bluuebunny/crossref_metadata_2025 huggingface dataset:
Biotin-avidin-amplified enzyme immunoassay for detection of herpes simplex virus antigen in clinical specimens
Authors: Adler-Storthz K , Kendall C , Kennedy R C , Henkel R D , Dreesman G R
Published: January 2020

A biotin-avidin-amplified enzyme-linked immunosorbent assay (B-A ELISA) has been developed to detect herpes simplex virus type 1 (HSV-1) and HSV-2 antigens in clinical specimens. The test was designed as a solid-phase, double-antibody, sandwich assay in which plates were coated with a polyclonal rabbit immunoglobulin G anti-HSV reagent, and the sandwich antibody was a biotin-labeled mouse immunoglobulin M monoclonal antibody that reacts with a common antigen associated with HSV-1 and HSV-2. The test can be completed in 4 h if antibody-coated plates are available. The detection limit of the B-A ELISA, determined by titration of virus stocks, was found to be approximately 90 PFU or 6 X 10(3) physical particles of either HSV-1 or HSV-2 per 50 microliter of virus stock. The following results were obtained in a study in which swabs were taken from a variety of lesions and assayed for infectivity in tissue culture and by B-A ELISA. Of 421 suspected HSV lesions tested, 69 were positive by both tests and 159 were negative by both tests. A total of 122 were positive by B-A ELISA but negative for infectivity. Seventy-one were negative by B-A ELISA but contained infectious virus. The HSV specificity of the assay was substantiated by partial blocking of reactivity with rabbit immunoglobulin G anti-HSV and by the absence of reactivity with a nonspecific biotin-labeled mouse immunoglobulin M monoclonal antibody.
Conventional and radiometric drug susceptibility testing of Mycobacterium tuberculosis complex
Authors: Laszlo A , Gill P , Handzel V , Hodgkin M M , Helbecque D M
Published: January 2020

A recently developed method of drug susceptibility testing of Mycobacterium tuberculosis which measures the evolution of labeled CO2 from [1-14C]palmitic acid (BACTEC 460 system) was compared to three conventional methods. The proportion method of drug susceptibility testing was the standard against which all test results were compared. Indirect drug susceptibility to isoniazid, streptomycin, rifampin, and ethambutol of 245 isolates belonging to the M. tuberculosis complex was determined. In 95% of the cases, results obtained by the radiometric method were available within 1 week, as opposed to 3 to 6 weeks needed in conventional methodology. Overall agreement was 96.4%. Specificity values ranged from 0.98 to 1.0; sensitivity values of 1.0 for rifampin, 0.96 for streptomycin, 0.91 for isoniazid, and 0.18 for ethambutol were obtained. The specificity of the absolute concentration and resistance ratio drug susceptibility testing methods were 0.99 and 1.0, respectively. The sensitivity of the former was higher than that of the radiometric method (0.99 verus 0.92), whereas that of the latter was lower (0.88 verus 0.96). Further testing indicated that the low sensitivity determined for ethambutol may be due to the choice of the critical concentration used, rather than to a shortcoming of the procedure. The radiometric method thus does not significantly differ in reliability from conventional methods of drug susceptibility testing of M. tuberculosis.
Serum antibody responses in naturally occurring influenza A virus infection determined by enzyme-linked immunosorbent assay, hemagglutination inhibition, and complement fixation
Authors: Madore H P , Reichman R C , Dolin R
Published: January 2020

Serum antibody responses to influenza A virus infection were examined in 388 normal subjects during a trial of chemoprophylaxis in an outbreak of influenza A in 1980-1981 in which both A/H1N1 and A/H3N2 viruses circulated. Paired serum specimens obtained over a 6-week period were tested for antibodies to both A/H1N1 and A/H3N2 viruses by conventional hemagglutination inhibition, complement fixation, and an enzyme-linked immunosorbent assay (ELISA). Antibody responses detected by ELISA were determined by calculation of the area generated between titration curves of paired sera (area method), as well as by a conventional endpoint dilution method (endpoint method). Forty-two significant antibody rises were detected; 42 by ELISA (area method), 33 by ELISA (endpoint method), 32 by hemagglutination inhibition, and 13 by complement fixation. ELISA (area method) detected rises more frequently than either ELISA (endpoint method) (P less than 0.01), hemagglutination inhibition (P less than 0.005), or complement fixation (P less than 0.001). Another sensitive assay, the microneutralization test, detected significantly fewer rises (33, P less than 0.025) than the ELISA (area method). In the 42 subjects with ELISA (area method) rises, corroborating evidence of influenza A infection by other techniques (virus isolation, microneutralization, hemagglutination inhibition, or complement fixation tests) were available for 39 (93%). ELISA (area method) rises were subtype specific in all serum pairs in which other documentation of subtype-specific infection was available (38 of 38). Thus, ELISA (area method) was the single most sensitive assay for detection of serum antibody rises in this setting and possessed a high degree of subtype specificity.
Evaluation of the Phadebact CSF test for detection of the four most common causes of bacterial meningitis
Authors: Drow D L , Welch D F , Hensel D , Eisenach K , Long E , Slifkin M
Published: January 2020

