothin. Five of the strains were isolated from wounds, three were from the respiratory tract, and one each was from a stool, knee fluid, water, and an unknown source. The clinical significance of this organism is not known; therefore, future studies should focus on its isolation and its relationship to human disease.
Specific and cross-reacting antigens of Staphylococcus aureus of human and canine origins
Authors: Live I
Published: January 2020

Biotype -specificity of Staphylococcus aureus of human and canine origins has been found to be associated with thermolabile agglutinogens represented in S. aureus strains 17 and 61218, respectively. Both strains also have exhibited a common thermostable antigen. On that basis, absorbed antisera have been developed for the differentiation of S. aureus of the two biotypes. In the present study, still another thermostable agglutinogen was established, shared by strain 17 and some S. aureus strains of canine origin, as represented by S. aureus strain 887. These findings led to modification and enhanced specificity of the serological method of distinguishing S. aureus of the human biotype from S. aureus of the canine biotype.
Escherichia fergusonii and Enterobacter taylorae, two new species of Enterobacteriaceae isolated from clinical specimens
Authors: Farmer J J , Fanning G R , Davis B R , O'Hara C M , Riddle C , Hickman-Brenner F W , Asbury M A , Lowery V A , Brenner D J
Published: January 2020

Escherichia fergusonii (formerly known as Enteric Group 10) and Enterobacter taylorae (formerly known as Enteric Group 19) are proposed as new species in the family Enterobacteriaceae. By DNA hybridization (32P, 60 degrees C, hydroxyapatite), strains of E. fergusonii were 90 to 97% related to the type strain (holotype) ATCC 35469. They were most closely related to Escherichia coli and more distantly related to species in other genera. E. fergusonii strains are positive for indole production, methyl red, lysine decarboxylase, ornithine decarboxylase, and motility. They ferment D-glucose with gas production and also ferment adonitol, L-arabinose, L-rhamnose, maltose, D-xylose, trehalose, cellobiose, and D-arabitol. They are negative for Voges-Proskauer, citrate utilization (17% positive), urea hydrolysis, phenylalanine deamination, arginine dihydrolase, growth in KCN, and fermentation of lactose, sucrose, myo-inositol, D-sorbitol, raffinose, and alpha-methyl-D-glucoside. By DNA hybridization (32P, 60 degrees C, hydroxyapatite), strains of E. taylorae were 84 to 95% related to the type strain (holotype) ATCC 35317. Their nearest relative was E. cloacae, to which they were 61% related. Other named species were more distantly related. Strains of E. taylorae are positive for Voges-Proskauer, citrate utilization, arginine dihydrolase, ornithine decarboxylase, motility, growth in KCN medium, and malonate utilization. They ferment D-glucose with gas production and also ferment D-mannitol, L-arabinose, L-rhamnose, maltose, D-xylose, trehalose, and cellobiose. They are negative for indole production, methyl red, H2S production on triple sugar-iron agar, urea hydrolysis, phenylalanine deamination, lysine decarboxylase, gelatin hydrolysis, and fermentation of adonitol, i-inositol, D-sorbitol, and raffinose. Both new species occur in human clinical specimens. Two strains of E. fergusonii were isolated from blood. Five stains of E. taylorae were isolated from blood, and one was from spinal fluid. These blood and spinal fluid isolates suggest possible clinical significance, but this point requires further study.
Comparison of enzyme-linked immunosorbent assay with enzyme-linked fluorescence assay with automated readers for detection of rubella virus antibody and herpes simplex virus
Authors: Shekarchi I C , Sever J L , Nerurkar L , Fuccillo D
Published: January 2020

The enzyme-linked immunosorbent assay (ELISA) was compared with the enzyme-linked fluorescence assay (ELFA) for the detection of rubella antibody and herpes simplex virus antigen. Test parameters, specimens, antigen or antibody, and conjugates for the two types of assays were identical except that p-nitrophenyl phosphate was used as the substrate for the ELISA and 4-methylumbelliferyl phosphate was used as the substrate for ELFA. Automated readers were used for both assays. Antibody titers and sensitivity of antigen detection were quite similar for ELISA and ELFA. ELFA for rubella antibody, however, could be conducted with less antigen or shorter substrate incubation time (5 min for ELFA versus 30 min for ELISA). For herpes simplex virus antigen detection, ELFA could also be read after a shorter substrate incubation time (15 min for ELFA versus 30 min for ELISA). Clear polystyrene microtiter plates routinely used for ELISA could be used for ELFA, but clear polyvinyl chloride plates had high background fluorescence. Black polystyrene and polyvinyl chloride plates gave lower background fluorescence than did clear plates. ELFA is of particular value as a substitute for ELISAs in which long substrate incubations are required or antigens of only low titer are available.
Lack of utility of a Limulus amoebocyte lysate assay in the diagnosis of urethral discharges in men
Authors: Judson F N , Werness B A , Shahan M R
Published: January 2020

