bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification of an antigen common to different species of the genus Campylobacter
Authors: Mills S D , Kurjanczyk L A , Penner J L
Published: January 2020

Affinity-purified antibody specific for a determinant on flagellin from Campylobacter jejuni was used to screen by immunoblotting strains of C. coli, C. laridis, C. fetus, "C. upsaliensis," C. pylori, and C. sputorum biovar fecalis. The antigen was detected in each of these species, but the molecular weights of the proteins bearing the common antigen varied considerably.
Influence of culture medium pH and proteinase inhibitors on extracellular proteinase activity and cell growth of Sporothrix schenckii
Authors: Tsuboi R , Sanada T , Ogawa H
Published: January 2020

Sporothrix schenckii produces two extracellular proteinases in albumin- or collagen-supplemented unbuffered liquid medium. Proteinase I had an optimal pH of 6.0, and its activity was strongly inhibited by chymostatin. Proteinase II had an optimal pH of 3.5, and its activity was strongly inhibited by pepstatin. Speculating that these two proteinases are key enzymes for fungal growth, we investigated the influences of culture medium pH and either or both of the proteinase inhibitors pepstatin and chymostatin on the cell growth of S. schenckii. In buffered medium at a pH of 3.5, an optimal pH for proteinase II, only proteinase II activities were observed, while at pH 6.0, an optimal pH for proteinase I, only proteinase I activities were observed. However, there was no cell growth inhibition. These results suggested that the regulation of the production of the two proteinases is dependent on environmental pH. The addition of pepstatin or chymostatin to the culture medium did not inhibit the cell growth of S. schenckii, but the addition of both inhibitors at a concentration of 10 micrograms/ml strongly inhibited growth. These results suggested that at least one of the two proteinases was expressed to allow fungal growth in albumin-supplemented media. The indirect fungistatic action of the proteinase inhibitors, which inhibit proteolysis, may be applied to a topical therapeutic agent in vivo.
Multicenter evaluation of four methods of yeast inoculum preparation
Authors: Pfaller M A , Burmeister L , Bartlett M S , Rinaldi M G
Published: January 2020

We initiated a comparative study of four methods of yeast inoculum preparation: a spectrophotometric method, the Wickerham card method, a hemacytometer method, and the Prompt inoculation system. The variability in inoculum size obtained when each method was applied to two strains each of Candida albicans, Candida tropicalis, Candida parapsilosis, Torulopsis glabrata, Cryptococcus neoformans, and Saccharomyces cerevisiae was analyzed in a single laboratory. Each method was performed in triplicate on the same day and on three separate days to provide estimates of within-day and between-day variations. Inoculum size was determined by viable colony counts. The greatest range of inoculum sizes was seen with the Wickerham card method. Viable counts ranged from 1.1 X 10(6) to 24.2 X 10(6) CFU/ml among the 12 yeast isolates. The greatest variation was observed with the Prompt system. Within-day coefficients of variation averaged 19% (range, 4 to 45%), and between-day coefficients of variation averaged 22% (range, 3 to 51%). Variation between laboratories was evaluated by comparing inoculum values obtained by each method in three different laboratories for two strains of C. albicans. The spectrophotometric method was the least variable and the Wickerham card and hemacytometer methods were the most variable methods between laboratories. The spectrophotometric method is recommended as the method of choice for preparation of a standardized inoculum suspension for susceptibility testing of yeasts.
False-positive DNA probe test for Legionella species associated with a cluster of respiratory illnesses
Authors: Laussucq S , Schuster D , Alexander W J , Thacker W L , Wilkinson H W , Spika J S
Published: January 2020

Between 11 November 1986 and 28 February 1987, legionellosis was diagnosed in 23 patients at one hospital with a recently marketed Legionella-specific DNA probe for respiratory secretions. Only 10 of the 23 probe-positive patients showed findings typical of Legionella pneumonia, including a temperature of greater than or equal to 100.5 degrees F (approximately 38.1 degrees C) and radiographic evidence of pneumonia. No differences were found in the results of laboratory studies, demographic features, or underlying risk factors for these 10 probe-positive patients when compared with the 13 probe-positive patients with nonpneumonic illnesses. A case-control study comparing probe-positive and -negative patients failed to identify any different features of disease or epidemiologic characteristics. Probes of repeat specimens of sputum were still positive 2 to 13 weeks after the initial test in 5 (50%) of the 10 probe-positive patients. The clinical features in most patients were atypical for legionellosis, and the diagnosis could not be confirmed by traditional laboratory tests performed on duplicate specimens processed at the Centers for Disease Control. This report emphasizes the need for clinical microbiology laboratories to confirm test results from new procedures by accepted diagnostic methods.
Multiple Candida strains in the course of a single systemic infection
Authors: Soll D R , Staebell M , Langtimm C , Pfaller M , Hicks J , Rao T V
Published: January 2020