A five-center collaborative study was undertaken to determine the suitability of the Phadebact CSF test kit and the Phadebact group B Streptococcus reagent for routine use by clinical laboratories to detect antigens of common organisms causing bacterial meningitis. The kits employ staphylococcal protein A coagglutination to detect the antigens of Haemophilus influenzae types a, b, c, d, e, and f, Neisseria meningitidis groups A, B, C, Y, and W135, Streptococcus pneumoniae (83 serotypes), and group B Streptococcus. A total of 2,817 individual tests were performed on 577 cerebrospinal fluid specimens. The percent positive specimens detected by coagglutination was as follows: overall, 84%; H. influenzae, 97%; group B Streptococcus, 75%; S. pneumoniae, 71%; and N. meningitidis, 58%. Eighty-five of the specimens were also tested by counterimmunoelectrophoresis. Coagglutination was more sensitive than counterimmunoelectrophoresis because it detected 74% of the positive specimens, whereas counterimmunoelectrophoresis detected only 65%. No false-positive results were obtained with coagglutination. The Phadebact CSF test kit is recommended for routine use in screening cerebrospinal fluid samples for antigens of the common organisms causing bacterial meningitis along with the Gram stain and culture for delayed confirmation of the rapid results.
Inguinal abscess caused by Rhizopus rhizopodiformis: successful treatment with surgery and amphotericin B
Authors: West B C , Kwon-Chung K J , King J W , Grafton W D , Rohr M S
Published: January 2020

Rhizopus rhizopodiformis has seldom been isolated from human mucormycosis. We report the first subcutaneous abscess to be caused by this fungus. It occurred in a diabetic man and presented as an inguinal mass, suggestive of a hernia, superficial to his cadaveric renal transplant. The fungus was readily isolated from pus inoculated onto blood and chocolate agars after a short incubation. The patient was cured by surgical drainage and treatment with 2.0 g of intravenous amphotericin B. Complete identification of such isolates is recommended.
Increased frequency of ColV plasmids and mannose-resistant hemagglutinating activity in an Escherichia coli K1 population
Authors: Agüero M E , Harrison H , Cabello F C
Published: January 2020

The expression of traits linked to pathogenicity was studied in a population of Escherichia coli K1 strains. It was found that E. coli K1 strains isolated from extraintestinal infection harbor the ColV plasmid and express mannose-resistant hemagglutinating activity type VI with a high frequency. The presence of these properties may play a role in the ability of some E. coli K1 serogroups to invade.
Plasmid profiles of clinical and environmental isolates of Legionella pneumophila serogroup 1
Authors: Maher W E , Plouffe J F , Para M F
Published: January 2020

Clinical and environmental Legionella pneumophila serogroup 1 isolates from a single water source in Columbus, Ohio, exhibited five different plasmid profiles. The multiplicity of plasmid profiles observed within a single geographic area and a skewed distribution of isolates bearing these plasmid profiles within this area suggest that plasmid analyses will be useful in the study of the epidemiology of Legionnaires disease and the environmental distribution of legionellae.
Enzyme-linked immunosorbent assay compared with neutralization tests for evaluation of live mumps vaccines
Authors: Sakata H , Hishiyama M , Sugiura A
Published: January 2020

Mumps-specific antibody levels before and after vaccination with live mumps vaccines were determined by enzyme-linked immunosorbent assay (ELISA) and neutralization tests. A correlation was found between neutralization titers and optical density in ELISA. However, postvaccination sera from some vaccinees who failed to seroconvert by neutralization contained significant levels of mumps-specific antibody detectable by ELISA. In some of these serum specimens, the antibody directed to the F polypeptide of mumps virus was predominant. Most sera positive in ELISA neutralized mumps virus upon the addition of fresh guinea pig serum to the virus-serum mixture.
Legionella bozemanii serogroup 2: a new etiological agent
Authors: Tang P W , Toma S , Moss C W , Steigerwalt A G , Cooligan T G , Brenner D J
Published: January 2020

A newly discovered bacterium, Toronto 3, isolated from a lung aspirate of a patient with pneumonia, has been characterized. The isolate was identified as Legionella bozemanii by chemical data from cellular fatty acid and ubiquinone analyses and by DNA relatedness studies. The isolate, however, differs phenotypically from L. bozemanii by its colonial characteristics and strong interspecies serological cross-reactions, which are unique among clinical isolates of legionellae. The name L. bozemanni serogroup 2 is proposed. The reference strain is Toronto 3. The clinical illness caused by L. bozemanii serogroup 2 was not distinguishable from other Legionella infections. L. bozemanii is the third Legionella species with more than one serogroup. Rapid laboratory diagnosis of this strain by direct fluorescent antibody test may be complicated in the absence of culture isolation.
Differentiation of Bacillus anthracis and other Bacillus species by lectins
Authors: Cole H B , Ezzell J W , Keller K F , Doyle R J
Published: January 2020