We evaluated a Limulus amoebocyte lysate assay (LALA) test kit for the diagnosis of gonorrhea in 883 unselected men with urethral discharge. Results were compared with those of Gram-stained smears and Martin-Lewis cultures. Of 331 men with gonococcal urethritis and 552 men with nongonoccal urethritis, 125 (37.8%) and 503 (91.1%), respectively, could not be evaluated by LALA owing either to insufficient discharge specimen to perform the test (569 or 64.4%) or to other exclusion criteria (59 or 6.7%). Of 255 LALA-evaluable discharges, LALA correctly diagnosed 252 (98.8%), compared with 244 (95.7%) for the Gram-stained smear. However, the Gram-stained smear also correctly diagnosed 96.5% of 456 men with insufficient discharge for LALA testing. The clinical utility of the LALA test kit is severely limited by performance criteria that exclude the majority of unselected men with urethritis. In addition, it is more technically cumbersome, time consuming, and costly than Gram-stained smears. Further test modifications are unlikely to overcome these inherent disadvantages of LALA.
Evaluation of the efficacy of reincubation and subsequent subculture of initially positive blood cultures in the detection of additional clinically significant isolates
Authors: Towne A R , Gay R M
Published: January 2020

Polymicrobial bacteremias are associated with higher mortality than are unimicrobial bacteremias, and their incidence appears to be increasing. Other researchers have recently shown that the prevalence of polymicrobial bacteremias may be underestimated when blood cultures are not evaluated further after identification of an initial isolate. We investigated this possibility by reincubating and further subculturing blood cultures initially positive for a single organism. We failed to show a clinically important increase in recovery of multiple isolates.
Comparison of methods for serotyping isolates of Haemophilus influenzae
Authors: Himmelreich C A , Barenkamp S J , Storch G A
Published: January 2020

This study compared four methods for serotyping isolates of Haemophilus influenzae. Slide agglutination with commercial antisera (Difco Laboratories and Wellcome Diagnostics), coagglutination (Phadebact Haemophilus Test [Pharmacia Fine Chemicals]), latex agglutination with affinity-purified anticapsular antibody, and counterimmunoelectrophoresis with multiple antisera were used to serotype 80 isolates of H. influenzae. Coagglutination and counterimmunoelectrophoresis correctly identified all 80 isolates as either type b or not type b. Slide agglutination and latex agglutination each successfully identified 76 of the 80 isolates; however, each of these two methods failed to type four isolates because of agglutination of controls. We recommend slide agglutination or coagglutination as the serotyping methods of choice in most laboratories because they are simple, accurate, and rapid. Slide agglutination with Difco antiserum can be performed at the lowest cost.
Enzyme-linked immunosorbent assay to measure antibodies to purified heat-labile enterotoxins from human and porcine strains of Escherichia coli and to cholera toxin: application in serodiagnosis and seroepidemiology
Authors: Levine M M , Young C R , Black R E , Takeda Y , Finkelstein R A
Published: January 2020