Species and strain variabilities have been monitored during the history of a prolonged Candida infection in a single compromised bone marrow transplant patient by analyzing sugar assimilation patterns, high-frequency switching repertoires, and Southern blot hybridization patterns with two cloned mid-repeat sequences (Ca3 and Ca7) which are species specific for Candida albicans and one cloned mid-repeat sequence (Ct13-8) which is species specific for Candida tropicalis. Evidence is presented that during the course of this infection (i) two strains of C. albicans and three strains of C. tropicalis were distinguished by their switching repertoires, Southern blot hybridization patterns, and sugar assimilation patterns; (ii) the three C. tropicalis strains were in a high-frequency mode of switching; (iii) two C. tropicalis strains coexisted in the blood and three C. tropicalis strains coexisted in the throat at different times during the history of the infection; (iv) amphotericin B treatment selectively removed one of two C. tropicalis strains coexisting in the blood and this strain exhibited greater susceptibility to amphotericin B in vitro (the remaining strain was subsequently removed from the blood by flucytosine treatment); and (v) both the strain removed from the blood by amphotericin B and the strain removed from the blood by flucytosine reappeared several days later at another site of infection. It is demonstrated for the first time that C. tropicalis is capable of high-frequency switching of colony morphology just as C. albicans is, that there is more than one strain-specific switching repertoire in C. tropicalis, and that a C. tropicalis mid-repeat sequence can be used for discriminating species and assessing strain relatedness, as previously demonstrated for C. albicans mid-repeat sequences.
Novel application of video image processing to biochemical and antimicrobial susceptibility testing
Authors: D'Amato R F , Isenberg H D , McKinley G A , Baron E J , Tepper R , Shulman M
Published: January 2020

ALADIN (Analytab Products, Plainview, N.Y.) is an automated instrument that uses video imaging (computer-assisted guided video camera) for the determination of biochemical and antimicrobial susceptibility test reactions. This collaborative investigation compared video-generated results obtained with ALADIN with visually determined findings. Both approaches were used to view identical reactions. Overall agreement for biochemical and antimicrobial susceptibility tests was greater than 95%. This study demonstrates that video imaging is an acceptable approach for determining microbial responses to biochemical and antimicrobial agents and may provide, with appropriate computer modifications, more accurate and reproducible results than are possible by visual scrutiny.
Antibody responses to serogroup B meningococcal outer membrane antigens after vaccination and infection
Authors: Rosenqvist E , Harthug S , Frøholm L O , Høiby E A , Bøvre K , Zollinger W D
Published: January 2020

Antibody responses of adult volunteers given a vaccine containing meningococcal capsular polysaccharides (serogroups A, C, Y, and W-135) noncovalently complexed with serotype 2b:P1.2 and 15:P1.16 outer membrane proteins have been studied. Sera were analyzed by enzyme-linked immunosorbent assay methods for immunoglobulin G (IgG), IgM, and IgA antibodies and for bactericidal activities against the homologous strains. The vaccination was performed as a double-blind experiment with 47 volunteers, of whom 23 received the protein-polysaccharide vaccine and 24 received the control preparation containing the polysaccharides only. Ten additional persons volunteered for the protein-polysaccharide vaccine. Before vaccination, carriers of meningococci had significantly higher levels of specific IgG and IgA and also higher bactericidal activities than noncarriers. At 2 weeks postvaccination we found significant IgG and bactericidal antibody responses against both the 2b:P1.2 and 15:P1.16 strains in about 70% of the protein-polysaccharide vaccinees. The immune response induced by disease was compared with that induced by vaccination by analyzing paired sera from 13 survivors of serogroup B serotype 15 meningococcal disease. We found that the mean specific IgG level in acute-phase sera was lower than average in prevaccination sera from the vaccinees but similar to that of healthy noncarriers before vaccination. The convalescent-phase sera showed IgG responses similar to those of the vaccinees, but the IgM response to disease was significantly higher than after vaccination. The immune response to disease caused by serogroup B serotype 15 meningococci was found by enzyme-linked immunosorbent assay analysis to be about the same with outer-membrane antigens from a serotype 2b strain as it was with antigens from a serotype 15 strain.
Human-isotype-specific enzyme immunoassay for antibodies to pneumococcal polysaccharides
Authors: Shyamala G N , Roberton D M , Hosking C S
Published: January 2020

A simple enzyme immunoassay has been developed to allow the quantitation of the human response to immunization with pneumococcal polysaccharide. The assay uses the 14-valent vaccine (Pneumovax) as a convenient antigen to adsorb to the solid-phase microdilution plate wells and commercially available isotype-specific antibody conjugates. The results have been expressed as arbitrary pneumococcal polysaccharide antibody units by reading off a standard curve constructed by using heterogeneous pooled serum. All nonimmunized subjects tested had immunoglobulin G (IgG) antibodies present in serum. All six control subjects who were immunized with Pneumovax demonstrated an IgG response, and the majority responded with a rise in IgA- and IgM-specific antibody concentrations at a mean of 6 weeks postimmunization. Five out of six cord sera tested contained IgG antibodies only, which were present in concentrations similar to those seen in adults, whereas in 6- to 12-month-old infants only low levels of IgG and IgM and no IgA antibodies were detected. Serum taken from 10 hypogammaglobulinemic patients immediately prior to infusion of immunoglobulin showed low to negative IgG antibody concentrations, and no IgA or IgM antibody was present.
Evaluation of a new latex test and a new enzyme immunoassay for determination of rubella immunity
Authors: Simor A E , Chua R , Low D E
Published: January 2020