Bacillus anthracis was agglutinated by several lectins, including those from Griffonia simplicifolia, Glycine max, Abrus precatorius, and Ricinus communis. Some strains of Bacillus cereus var. mycoides (B. mycoides) were strongly reactive with the lectin from Helix pomatia and weakly reactive with the G. max lectin. The differential interactions between Bacillus species and lectins afforded a means of distinguishing B. anthracis from other bacilli. B. cereus strains exhibited heterogeneity with respect to agglutination patterns by lectins but could readily be differentiated from B. anthracis and the related B. mycoides. Spores of B. anthracis and B. mycoides retained lectin receptors, although the heating of spores or vegetative cells at 100 degrees C resulted in a decrease in their ability to be specifically agglutinated. Fluorescein-conjugated lectin of G. max stained vegetative cells of B. anthracis uniformly, suggesting that the distribution of lectin receptors was continuous over the entire cellular surface. B. anthracis cells grown under conditions to promote the production of capsular poly(D-glutamyl peptide) were also readily agglutinated by the lectins, suggesting that the lectin reactive sites penetrate the polypeptide layer. Trypsin, subtilisin, lysozyme, and mutanolysin did not modify the reactivity of B. anthracis with the G. max agglutinin, although the same enzymes markedly diminished the interaction between the lectin and B. mycoides. Because the lectins which interact with B. anthracis are specific for alpha-D-galactose or 2-acetamido-2-deoxy-alpha-D-galactose residues, it is likely that the bacteria possess cell surface polymers which contain these sugars. Lectins may prove useful in the laboratory identification of B. anthracis and possibly other pathogenic Bacillus species, such as B. cereus.
Susceptibility testing of filamentous fungi to amphotericin B by a rapid radiometric method
Authors: Merz W G , Fay D , Thumar B , Dixon D
Published: January 2020

A rapid, radiometric method was developed to determine the susceptibility of filamentous fungi to amphotericin B. The rapid, radiometric method depended on measurement of the inhibition of 14CO2 production in the presence of amphotericin B. Thirty isolates of filamentous fungi were tested by the rapid, radiometric method and a reference agar dilution method. There was 93% agreement between the two methods when an 80% or greater decrease in CO2 production was used to calculate the minimal inhibitory concentration with the rapid, radiometric method. Minimal inhibitory concentrations, based on 80% decrease of CO2 production, were achieved within 24 h of incubation with all of the fungi tested.
Enzyme immunoassay for diagnosis of gonorrhea
Authors: Schachter J , McCormack W M , Smith R F , Parks R M , Bailey R , Ohlin A C
Published: January 2020

An enzyme immunoassay (EIA; Gonozyme [Abbott Laboratories]) for gonococcal antigen was assessed for the rapid diagnosis of gonorrhea. Patients attending two sexually transmitted disease clinics were tested by EIA and culture on Thayer-Martin medium. EIA was highly effective in detecting gonococcal infection among symptomatic men, with 70 of 75 (93.3%) culture-positive men having positive tests and no false-positive reactions. The performance of the test was not as good in detecting cervical gonorrhea; the best result obtained was a sensitivity of 87% (33 of 38) for EIA compared with culture. EIA false-positives occurred at a relatively low rate for women, with the test having a specificity of ca. 97%. The test clearly is capable of detecting gonococcal antigen in cervical and urethral specimens, but its role in routine diagnosis is not clear. Its performance seems equal to that of the Gram stain for men, but it seems to be less sensitive than culture for cervical gonorrhea--a drawback in high-risk populations. The low false-positive rate could be an important issue in screening low-prevalence populations.
Influence of culture conditions on expression of the mucoid mode of growth of Pseudomonas aeruginosa
Authors: Chan R , Lam J S , Lam K , Costerton J W
Published: January 2020

Five strains of Pseudomonas aeruginosa that are routinely distinguished by diagnostic laboratories on the basis of their colony morphology on agar media were grown in different media to assess the effects of culture conditions on mucoid growth, which we define as the copious production of exopolysaccharide. On brain heart infusion agar, only two of these strains (mucoid and gelatinous) grew as slimy mucoid colonies. None of the five strains produced a mucoid pattern of growth in Mueller-Hinton broth, in which all grew as turbid, nonmucoid, homogeneous suspensions of bacterial cells. When Mueller-Hinton broth was supplemented with Mg2+, all of the strains produced some mucoid aggregated growth, but growth in a modified version of the chemically defined medium of Vogel and Bonner, with elevated levels of Mg2+ and gluconate, produced patently mucoid growth in all strains. This mucoid growth in a liquid medium takes the form of large, coherent, slimy aggregates within the medium and of a "collar" of adherent microcolonies at the air-medium interface. Direct observations by light and electron microscopy showed the submerged aggregates and the adherent microcolonies to be composed of bacterial cells enmeshed in a copious exopolysaccharide matrix. When agar was added to the supplemented medium of Vogel and Bonner, only the mucoid and gelatinous strains produced slimy mucoid colonies on its surface. We conclude that both chemical and physical factors affect exopolysaccharide production by these clinical strains of P. aeruginosa and that the colony morphology on a single agar medium is an insufficient criterion for the designation of a given isolate as being mucoid or nonmucoid.
Comparison of the suitability of three common bacterial media for susceptibility testing of trimethoprim-sulfamethoxazole
Authors: Ferguson R W , Weissfeld A S
Published: January 2020

The in vitro inhibitory activity of trimethoprim-sulfamethoxazole is inversely related to the amount of thymine and thymidine in the test medium; manufacturers must adequately control their media to avoid spurious antimicrobial susceptibility results. No differences were observed when commercial Mueller-Hinton broth and agar were compared with a semisynthetic broth medium by the use of microdilution and disk diffusion techniques.
Isolation of mucoid Vibrio parahaemolyticus strains
Authors: Lam S , Monteiro E
Published: January 2020

Mucoid strains of Vibrio parahaemolyticus were isolated from the stools of two asymptomatic carriers and a patient with gastroenteritis. The strains demonstrated biochemical reactions and antibiotic susceptibility typical of nonmucoid strains of V. parahaemolyticus isolated locally. The slime substance was typed by coagglutination and was antigenically similar to the capsular antigen of the same strain. Three different serotypes (O10:K24, O5:K17, and O5:K15) were involved.
Rapid detection of Clostridium perfringens type A enterotoxin by enzyme-linked immunosorbent assay
Authors: McClane B A , Strouse R J
Published: January 2020