Serum immunoglobulin G antibodies to purified heat-labile enterotoxin (LT) from human (LTh) and porcine (LTp) Escherichia coli strains and cholera enterotoxin (CT) were measured by an enzyme-linked immunosorbent assay. Sera from patients with LTh E. coli infection showed a prominent response with LTh, an intermediate response with LTp, and a meager response with CT. Of 47 persons with clinical LTh-producing E. coli (herein shortened to LTh E. coli) infections, significant rises in antitoxin were detected against LTh in 36 (77%), against LTp in 30 (64%), and against CT in only 13 (28%) patients; seroconversions also occurred in 11 of 14 (79%) patients with subclinical LTh E. coli infections. In North Americans with experimental LTh E. coli infection, anti-Lth did not remain at high levels for more than 3 months. Persons with cholera manifested antitoxin responses that were similarly potent against all three toxin antigens; in fact, net optical density values were often slightly higher against LTh than against CT. The ratio of CT/LTh ELISA net optical density in convalescent sera proved to be a sensitive means to differentiate LT E. coli from cholera infection. All 11 cholera patients tested had CT/LTh ratios of greater than 0.70, whereas in only 1 of 47 LTh E. coli infections did the ratio exceed that value (it was 0.71) (P less than 0.0000000001). In single serum specimens, a net optical density of greater than or equal to 0.30 against LTh was shown to be a useful cutoff in screening sera for recent LTh E. coli or past cholera infection. The CT/LTh ratio was then used to differentiate definitively. Sera from healthy 3- to 5-year -olds and 15- to 19-year-olds in Maryland, Chile, and Bangladesh were tested against LTh and CT. The serological results fit known epidemiological observations. (i) LTh infections are rare in the United States (only 2 of 60 sera had LTh net optical density values of >/= 0.30. (ii) In contrast, evidence of recent LTh E. coli infections was very common in Chilean (69%) and Bangladeshi (57%) 3- to 5-year-olds and not uncommon in 15- to 19-year-olds (38 and 31%, respectively) in those countries. (iii) Only Bangladeshi sera showed serological evidence of cholera infections (CT/LTh ratios of > 0.70). The immunoglobulin G enzyme-linked immunosorbent assay measuring antibodies to purified LTh and CT represents a practical and effective tool for the serological study of LTh E. coli and cholera diarrheal infections.
Improvement of two toluidine blue O-mediated techniques for DNase detection
Authors: Waller J R , Hodel S L , Nuti R N
Published: January 2020

Two DNase detection techniques in which the metachromatic dye toluidine blue O (TBO) is used have been improved, and a potential source of difficulty for personnel attempting to use TBO-related methods has been identified. Reducing the concentration of TBO in the Streitfeld plate-flooding method from 0.1 to 0.05% resulted in easier control of staining intensity, less masking of DNase-positive reactions due to overstaining, sharper delineation of zones of DNase activity, and more sensitive detection of weak DNase reactions. Incorporation of 0.005% TBO in DNase agar, rather than the recommended 0.01%, allowed growth and expression of DNase activity by gram-positive as well as gram-negative bacteria. The reduced dye content in the agar also enhanced expression of DNase activity by some organisms and provided sharper delineation of DNase-positive reactions. Because optimum expression of DNase activity depends upon exact TBO concentrations in both the flooding and agar incorporation techniques, strict attention must be paid to the dye content of commercially available TBO dye powders. TBO concentrations must reflect actual dye content; therefore, calculations must include a conversion factor that accounts for the true dye content of the commercial preparation. The conversion factor that we developed is determined by dividing 100 by the percentage of dye in the commercial powder. The grams of commercial dye powder required per 100 ml of dye mixture is calculated by multiplying the percentage of dye required in the dye mixture by the conversion factor.
Radiometric method for testing susceptibility of mycobacteria to pyrazinamide in 7H12 broth
Authors: Heifets L B , Iseman M D
Published: January 2020

The test of susceptibility to pyrazinamide requires an acid environment (pH less than or equal to 5.5). This, however, is not favorable to the growth of Mycobacterium tuberculosis, especially in solid agar media. To obviate this difficulty, we developed a testing method with 7H12 broth medium and based on radiometric readings of the growth. The radiometric method employed in this study (BACTEC system) provides an opportunity to detect the dynamics of growth by daily recording of the growth index, which reflects the metabolic activity of the multiplying bacteria. In our technique, M. tuberculosis isolates were initially cultivated at pH 6.8. After logarithmic growth had begun, phosphoric acid solution was added to obtain pH 5.5. When pyrazinamide was added simultaneously with the acid, the growth index of susceptible cultures decreased, whereas it continued to increase in pH 5.5 control vials and in tests with pyrazinamide-resistant strains.
Effect of the source of Mueller-Hinton agar and resistance frequency on the detection of methicillin-resistant Staphylococcus aureus
Authors: Hindler J A , Inderlied C B
Published: January 2020