A new enzyme immunoassay (Rubenostika; Organon Teknika, Turnhout, Belgium), a new latex agglutination test (Rubalex; Orion Diagnostica, Espoo, Finland), and three other accepted methods for the determination of rubella immunity were compared with a standard hemagglutination inhibition assay. Of 224 serum samples tested, 54 (24%) were nonreactive and 24 (11%) were low titered. All procedures were very specific (94 to 100%). Rubenostika was the least sensitive method (88%), and Rubalex was the most sensitive (98%).
Fatal infection caused by a multiply resistant type 3 pneumococcus
Authors: Lawrenson J B , Klugman K P , Eidelman J I , Wasas A , Miller S D , Lipman J
Published: January 2020

The most virulent pneumococcal serotype (type 3) has not to date been associated with multiple antimicrobial resistance. We report an unusual gastrointestinal presentation of fatal septicemia caused by a multiply resistant type 3 pneumococcus in a setting of increasing prevalence of multiple resistance, including resistance to erythromycin, clindamycin, and tetracycline.
Cellular fatty acid composition of Kingella species, Cardiobacterium hominis, and Eikenella corrodens
Authors: Wallace P L , Hollis D G , Weaver R E , Moss C W
Published: January 2020

We determined the cellular fatty acid composition of reference strains and clinical isolates of each of the three Kingella species, Cardiobacterium hominis, and Eikenella corrodens by using capillary gas chromatography. Kingella denitrificans and Kingella kingae contained myristic (14:0) and palmitic (16:0) acids as major acids, whereas cis-vaccenic (18:1 omega 7c) and palmitic acids were the major acids in Kingella indologenes, C. hominis, and E. corrodens. C. hominis differed from the other four species by the absence of 3-hydroxylauric (3-OH-12:0) acid, from K. indologenes by the presence of 3-hydroxypalmitic (3-OH-16:0) acid, and from E. corrodens by the presence of 3-hydroxymyristic (3-OH-14:0) acid. E. corrodens contained a small amount (2%) of myristic acid, while the other four species contained moderate to large amounts (11 to 31%) of this acid.
Mannose-resistant hemagglutination of human erythrocytes by enterotoxigenic Escherichia coli with colonization factor antigen II
Authors: Evans D J , Evans D G , Diaz S R , Graham D Y
Published: January 2020

The human erythrocyte receptor which mediates mannose-resistant hemagglutination by enterotoxigenic Escherichia coli possessing colonization factor antigen II is not universally distributed among donors. Mannose-resistant hemagglutination-positive erythrocytes are more common among black donors than nonblack donors; tests with erythrocytes of known antigenic makeup confirm this correlation. Colonization factor antigen II receptor activity of mannose-resistant hemagglutination-positive erythrocytes is unstable when whole blood is stored at 4 degrees C. Also, screening of donors is best performed with enterotoxigenic E. coli possessing colonization factor antigen II composed of the coli surface antigen 1 (CS1) plus CS3, since these consistently produce stronger hemagglutination reactions than strains with colonization factor antigen II composed of either CS2 plus CS3 or CS3 only.
Characterization of Candida isolates from pediatric burn patients
Authors: Neely A N , Odds F C , Basatia B K , Holder I A
Published: January 2020

To provide more detailed information about Candida epidemiology and pathogenesis in pediatric burn patients, Candida isolates from 113 patients collected over 3 years were identified at the species level and the serotypes and biotypes of the C. albicans isolates were determined. A total of 85% of the patients were colonized or infected by C. albicans, 18% by C. tropicalis, and 11% by C. parapsilosis. Although colonization or infection often was found at multiple sites and times, 87% of the patients were colonized or infected by only one Candida species or strain; the other 13% showed multiple colonizations or infections, some of which occurred simultaneously at the same site. C. albicans biotyping determined the tolerance of the isolates to pH (pH 1.4) and salt; flucytosine, borate, and safranine resistance; and ability to produce proteinase and assimilate urea, sorbose, and citrate; results are expressed as three-digit numbers. For isolates from three different anatomical sites, the distribution of the nine biotype characteristics was similar in all cases but one. Significantly more fecal than wound or throat isolates were resistant to safranine. Sixty-four different serotype-biotype combinations were found in the 96 patients with C. albicans infections or colonizations. Twenty-nine percent of all C. albicans isolates had the partial biotype -57, while 20 of the 96 patients had specifically serotype B, biotype 557 colonizations or infections. Eleven patients had the B557 infection when admitted; nine patients acquired the yeast in-house. Thirty percent of the C. albicans isolated from 23 adult patients at a nearby hospital also showed the -57 biotype pattern, suggesting that C. albicans isolates expressing this biotype are either extremely prevalent in nature or are more virulent than other C. albicans isolates.
In vitro quantitative model of catheter infection during simulated parenteral nutrition
Authors: Merlino R , Gaillard J L , Fauchere J L , Chaumont P , Droy-Lefaix M T , Descamps P , Ricour C , Veron M
Published: January 2020