Clostridium perfringens type A enterotoxin was specifically detected and readily quantified by indirect and four-layer sandwich enzyme-linked immunosorbent assays (ELISAs). With the indirect ELISA, enterotoxin was detected in quantities of as low as 2.5 ng (25 ng/ml). When the more sensitive sandwich ELISA procedures was used, 100 pg (1 ng/ml) of enterotoxin was detected. The sandwich ELISA procedure specifically detected enterotoxin in human fecal extracts. Additionally, the sandwich ELISA specifically differentiated enterotoxin-positive strains from enterotoxin-negative strains of C. perfringens. Both the indirect and sandwich ELISA procedures described for C. perfringens enterotoxin in this report are rapid, specific, sensitive, and easily adaptable for large-scale use by clinical or research laboratories.
Comparative studies and laboratory diagnosis of Vibrio vulnificus, an invasive Vibrio sp
Authors: Desmond E P , Janda J M , Adams F I , Bottone E J
Published: January 2020

Vibrio vulnificus was isolated from a bacteremic patient. This strain, together with other isolates of V. vulnificus, was compared with V. alginolyticus, V. fluvialis, and V. parahaemolyticus with regard to growth characteristics on enteric agar media (enabling isolation and identification) and production of exoenzymes which could correlate with invasive potential. V. vulnificus grew well on MacConkey. Endo, xylose-lysine deoxycholate, and Hektoen enteric agar plates. Because V. vulnificus colonies resembled those of lactose-fermenting strains of the family Enterobacteriaceae, however, isolation of this vibrio from mixed specimens or stools may require the use of thiosulfate-citrate-bile salts-sucrose agar. V. vulnificus produced numerous exoenzymes (protease, DNase, lipase, and esterase) but not elastase or lecithinase. Although differences in exoenzyme production were observed among the four vibrio species, no single exoenzyme could be linked to the invasive potential of V. vulnificus.
Clinical evaluation of the lysis-centrifugation blood culture system for the detection of fungemia and comparison with a conventional biphasic broth blood culture system
Authors: Bille J , Edson R S , Roberts G D
Published: January 2020

In a comparative fungal blood culture study, a lysis-centrifugation system (Isolator; Du Pont Co., Wilmington, Del.) detected 89% of all episodes of fungemia; the lysis-centrifugation system detected fungemia exclusively or significantly earlier than did a biphasic brain heart infusion bottle system 83% of the time. The lysis-centrifugation system was particularly useful in the early detection of fungemia caused by Candida tropicalis and Candida glabrata. In 53% of the clinically significant episodes, the earlier detection was directly helpful in the management of patients with fungemia. High-magnitude candidemia (greater than 5 CFU/ml of blood) was significantly associated with the presence of an infected intravascular catheter and with Candida species other than Candida albicans. The lysis-centrifugation system was sensitive in the detection of fungemia during the monitoring of patients receiving antifungal agents or after removal of an infected intravascular catheter.
Immunity to human cytomegalovirus measured and compared by complement fixation, indirect fluorescent-antibody, indirect hemagglutination, and enzyme-linked immunosorbent assays
Authors: Brandt J A , Kettering J D , Lewis J E
Published: January 2020

The complement fixation test is currently the test employed most frequently to determine the presence of antibody to human cytomegalovirus. Several other techniques have been adapted for this purpose. A comparison of cytomegalovirus antibody titers was made between the complement fixation test, a commercially available enzyme-linked immunosorbent assay, an indirect immunofluorescent technique, and a modified indirect hemagglutination test. Forty-three serum samples were tested for antibodies by each of the above procedures. The enzyme-linked immunosorbent, immunofluorescent, and indirect hemagglutination assays were in close agreement on all samples tested; the titers obtained with these methods were all equal to or greater than the complement fixation titer for 38 of the 41 samples (92.6%). Two samples were anticomplementary in the complement fixation test but gave readable results in the other tests. The complement fixation test was the least sensitive of the procedures examined. The commercial enzyme-linked immunosorbent assay system was the most practical method and offered the highest degree of sensitivity in detecting antibodies to cytomegalovirus.
Effects of overnight refrigeration on the microscopic evaluation of sputum
Authors: Penn R L , Silberman R
Published: January 2020

Microscopic evaluation of sputum permits selection of specimens suitable for culture, assessment of likely pathogens, and the best interpretation of culture results. We prospectively evaluated 50 sputum specimens which were promptly submitted to our clinical laboratory; smears and cultures were performed both immediately and after 20 h of refrigeration. Specimens were grouped according to the numbers of squamous epithelial cells and neutrophils per low-power field present on coded Gram-stained smears. The numbers of bacteria in five oil immersion fields were used to characterize smears for predominant, mixed, or scanty forms. After refrigeration, only three specimens changed group from a definite loss of squamous epithelial cells, and only two changed group from a definite loss of neutrophils. Based on cellular composition, the majority of samples would have been processed identically both before and after refrigeration. In contrast, organism forms detected on smears and their relative quantities were dramatically altered after refrigeration. A predominant smear form was gained in 11 and lost in 8 refrigerated specimens. The frequent changes on smears observed overall resulted from both increases and decreases in numbers of bacteria and yeasts. The majority of sputum culture results were insignificantly affected by the refrigeration of specimens. We conclude that 20 h of refrigeration renders sputum useless for the microscopic evaluation of potential pathogens and the subsequent interpretation of culture results. However, overnight refrigeration does not affect the determination from smears of sputum suitability for culture based on cellular composition.
Comparative evaluation of the API 20S system and the automicrobic system gram-positive identification card for species identification of streptococci
Authors: Appelbaum P C , Jacobs M R , Heald J I , Palko W M , Duffett A , Crist R , Naugle P A
Published: January 2020