Inconsistencies in the results of disk diffusion tests of oxacillin against Staphylococcus aureus that occurred when using commercially prepared Mueller-Hinton agar from different sources led us to evaluate the ability of media from different sources to detect resistance to oxacillin, methicillin, and nafcillin in S. aureus. Mueller-Hinton agar from five manufacturers was prepared in our laboratory and used for standard disk diffusion and agar dilution tests. Ten oxacillin-resistant S. aureus isolates, of which three were definitive-resistant and seven were occult-resistant, were examined. All definitive-resistant strains were resistant to all three antimicrobial agents on four out of five agars. The occult-resistant strains were consistently detected as resistant on only one of the agars. With only slight differences, oxacillin, methicillin, and nafcillin resistance was more readily detected by disk diffusion and agar dilution when initially incubated at 30 degrees C, and extended incubation improved the detection. The frequency of resistance within a population of occult-resistant cells was low compared with the frequency within a population of definitively resistant cells. The heterogeneity of colony morphology and apparent growth rates within a population of occult-resistant cells contributed to the problem of detecting some resistant isolates. Definitive-resistant isolates were characterized by a very high and stable frequency of resistance. Occult-resistant strains were characterized by a lower frequency of resistance, although the true frequency of resistance may be difficult to ascertain because of heterogeneity in growth rates.
Illness associated with Campylobacter laridis, a newly recognized Campylobacter species
Authors: Tauxe R V , Patton C M , Edmonds P , Barrett T J , Brenner D J , Blake P A
Published: January 2020

Campylobacter laridis, a recently described thermophilic Campylobacter species found principally in seagulls, has not previously been linked to illness in humans. Six clinical isolates of this species were referred to the national campylobacter reference laboratory in 1982 and 1983. Each isolate was confirmed by biochemical characterization and by DNA relatedness studies. The six isolates were obtained during an illness: enteritis in four, severe crampy abdominal pain in one, and terminal bacteremia in an immunocompromised host in one. The infections occurred in persons 8 months to 71 years old. Neither the geographic distribution nor the reports of the patients suggest that seagulls played a direct role in the epidemiology of these infections. This potential human enteric pathogen appears to be clinically, epidemiologically, and microbiologically similar to Campylobacter jejuni and may be mistaken for it if nalidixic acid susceptibility screening is not routinely performed.
Effect of fibronectin on adherence of neutrophils from newborn infants
Authors: Harris M C , Levitt J , Douglas S D , Gerdes J S , Polin R A
Published: January 2020

Neutrophils (polymorphonuclear leukocytes [PMNs]) from infants and adults increased in adherence over a 20-min incubation as measured in a morphological assay. At each time point, PMNs from healthy infants had significantly diminished adherence when compared with those of adults; PMNs from sick newborn infants showed intermediate adherence values. Moreover, PMNs from healthy infants had significantly diminished membrane fluorescence for fibronectin when compared with those of adults. Preincubation with exogenous fibronectin did not augment the adherence of PMNs from infants or adults. Deficient adherence of PMNs from newborns may be related to decreased membrane fibronectin.
Isolation of Campylobacter fetus from a pet turtle
Authors: Harvey S , Greenwood J R
Published: January 2020

During the course of a Salmonella agona case investigation, Campylobacter fetus was isolated from a pet turtle. This is the first reported isolation of C. fetus from a turtle and suggests that turtles, in addition to being reservoirs for Salmonella species, may also be reservoirs for C. fetus.
Antibacterial activity in blood cultures
Authors: Rodriguez F , Lorian V
Published: January 2020

A total of 2,010 blood samples inoculated into tryptic soy broth were examined for antibacterial activity by means of a bioassay plate seeded with Bacillus subtilis spores. The size of the zone of inhibition on this plate was indicative of the degree of antibacterial activity. Current antibiotic therapy was confirmed by examination of chart records. Of the 2,010 blood cultures tested, 147 (7.3%) contained detectable levels of antibiotics; of these 147, 14 (9.5%) yielded growth of bacteria, and 133 (90.5%) remained negative. When the Antibiotic Removal Device (Marion Scientific, Div. Marion Laboratories, Inc., Kansas City, Mo.) was used, it eliminated the antibacterial activity but did not improve the recovery of bacteria from these cultures. Only bacteria resistant to the respective antibiotic were recovered from blood cultures that showed high levels of antibacterial activity (beta-lactam antibiotics, greater than 0.60 micrograms/ml; aminoglycosides, greater than 2 micrograms/ml; and tetracycline, greater than 4 micrograms/ml). Blood cultures showing low levels of antibacterial activity yielded both resistant and susceptible bacteria.
Isolation and enumeration of Clostridium botulinum by direct inoculation of infant fecal specimens on egg yolk agar and Clostridium botulinum isolation media
Authors: Glasby C , Hatheway C L
Published: January 2020

Direct plating of stool specimens on selective (Clostridium botulinum isolation) and nonselective (egg yolk agar) media was evaluated as an aid in confirming infant botulism. C. botulinum was isolated from 13 of 14 culture-positive specimens with C. botulinum isolation and from 8 of 14 egg yolk agar. No lipase reaction was seen on plates of 31 culture-negative specimens.