We developed a quantitative in vitro model of catheter infection. Colonization was initiated by inoculating the catheter lumen with a small number of bacteria (approximately 5 x 10(3) viable organisms). Then the inoculated catheters were used for simulated total parenteral nutrition therapy consisting of infusions for 9 h a day, and bacteria were counted in the effluent fluid against time, enabling us to monitor catheter colonization quantitatively. Bacterial colonization of prosthetic devices is a progressive process, as evidenced by the slow day-to-day increase of bacterial release seen here. On the other hand, bacterial strains of various representative species exhibited significant differences in their ability to infect catheters. These results suggest that the in vitro model presented here is a reliable tool for monitoring the degree of catheter colonization under standardized conditions and could be used for further studying the main factors of catheter-related sepsis or the treatment of this information.
Simple genetic method to identify viridans group streptococci by colorimetric dot hybridization and fluorometric hybridization in microdilution wells
Authors: Ezaki T , Hashimoto Y , Takeuchi N , Yamamoto H , Liu S L , Miura H , Matsui K , Yabuuchi E
Published: January 2020

Simple dot hybridization and fluorometric hybridization methods in microdilution wells were designed and established for rapid and routine genetic identification of viridans group streptococci. Reference DNA extracted from each strain of 24 reference Streptococcus species was fixed both on a nitrocellulose filter and in a microdilution well. A 1-ml portion of the bacterial suspension which matched the turbidity of McFarland no. 2 standard was prepared when a streptococcal strain was isolated. It was lysed with achromopeptidase, and the DNA was quickly labeled with photobiotin under a sunlamp for 15 min. Dot hybridization and fluorometric hybridization were then carried out between the labeled DNA of the unknown organism and 24 unlabeled reference DNAs. Hybridized fragments on a nitrocellulose filter were detected by using alkaline-phosphatase-conjugated streptavidin and analyzed with a color graphic analyzer. Hybridized fragments in microdilution wells were quantitatively detected by using an enzyme, streptavidin-conjugated beta-D-galactosidase, and a fluorogenic substrate, 4-methylumbelliferyl-beta-D-galactoside. Strains belonging to each genetically distinct species could be identified by this dot blot hybridization test. However, some clinical strains cross-hybridized with two or more reference species, and then they were difficult to differentiate by dot blot hybridization. In such a case, fluorometric identification provided reliable results because the fluorometric method was more quantitative than dot blot identification. By these methods, it was possible to determine species assignment within the viridans group.
Humoral immune response in human tuberculosis: immunoglobulins G, A, and M directed against the purified P32 protein antigen of Mycobacterium bovis bacillus Calmette-Guérin
Authors: Turneer M , Van Vooren J P , De Bruyn J , Serruys E , Dierckx P , Yernault J C
Published: January 2020

The P32 protein antigen of Mycobacterium bovis BCG, identified as antigen 85A in the BCG reference system, was used to investigate the humoral immune response in human tuberculosis (TB). Immunoglobulin G (IgG), IgA, and IgM directed against P32 were measured by an enzyme-linked immunosorbent assay. Mean IgG and IgA antibody levels differed significantly (P less than 0.001) between active-TB patients (50 untreated and 52 treated) and healthy control subjects (111 unvaccinated tuberculin negative, 38 unvaccinated tuberculin positive, and 72 recently BCG vaccinated). Mean IgG antibody levels, but not mean IgA antibody levels, were higher (P less than 0.05) in patients with positive microscopic examination for acid-fast bacilli than in patients with negative microscopic examination. A positive relation was found between mean levels and the extent of disease. There was no difference in mean IgM antibody levels between patients and controls. By setting the upper normal limit at the 95th percentile of the 221 healthy subjects, the sensitivities were 46% in untreated and 63% in treated patients for IgG and 30 and 50%, respectively, for IgA. Of the untreated patients, 56% were positive for either IgG or IgA antibodies. Among the untreated patients with negative direct smear, 35% were positive for IgG and 24% were positive for IgA. When both immunoglobulin classes were combined, the serological test was positive in 47% of those patients. Neither naturally acquired tuberculin hypersensitivity nor BCG vaccination affected positivity frequencies in healthy subjects. Only active TB seemed to induce significant anti-P32 antibody levels and to be associated with positivity. A serological test with P32 as the antigen might therefore be helpful for the rapid diagnosis of TB.
Candida albicans- and Candida stellatoidea-specific DNA fragment
Authors: Cutler J E , Glee P M , Horn H L
Published: January 2020

DNA was isolated from whole cells of Candida albicans and digested with MspI restriction enzyme. In addition to the expected large number of low-molecular-weight DNA pieces resulting from the digestion, multiple high-molecular-weight (greater than 3.0 kilobase pairs) fragments were generated by this enzyme, which cleaves DNA at CCGG sequences. Some of these fragments appeared highly repeated. An MspI fragment which was similar in size to one of the repeat elements (2.9 kilobase pairs) was cloned into the ClaI site of the plasmid vector pBR322 and replicated in a suitable Escherichia coli host strain. The candidal fragment was radiolabeled and used to probe Southern blots of DNA from several Candida species, various other fungi, and mouse and human cells. Only DNA from C. albicans and a strain of Candida stellatoidea was found to contain sequences of significant homology for hybridization. The cloned fragment may possibly be of use as a DNA probe for detection of the presence of C. albicans.
Campylobacter pylori isolated from the stomach of the monkey, Macaca nemestrina
Authors: Bronsdon M A , Schoenknecht F D
Published: January 2020