Two commercial methods, the API 20S system (API; Analytab Products, Inc., Plainview, N.Y.) and the Gram-Positive Identification Card (GPI; Vitek Systems, Inc., Hazelwood, Mo.), were evaluated without additional tests for the identification of 241 streptococcus strains. Organisms included 60 beta-hemolytic strains, 36 group D strains, 26 Streptococcus pneumoniae strains, and 119 viridans streptococcus strains. API correctly identified to species 68.3% of beta-hemolytic strains, 86.1% of group D strains, 53.9% of S. pneumoniae strains, and 12.6% of viridans streptococci. This method provided excellent identification of group A and B and S. faecalis strains. Overall, API correctly identified 41.9% of strains to species, with 41.1% good likelihood but low selectivity, 15.8% incorrect, and 1.2% not identified. GPI correctly identified to species 58.3% of beta-hemolytic strains, 97.2% of group D strains, 80.8% of S. pneumoniae strains, and 57.2% of viridans streptococci. Group A, B, and D strains were all accurately identified by this system. Overall, GPI correctly identified to species 66.0% of strains, with 8.7% correct preliminary identification, 20.8% incorrect, and 4.6% not identified. Both methods represent a worthwhile advance in streptococcal identification. Neither system, however, can be recommended for species identification of the viridans group at this time.
Blood-free selective medium for isolation of Campylobacter jejuni from feces
Authors: Bolton F J , Hutchinson D N , Coates D
Published: January 2020

A blood-free selective agar is described which contains charcoal, ferrous sulfate, sodium pyruvate, casein hydrolysates, cefazolin, and sodium deoxycholate (CCD agar). CCD agar was compared with Preston medium for isolation of Campylobacter jejuni from human feces, and isolation rates were similar on both media, but CCD agar was less selective. Temperature studies at 37 and 42 degrees C confirmed that incubation of direct plates at 42 degrees C for 48 h was necessary for maximum isolation of C. jejuni.
Methylumbelliferyl-beta-D-glucuronide-based medium for rapid isolation and identification of Escherichia coli
Authors: Trepeta R W , Edberg S C
Published: January 2020

Escherichia coli is the most common gram-negative microbe isolated and identified in clinical microbiology laboratories. It can be identified within 1 h by oxidase, indole, lactose, and beta-glucuronidase tests. The oxidase and indole tests are performed as spot tests, and lactose fermentation is read directly from MacConkey agar. It was found that 4-methylumbelliferyl-beta-D-glucuronide could be incorporated directly into a modified MacConkey agar to directly detect the presence of beta-glucuronidase. Other characteristics of MacConkey agar were not affected. The incorporation of 4-methylumbelliferyl-beta-D-glucuronide into modified agar obviated the need for manufacture, quality control, and incubation of reagent-containing test tubes. The time needed to identify E. coli strains was reduced from 1 h to 5 min, and the ability to detect this species in mixed specimens was also enhanced.
Quantitation of bacteria in cerebrospinal fluid and blood of children with meningitis and its diagnostic significance
Authors: La Scolea L J , Dryja D
Published: January 2020

Cerebrospinal fluid (CSF) specimens from pediatric patients with meningitis were examined for their concentration of microbes and the relationship of this count to the bacteremia levels, microscopy results, and polymorphonuclear leukocyte concentration. A total of 2,031 consecutive CSF specimens were analyzed, of which 63 (3.1%) were positive by culture from the same number of patients. We observed that 85% of the total CSF specimens positive for Haemophilus influenzae type b, Streptococcus agalactiae, Streptococcus pneumoniae, and Neisseria meningitidis had counts in excess of 10(3) CFU/ml, with 56% of the specimens exceeding 10(5) CFU/ml. A correlation existed between the number of organisms present in the CSF and blood. For example, from a total of 22 patients who had counts of H. influenzae greater than 10(3) CFU/ml in the CSF, 16 or 73% had levels of bacteremia greater than 10(3) CFU/ml. It was also noted that the bacterial concentration had a profound effect on the sensitivity of microscopy. The percentage of positive results increased from 25% with less than or equal to 10(3) CFU/ml to 60% in the range of greater than 10(3) to 10(5) CFU/ml and to 97% at concentrations of greater than 10(5) CFU/ml. Furthermore, a significant correlation (P less than 0.01) was noted between the concentration of bacteria in the CSF and the number of polymorphonuclear leukocytes observed on microscopy.
Comparison of two commercially available test methods with conventional coagulase tests for identification of Staphylococcus aureus
Authors: Jungkind D L , Torhan N J , Corman K E , Bondi J M
Published: January 2020