Campylobacter pylori was isolated from the gastric mucosa in 6 of 24 pigtailed macaques (Macaca nemestrina) examined by gastric biopsy and culture; 3 isolates were recovered during gastroendoscopy, and 3 were recovered at necropsy. The isolates were morphologically and biochemically similar to the human type strain NCTC 11638, differing only in colony diameter, pigmentation, and rate of growth. Identity of the isolates was confirmed by whole-genomic DNA-DNA hybridization with the type strain. Colonization of the monkey stomachs was associated with hypochlorhydria and histologic features resembling type B chronic gastritis in humans. Host animals exhibited no morbid clinical effects of colonization, although endoscopy revealed inflammation, erythema, and friable tissue in some animals. The discovery of C. pylori occurring spontaneously in M. nemestrina extends the known range of the hosts of the organism and offers the possibility of a natural or experimental model of the infection in monkeys.
Staining characteristics of six commercially available monoclonal immunofluorescence reagents for direct diagnosis of Chlamydia trachomatis infections
Authors: Cles L D , Bruch K , Stamm W E
Published: January 2020

Using purified elementary bodies of 14 Chlamydia trachomatis serovars in an in vitro assay, we compared the staining characteristics of six commercially available monoclonal antibody reagents used for direct immunofluorescent staining of patient smears. Considerable variation in the degree of brightness, consistency of staining, and specificity of the six reagents was found. Monoclonal antibodies against the major outer membrane proteins of C. trachomatis produced brighter fluorescence, more consistent elementary body morphology, and less nonspecific staining than did monoclonal antibodies directed against chlamydial lipopolysaccharide.
Laboratory diagnosis of peritonitis in patients on continuous ambulatory peritoneal dialysis
Authors: Ludlam H A , Price T N , Berry A J , Phillips I
Published: January 2020

The clinical course and laboratory diagnosis of peritonitis in patients undergoing continuous ambulatory peritoneal dialysis was studied in 32 consecutive episodes. Peritonitis was associated with a failure in aseptic technique in eight episodes and with an exit-site infection in four episodes. Intraperitoneal vancomycin and ceftazidime were safe, effective, and convenient. Most patients administered their antibiotics at home, and symptoms usually resolved by day 4. Culture of the deposit obtained by centrifugation of 50 ml of effluent after leukocyte lysis provided the best rate of recovery (84% culture positive) but was technically demanding. Filtration of the same volume without leukocyte lysis was simple to perform and almost as effective. Enrichment was less satisfactory (65% culture positive) owing to the presence of antibiotic or infection with fastidious microorganisms. Culture of 50 ml of effluent after concentration by a commonly used laboratory technique, centrifugation without leukocyte lysis, performed poorly (59% culture positive at 48 h), as this method caused sequestration and death of microorganisms within the leukocytes. Culture of nearly 1 liter of effluent from 33 asymptomatic patients by the same techniques yielded no microorganisms.
Pneumocystis carinii antigen detection in rat serum and lung lavage
Authors: McNabb S J , Graves D C , Kosanke S D , Moyer M J , Ivey M H
Published: January 2020

We developed a modified double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) that detected relatively low concentrations of known Pneumocystis carinii antigen added to buffer or rat sera. Artificial immunization-derived polyclonal rabbit anti-P. carinii antibody was used on the solid phase to capture the antigen. Infection-derived (after P. carinii pneumonia) polyclonal rat anti-P. carinii antibody or a mixture of five murine monoclonal antibodies was used as the antigen detector antibody. Rabbit anti-rat immunoglobulin G antibody or goat anti-mouse immunoglobulin G antibody conjugated to alkaline phosphatase was used as the final antibody. After standardization and optimization of the various reactants in this ELISA system, approximately 53 ng of known P. carinii antigen per ml suspended in phosphate-buffered saline-Tween 20 buffer or 210 ng of antigen per ml suspended in normal rat serum diluted 1:4 could be detected. In addition, an indirect ELISA for P. carinii antibody measurement was developed, using as the antigen a soluble supernatant from a sonicated preparation of Percoll-purified whole cysts and trophozoites to coat the solid phase. Limited studies with sera from a small number of caesarian-obtained, barrier-sustained rats from Charles River Breeding Laboratories, Inc., and the National Institutes of Health and sera from normal and heavily infected rats indicated that the caesarian-obtained, barrier-sustained rats had negligible levels of antibody. The normal and heavily infected rats had variable antibody titers. A significantly high level of P. carinii antigenemia was detected in only 2 (11%) of 18 heavily infected rats. Extensive studies of the P. carinii pneumonia rat model with the ELISA did not reveal significant serum P. carinii antigenemia during the acute stage of infection. However, soluble P. carinii antigen was detected by the ELISA and Western blot assays in the supernatant of lavage fluid after centrifugation to sediment intact organisms. As expected, P. carinii antigens were detected by these assays in the lavage pellet recovered after centrifugation. In conclusion, the antigen assay used in this study detected P. carinii antigen in lung lavage but failed to detect P. carinii antigen in rat serum during the acute phase of infection.
Comparison of diploid fibroblast and rabbit kidney tissue cultures and a diploid fibroblast microtiter plate system for the isolation of herpes simplex virus
Authors: Langenberg A , Zbanyszek R , Dragavon J , Ashley R , Corey L
Published: January 2020

We evaluated the relative sensitivities of two cell systems (rabbit kidney [RK] and human diploid fibroblast [DF; human embryonic tonsil]) in standard tube cultures versus DF cells in a 48-well microtiter plate system for the detection of both symptomatic and asymptomatic herpes simplex virus (HSV) infection. At least one system isolated HSV in 111 of 809 specimens (13.7%). HSV was isolated in RK tube cultures from 110 specimens (99%), in DF tube cultures from 91 specimens (82%), and in DF microtiter plates from 95 specimens (86%). The frequency of HSV isolation varied with the anatomic site and the presence or absence of a herpetic lesion. The sensitivities of the three culture systems remained similar whether the specimens were obtained from lesions or whether the specimens were taken to determine if asymptomatic excretion of HSV was present. While RK tube cultures were more sensitive than DF tube cultures, the DF microtiter plate system was as sensitive as DF tube cultures and its use is supported as a cheaper and less labor-intensive method for the detection of HSV.
Semiquantitation of bacteria in sputum gram stains
Authors: Valenstein P N
Published: January 2020

In many clinical laboratories, bacteria seen in Gram-stained sputum smears are reported semiquantitatively, using a three- or four-category scale consisting of ratings such as numerous, moderate, rare, and none seen. The consistency with which these categories are assigned was evaluated by repeatedly presenting coded smears to seven experienced microbiology technologists. Technologists rated the same smear twice, pairs of smears prepared from the same specimen, and smears prepared after specimen refrigeration. Agreement was assessed with the weighted kappa test. Semiquantitation of gram-negative rods, gram-positive diplococci, and gram-positive cocci in clusters all showed poor reproducibility (kappa = 0.32, 0.34, and 0.17, respectively). Twenty-four percent of paired ratings differed by two or more categories. Lack of reproducibility was due mainly to the inability of the technologists to render a consistent rating when viewing the same slide on separate occasions (P less than 0.001). Variation in the rating styles of different technologists, differences between smears prepared from the same specimen, and specimen refrigeration tended to further decrease the consistency of ratings, but the reductions were not statistically significant. The quantity of potentially pathogenic bacteria in sputum smears is not estimated consistently with standard microscopy procedures and should not be reported.
Detection of Vibrio cholerae with monoclonal antibodies specific for serovar O1 lipopolysaccharide
Authors: Adams L B , Henk M C , Siebeling R J
Published: January 2020

Six hybridoma cell lines, each of which produced a monoclonal antibody (MAb) against Vibrio cholerae O1 lipopolysaccharide (LPS), were established. Each MAb was active serologically by both enzyme-linked immunosorbent assay (ELISA) and the slide agglutination test. In the ELISA, each MAb was tested against 7 O1 and 9 non-O1 LPS preparations. Three MAbs reacted with both Inaba and Ogawa serovars (A antigen), two MAbs reacted with the Ogawa serovars only (B antigen), and one MAb reacted with the Inaba serovars only (C antigen). Each MAb was also tested in the ELISA against whole-cell preparations of 37 O1 and 52 non-O1 V. cholerae serovars, 20 heterologous Vibrio species, and 37 heterologous bacterial species. The MAbs reacted with V. cholerae O1 cells only, except for one anti-A antigen MAb which reacted weakly with five V. cholerae non-O1 serovars and Serratia marcescens. Each anti-A antigen MAb was labeled with fluorescein isothiocyanate (FITC) and tested by direct immunofluorescence against selected O1 and non-O1 serovars. Each MAb-FITC conjugate, when tested alone, exhibited O1-specific fluorescence; however, mixtures of the MAb-FITC dramatically enhanced fluorescence intensity on O1 cells. This finding was also visualized by immunoelectron microscopy on both thin-sectioned and negatively stained O1 cells by using an anti-mouse immunoglobulin-colloidal gold conjugate. These results suggest that the A antigen can be described by more than one epitope and that a superior serotyping reagent can be prepared from a defined mixture of MAbs.
Screening for hepatitis B virus in healthy blood donors by molecular DNA hybridization analysis
Authors: Sun C F , Pao C C , Wu S Y , Liaw Y F
Published: January 2020

A DNA molecular hybridization technique employing a purified adw subtype hepatitis B virus (HBV) cloned DNA of 3.2 kilobase pairs as a probe was used to screen for the presence of HBV DNA in blood samples collected from 486 apparently healthy blood donors. Eighteen of 104 (17.3%) hepatitis B surface antigen (HBsAg) carriers and 7 of 382 (1.8%) HBsAg-negative individuals had circulating HBV DNA in their sera. Among the seven individuals who were positive for HBV DNA but negative for HBsAg, three had antibodies against both HBsAg (anti-HBsAg) and hepatitis B core antigen, one had only anti-HBsAg, one had both anti-hepatitis B core antigen and anti-hepatitis B e antigen and two were negative for all the above HBV markers. The results suggest that the absence of HBsAg in otherwise apparently healthy individuals may not be enough to ensure lack of circulating HBV.
Method for in vitro conversion of rough strains of Pseudomonas aeruginosa to smooth strains
Authors: Liu P V
Published: January 2020