The API STAPHase (Analytab Products, Inc., Plainview, N.Y.) and SeroSTAT Staph (Scott Laboratories, Fiskville, R.I.) tests were compared to the conventional tube coagulase test and a slide coagulase test by using fresh isolates of members of the family Micrococcaceae. The 4-h, 24-h, and combined readings of the tube coagulase test detected 94.5, 99.5 and 100%, respectively, of 219 Staphylococcus aureus isolates. The API STAPHase, SeroSTAT Staph, and slide coagulase tests detected 95.9, 95.4 and 95.9% of the isolates of S. aureus, respectively. There were no false-positive results with any of the systems when tested with 103 strains of members of the family Micrococcaceae other than S. aureus. We concluded that the STAPHase and SeroSTAT Staph tests were equal in accuracy to the slide coagulase and 4-h tube coagulase tests and were suitable for use in the clinical microbiology laboratory. However, SeroSTAT Staph gave faster results than the API STAPHase, and the test was easier to perform. Also, the false-negative rate was high enough with the STAPHase, SeroSTAT Staph, and the slide coagulase tests that all negative reactions should be confirmed with a tube test.
Cultivation and subgroup determination of human rotaviruses from Egyptian infants and young children
Authors: Naguib T , Wyatt R G , Mohieldin M S , Zaki A M , Imam I Z , DuPont H L
Published: January 2020

Primary African green monkey kidney cells were more sensitive than primary cynomolgus monkey kidney and MA104 cells for supporting the growth of human rotaviruses detected in diarrheal stools of Egyptian infants and young children. In attempts to characterize these Egyptian rotaviruses, only 31% of the strains tested in the form of fecal suspensions were identified as subgroup 1 or 2. After one passage in African green monkey kidney cells, 80% of the strains were identified as subgroup 1 or 2. Of these 43 rotaviruses for which the subgroup was determined, 28% were subgroup 1 and 72% were subgroup 2. Thus, cultivation in African green monkey kidney cell cultures facilitated the antigenic characterization of rotaviruses by subgrouping; cultivation also represents an initial step in determining serotype and in developing potential vaccine candidates.
Development and evaluation of a coculture technique for identification of anaerobic organisms
Authors: Hussain Z , Bürger H , Lannigan R , Groves D J
Published: January 2020

We investigated a method to reduce oxygen tension and produce anaerobic conditions conveniently by coculture with a nonfermentative organism. The biochemical reactions of 94 strains representing 41 different species were determined using prereduced media and gas chromatography according to standard methods. The standard reactions were compared with those obtained by preparing similar media, modified by containing no cysteine or resazurin, under aerobic conditions. Preincubation of the modified biochemical media with a standard suspension of a culture of Acinetobacter lwoffi provided reducing conditions. In 1,449 individual tests using 22 different biochemicals, 1,349 (93%) of the reactions agreed. Only two misidentifications resulted when using the modified media, one each at the species and genus levels. Two strains of Actinomyces odontolyticus failed to grow in coculture.
Detection of human immunoglobulins G and M antibodies to Rift Valley fever virus by enzyme-linked immunosorbent assay
Authors: Niklasson B , Peters C J , Grandien M , Wood O
Published: January 2020

Rift Valley fever virus (RVFV) is an important human and animal pathogen in Africa and has been responsible for infections in travelers. Because of the aerosol infectivity and risk of dissemination of the virus, a need exists for simple, safe, serological tests for diagnosis. An enzyme-linked immunosorbent assay (ELISA) was developed to detect RVFV-specific immunoglobulins (immunoglobulin G [IgG] and IgM). In the test, a betapropiolactone-inactivated, sucrose-acetone-extracted, suckling mouse liver RVFV antigen was captured by mouse RVFV antibodies adsorbed to polystyrene plates. The test sample (human serum) was then added, and the binding of specific antibodies was indicated by alkaline phosphatase-conjugated swine anti-human IgG or IgM. A mu-capture IgM ELISA was also developed by using polystyrene plates coated with goat anti-human IgM incubated successively with serum sample, RVFV antigen, and indicator antibodies. The ELISA for RVFV-specific IgG proved to be more sensitive than hemagglutination inhibition or complement fixation tests and almost as sensitive as the plaque reduction neutralization test in detecting specific antibodies in human sera after vaccination. The two ELISA IgM tests could detect specific IgM antibodies during the first 6 weeks after RVFV vaccination. Three injections of inactivated vaccine were given on days 0, 6 to 8, and 32 to 34. ELISA IgM values for sera obtained on days 6 to 8 were negative or in the lower range of significance, on days 32 to 34 they were strongly positive, and on days 42 to 52 they were waning. Later sera were negative. The plague reduction neutralization test was negative on days 6 to 8 but rose progressively in later samples. These findings suggest that the three doses of RVFV vaccine induce a prolonged primary antibody response. The ELISA IgM could become an important tool for early diagnosis in acute human infection. A number of African sera, some of which were positive for RVFV by plaque reduction neutralization test, were also tested by ELISA IgG. There was good agreement between both tests.
Herpes simplex virus type-selective enzyme-linked immunosorbent assay with Helix pomatia lectin-purified antigens
Authors: Svennerholm B , Olofsson S , Jeansson S , Vahlne A , Lycke E
Published: January 2020

Helix pomatia lectin-purified antigens with specific reactivity to herpes simplex virus type 1 (HSV-1) and HSV-2 antibodies in human sera were used in an enzyme-linked immunosorbent assay. The type specificity of the antigens was assessed by double immunodiffusion precipitation in gel against rabbit HSV-1 and HSV-2 hyperimmune sera, and by enzyme-linked immunosorbent assay with human reference sera containing antibodies to either type of HSV. Fifty-two sera from patients with documented infection with either HSV-1 or HSV-2 were assayed for HSV type-specific immunoglobulin G antibodies. The reactivity of the sera against lectin-purified antigens correlated completely with the results of virus typing. We conclude that HSV type-specific immunoglobulin G antibodies can easily be measured by enzyme-linked immunosorbent assay with the use of Helix pomatia lectin-purified HSV-1 and HSV-2 antigens.
Immunoglobulin A protease activity of Ureaplasma urealyticum
Authors: Robertson J A , Stemler M E , Stemke G W
Published: January 2020

All of 14 serotype standards and 34 of 35 wild-type strains of Ureaplasma urealyticum isolated from humans demonstrated an immunoglobulin A (IgA) protease activity. This activity degraded radiolabeled human IgA including IgA1 but not IgG or azocasein. The IgA fragments were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, followed by radioautography, and they had molecular weights of about 110,000 and 50,000. The IgA protease activity persisted in 25 mM EDTA but was sensitive to trypsin; it was presumed to be protein. This is the fourth microbial genus and the first myocoplasma species in which an IgA protease activity has been identified. Such activity was absent in Mycoplasma pneumoniae, Mycoplasma hominis, and Acholeplasma laidlawii.
Rapid serotyping of human rotavirus strains by solid-phase immune electron microscopy
Authors: Gerna G , Passarani N , Battaglía M , Percivalle E
Published: January 2020

Nine cell culture-adapted, as well as 30 clinical, human rotavirus (HRV) strains from fecal extracts of children with primary HRV infection were typed by rapid solid-phase immune electron microscopy with protein A and absorbed DS-1 (HRV serotype 2), Wa (serotype 1), and VA70 (assumed serotype 3) rabbit immune sera. As a reference typing test for cell culture-adapted strains, the neutralization assay was used, whereas for noncultivatable strains typing was done for comparison, indirectly, based upon the differential neutralization reactivity of convalescent-phase serum samples from patients with primary HRV infection versus the three reference HRV serotypes. Typing results by solid-phase immune electron microscopy for all strains examined were in complete agreement with those obtained by the neutralization assay, both on cell culture-adapted strains with the three reference rabbit antisera and on three reference HRV strains with human convalescent-phase serum samples. Since adaptation to growth in cell cultures of clinical HRV strains from stool specimens is a time-consuming procedure and is often unsuccessful, solid-phase immune electron microscopy is preferred over the neutralization assay, giving results in about 16 h and also allowing typing of HRV strains from stool specimens low in virus particles. In addition, HRV strains reacting differently from the three reference serotypes may be easily selected by solid-phase immune electron microscopy for further characterization, as was the case for one strain in this study.
Naturally acquired rabies in an eastern chipmunk (Tamias striatus)
Authors: Dowda H , DiSalvo A F
Published: January 2020

Rabies in an Eastern chipmunk was detected by fluorescent-antibody testing and mouse inoculation. The results were independently confirmed, and the virus was recovered from tissue culture.
Whole-cell peroxidase test for identification of Legionella pneumophila
Authors: Pine L , Hoffman P S , Malcolm G B , Benson R F , Gorman G W
Published: January 2020

A simple combined peroxidase-catalase test has been developed which is applicable to live bacterial cells. Known strains of Legionella pneumophila were differentiated from other species of Legionella by being peroxidase positive and catalase negative.
Improved preservation medium for Campylobacter jejuni
Authors: Nair G B , Chowdhury S , Das P , Pal S , Pal S C
Published: January 2020

An egg-based medium was found to be superior to the conventional Wang transport medium and the recently developed biphasic medium for the preservation of Campylobacter jejuni in the laboratory. Strains of C. jejuni preserved in egg-based medium maintained at 4 degrees C were viable for over 3 months. The survival of C. jejuni in egg-based medium held at room temperature (27 +/- 2 degrees C) was also relatively longer than in Wang transport medium and biphasic medium.
Evaluation of the Dynatech MR 580 microplate reader for routine antimicrobial susceptibility testing
Authors: Mazens M F , Bartlett R C
Published: January 2020

In microdilution antimicrobial susceptibility tests in our laboratory, only two concentrations of each antimicrobial agent are used. Plate reading with the Dynatech MR 580 Microplate Reader was compared with visual inspection. An Apple II computer printed results as S, I, or R. Reproducibility between visual and MR 580 readings was 99.4%.
Use of the Autobac IDX system for rapid identification of Enterobacteriaceae and nonfermentative gram-negative bacilli
Authors: Costigan W J , Hollick G E
Published: January 2020

The Autobac IDX system was evaluated for its ability to accurately identify 290 gram-negative bacilli from 18 different genera. Excluding isolates with a low identification probability, the overall sensitivity of the system was found to be 95.8%. Late lactose-fermenting Escherichia coli, Citrobacter freundii, and Proteus mirabilis accounted for over 90% of the misidentifications. The Autobac IDX system offers a rapid and reliable method for the identification of gram-negative bacilli.
Contamination of cultures processed with the isolator lysis-centrifugation blood culture tube
Authors: Thomson R B , Vanzo S J , Henry N K , Guenther K L , Washington J A
Published: January 2020