Rough and serologically nontypable strains of Pseudomonas aeruginosa were found to become smooth and typable in vitro when they were maintained in a soft agar containing 0.5% peptone in distilled water and the dried out agar cultures were replenished monthly with a similar peptone solution in distilled water. After about a year, all of the nine rough strains of P. aeruginosa maintained this way became smooth and typable, and their pigmentations also changed significantly. Some melanogenic strains became fluorescine producers, and some weakly chromogenic strains became strong producers of fluorescine and pyocyanin.
Identification of Actinomyces (Corynebacterium) pyogenes with the API 20 Strep system
Authors: Morrison J R , Tillotson G S
Published: January 2020

A total of 62 strains of Actinomyces pyogenes (previously Corynebacterium pyogenes) were examined by the API 20 Strep system (API System, La Balme Les Grottes, Montalieu-Vercieu, France). The system was shown to be reliable and rapid when the tests were compared with standard identification methods. No confusion occurred with streptococcal profiles in the current API 20 Strep data base.
Mycobacterial cross contamination during radiometric culturing
Authors: Vannier A M , Tarrand J J , Murray P R
Published: January 2020

Pseudobacteremias in blood cultures performed on the BACTEC radiometric blood culture system have been reported. We report three cases of cross contamination with Mycobacterium avium that occurred when mycobacteria were cultured with the BACTEC 460 TB system. Malfunction of the needle sterilization heating block was demonstrated.
Comparison of four methods to detect Trichomonas vaginalis
Authors: Thomason J L , Gelbart S M , Sobun J F , Schulien M B , Hamilton P R
Published: January 2020

Four methods for the detection of Trichomonas vaginalis in vaginal secretions from 88 symptomatic patients were compared: wet-mount examination, Kupferberg liquid medium, Hirsch charcoal agar, and the Papanicolaou smear. Hirsch diphasic slants and Kupferberg medium did not significantly differ in sensitivity from direct examination of wet mounts. The Papanicolaou smear identification of trichomonads was found to be the least sensitive method evaluated.
Enzyme-linked immunosorbent assays for measurement of reovirus immunoglobulin G, A, and M levels in serum
Authors: Richardson S C , Bishop R F , Smith A L
Published: January 2020

Enzyme-linked immunosorbent assays were developed for the quantitation of anti-reovirus immunoglobulin G (IgG), IgA, and IgM in serum. The assays were specific, sensitive, and reproducible and measured antibodies to the reovirus group antigen. They should permit diagnosis of recent reovirus infection and longitudinal study of the development of reovirus group antibodies.
Evaluation of immunoglobulin G preparations for anti-cytomegalovirus antibodies with reference to neutralizing antibody in the presence of complement
Authors: Eizuru Y , Ueno I , Minamishima Y
Published: January 2020

Eighteen commercially available immunoglobulin G (IgG) preparations were evaluated by various methods for anti-cytomegalovirus (CMV) antibodies. The modifications of immunoglobulin G molecules, such as sulfonation, alkylation, and enzyme treatments, resulted in profound reductions in anti-CMV antibody titers. However, after reversion in vitro, sulfonated IgG preparations which have been shown to be reverted in vivo (Y. Masuho, K. Tomibe, K. Matsuzawa, T. Watanabe, S. Ishimoto, S. Tsunoda, and T. Noguchi, J. Biochem. 79:1377-1379, 1976) restored the anti-CMV antibody titers to levels comparable with those of nonsulfonated IgG preparations. There is no good correlation among the titers tested by different methods. As a method which reflects the biological activities in vivo, the neutralization test in the presence of complement is suitable for the evaluation of IgG preparations.
Detection of cytomegalovirus in shell vial cultures by using a DNA probe and early nuclear antigen monoclonal antibody
Authors: Scott A A , Walker K A , Hennigar L M , Williams C H , Manos J P , Gansler T
Published: January 2020

An in situ biotinylated DNA probe assay was evaluated as an adjunct to anti-cytomegalovirus early nuclear antigen indirect immunofluorescence and cytopathic effect on cytomegalovirus-infected monolayers in shell vial cultures. Viral infection was detected by early nuclear antigen indirect immunofluorescence at 24 h and by DNA probe assay and shell vial cytopathic effect at 5 days.
Serum antibody response to the 70,000-molecular-weight neisserial common antigen in humans infected by Neisseria gonorrhoeae
Authors: Aoun L , Cremieux A C , Casin I , Morel P , Martin P M
Published: January 2020

We evaluated the presence of antibodies directed against a 70,000-molecular-weight (70K) common neisserial antigen in sera from patients with first or repeated gonococcal infections and in sera from healthy controls. Sera were taken as soon as possible after the onset of disease, and anti-70K antibodies were detected by Western blot (immunoblot). Results show that significantly fewer patients with gonococcal infection possessed anti-70K antibodies than controls (P less than 0.001). This suggests a possible role of anti-70K antibodies in natural immunity against Neisseria gonorrhoeae.
Efficacies of rapid agglutination tests for identification of methicillin-resistant staphylococcal strains as Staphylococcus aureus
Authors: Piper J , Hadfield T , McCleskey F , Evans M , Friedstrom S , Lauderdale P , Winn R
Published: January 2020