Overall contamination (on- plus off-streak) of the Isolator (Du Pont Co.) blood culture tube (23%) was greater than that of a conventional broth blood culture bottle (0.6%) or that of a biphasic blood culture bottle (1.3%). To determine the source of this contamination, Isolator cultures of blood from 59 healthy volunteers and of sterile broth from 60 vials were made. A total of 37% of the blood cultures and 22% of the broth cultures were contaminated (P = 0.06). Staphylococcus epidermidis-contaminated cultures represented 31 and 10% of the blood and broth cultures, respectively (P = 0.06). Contamination of plates processed on a bench top, in front of horizontal laminar flow, and in a biological safety cabinet with vertical laminar flow were compared. Processing plates in a biological safety cabinet resulted in a significant reduction in the number of contaminated plates (P less than 0.05). The contamination rate for 7,874 Isolator blood cultures processed in the biological safety cabinet was significantly decreased to 6.7% on-streak (9.3% on- plus off-streak). Contamination of Isolator-processed blood cultures originated from the laboratory and the patient. The former can be reduced by inoculating plates in a vertical laminar flow biological safety cabinet and by maintaining adequate quality control of media. The latter may be unavoidable.
Modification of interpretive breakpoints for netilmicin disk susceptibility tests with Pseudomonas aeruginosa
Authors: Barry A L , Miller G H , Hare R S , Jones R N , Thornsberry C
Published: January 2020

Regression analysis of data correlating 30-micrograms netilmicin disk zone diameters with microdilution MICs, obtained by testing close-interval dilution steps, was performed with 77 selected strains of Pseudomonas aeruginosa, each tested in three independent laboratories. A zone of greater than or equal to 15 mm correlated with an MIC of less than or equal to 12 micrograms/ml (susceptible), and a zone of less than or equal to 12 mm correlated with an MIC of greater than 16 micrograms/ml (resistant). Additional disk tests were performed with 256 strains having known resistance mechanisms and 280 susceptible strains: the majority were appropriately categorized by these interpretive zone standards. The previously recommended standards of greater than or equal to 17 mm (MIC, less than or equal to 8.0 micrograms/ml) for the susceptible category inappropriately placed a significant number of truly susceptible P. aeruginosa strains in the intermediate category.
Comparison of the Roche Septi-Chek blood culture bottle with a brain heart infusion biphasic medium bottle and with a tryptic soy broth bottle
Authors: Henry N K , Grewell C M , McLimans C A , Washington J A
Published: January 2020

In a comparison of 1,368 positive blood cultures, a vented Roche Septi-Chek (V-RSC) blood culture bottle was superior to an unvented tryptic soy broth-containing bottle (Difco) for the recovery of all aerobic and facultatively anaerobic microorganisms. Anaerobic bacteria were recovered more frequently and earlier in the unvented tryptic soy broth-containing bottle. A separate comparison of 529 positive blood cultures was conducted to examine the performance of the V-RSC bottle with that of a vented brain heart infusion biphasic medium. The V-RSC bottle recovered significantly more isolates of Enterobacteriaceae and of anaerobic bacteria than did the vented brain heart infusion biphasic medium. The V-RSC bottle is a reliable blood culture system for all aerobic and facultatively anaerobic microorganisms. Because of its suboptimal recovery of anaerobic bacteria, it is recommended that the V-RSC bottle be used in combination with an unvented vacuum blood culture bottle.
Preparing inoculum for susceptibility testing of anaerobes
Authors: Swenson J M , Thornsberry C
Published: January 2020

We studied six methods for preparation of inoculum to use in susceptibility tests of anaerobic bacteria to determine viable counts of 15 different species of anaerobes. We counted viable bacteria for each method-organism combination. Methods studied included those used for the more routine tests (broth microdilution and broth-disk elution) and for the National Committee for Clinical Laboratory Standards standard reference agar method. Inocula prepared from broth suspensions of organisms from 24- and 48-h anaerobe blood agar plates and adjusted to the turbidity of a 0.5 McFarland standard gave the most consistent counts for all organisms and also the highest numbers. Counts from these suspensions were higher than those from overnight growth in thioglycolate or Schaedler broth when all were adjusted against the same turbidity standard. Preparing inoculum directly from agar plates may also speed up the sometimes lengthy process of susceptibility testing of anaerobes and thus make results more clinically useful.
Investigation of a Campylobacter jejuni outbreak by serotyping and chromosomal restriction endonuclease analysis
Authors: Bradbury W C , Pearson A D , Marko M A , Congi R V , Penner J L
Published: January 2020

Fifty Campylobacter jejuni isolates, including 29 from humans associated with an outbreak of enteritis, 20 from cattle, and 1 from a milk source, were serotyped on the basis of extractable thermostable antigens and examined by bacterial chromosomal restriction endonuclease digest analysis. Serotyping showed specific differences between the human isolates and the milk isolates, but each of these generally, although not consistently, reacted with 4 of the 42 C. jejuni typing antisera. Restriction patterns of all of the human isolates and some of the cattle isolates were indistinguishable, confirming the suspected link between the cattle and the human outbreak. The single milk isolate had a restriction pattern unlike those of the human isolates, and therefore its involvement in the transmission of infection from the cattle to the humans could not be confirmed.
Enzyme-linked immunosorbent assay for detection of equine infectious anemia virus p26 antigen and antibody
Authors: Shane B S , Issel C J , Montelaro R C
Published: January 2020

A sensitive specific enzyme-linked immunosorbent assay utilizing purified p26 antigen was developed for the detection of antibodies to equine infectious anemia virus in naturally and experimentally infected horses. Generally, antibodies to the virus could be detected by the enzyme-linked immunosorbent assay 3 to 4 days earlier than by the standard agar gel immunodiffusion test, and they could be detected more reliably in horses with weak or equivocal agar gel immunodiffusion test reactions. The enzyme-linked immunosorbent assay was also successfully applied to the detection of p26 antigen in tissue culture fluids.