Four commercially available rapid agglutination tests for the identification of Staphylococcus aureus were compared with the tube coagulase test for the identification of 300 methicillin-resistant isolates of staphylococci. Isolates tested included 207 methicillin-resistant S. aureus and 93 coagulase-negative staphylococci, collected from five medical centers. Strain variability was documented by phage typing and antimicrobial susceptibility patterns. Results of rapid identification tests ranged between 82 and 86% sensitivity, significantly poorer than the 98% sensitivity which the tube coagulase test provided.
New selective medium for Pseudomonas aeruginosa with phenanthroline and 9-chloro-9-[4-(diethylamino)phenyl]-9,10-dihydro-10- phenylacridine hydrochloride (C-390)
Authors: Campbell M E , Farmer S W , Speert D P
Published: January 2020

A new selective medium (PC agar) for the isolation of Pseudomonas aeruginosa was developed, consisting of 30 micrograms of 9-chloro-9-[4-(diethylamino)phenyl]-9,10-dihydro-10-phenylacridine hydrochloride (C-390) per ml and 30 micrograms of phenanthroline per ml in Columbia agar. PC agar was superior to phenanthroline, C-390, acetamide, and cetrimide agars for the selective growth of P. aeruginosa.
Surveillance of invasive Streptococcus pneumoniae infection in Quebec, Canada, from 1984 to 1986: serotype distribution, antimicrobial susceptibility, and clinical characteristics
Authors: Jetté L P , Lamothe F
Published: January 2020

Four hundred and sixty-eight strains of Streptococcus pneumoniae isolated from blood or normally sterile body fluids in 14 Quebec, Canada, hospitals between 1 July 1984 and 31 December 1986 were serotyped, their susceptibilities to antimicrobial agents were determined, and the laboratory data were correlated with the clinical information. The distribution pattern of the serogroups and serotypes was similar to that observed previously in Quebec and to that in other parts of Canada and in the United States. No regional variation was observed. The distribution of serogroups and serotypes was different in pediatric patients (less than 18 years old) when compared with that of adults (P less than 0.001). Overall, 94% of the strains were represented in 23-valent vaccine. The total mortality rate was 12.8% and increased with age: 1.6% in pediatric patients, 14.8% in patients 18 to 64 years old, and 31% in those greater than or equal to 65 years old (P less than 0.001). In patients 18 to 64 years old, the mortality rate was higher when an underlying condition that could have justified prior vaccination was present (P = 0.008). In patients greater than or equal to 65 years old, the mortality rates were similar in those with and those without underlying conditions, suggesting that vaccine use in all patients greater than or equal to 65 years old might be appropriate. Only six patients had received pneumococcal vaccine before infection. Only 15 strains (3.2%) were moderately susceptible or resistant to one or more of the antimicrobial agents tested. Six strains were moderately susceptible to penicillin G, and none was fully resistant.
Optimization of human papillomavirus genotype detection in cervical scrapes by a modified filter in situ hybridization test
Authors: Melchers W J , Herbrink P , Walboomers J M , Meijer C J , vd Drift H , Lindeman J , Quint W G
Published: January 2020

Optimal conditions for the screening of cervical scrapes for human papillomavirus (HPV) were investigated by using filter in situ hybridization. Since integrated and episomal HPV can be found, cell lines containing viral DNA in an integrated form (HPV in CaSki) or in an episomal state (BK virus-induced hamster tumor cells) were used for optimization experiments. An increase in sensitivity was achieved by alkaline denaturation and neutralization before the specimens were spotted onto the membrane. This increase was 5-fold for the episomal virus and 16-fold for the integrated virus in the model system, as compared with other methods. To evaluate this method on clinical material, 1,963 cervical scrapes were screened for the presence of HPV 6/11 and HPV 16. Nineteen scrapes were positive for HPV 6/11 or HPV 16; and in 1,810 scrapes, no HPV 6/11 or HPV 16 could be detected by the modified filter in situ hybridization technique. Scrapes from which the interpretation of the modified filter in situ hybridization results were equivocal (n = 71, 3.6%) or in which positivity was detected for both HPV 6/11 and HPV 16 (n = 63, 3.2%) were further analyzed by the DNA dot spot technique. Eight scrapes with an equivocal result and only one scrape showing a double positivity by the modified filter in situ hybridization technique could be confirmed in the dot spot assay. In the total group 12 scrapes were positive for HPV 6/11 DNA, 15 were positive for HPV 16 DNA, and 1 was positive for both HPV 6/11 and HPV 16 DNA. Southern blot analysis on modified filter in situ hybridization-positive and -negative scrapes revealed a 100% correlation.
Antibody response to Haemophilus somnus Fc receptor
Authors: Yarnall M , Corbeil L B
Published: January 2020

To characterize the bovine immune response to an Haemophilus somnus antigen known to be recognized by convalescent-phase serum, we studied isotypic antibody titers to the 270-kilodalton protein, which we had previously shown to be an immunoglobulin Fc receptor. With a modified immunodot procedure, an immune response was detected after experimental H. somnus abortion, experimental H. somnus pneumonia, or vaccination with commercial H. somnus vaccine, with the greatest titer found within the immunoglobulin G2 isotype. With protein A peroxidase conjugate, which detects primarily bovine immunoglobulin G2, we showed that cattle with H. Somnus disease could be distinguished from clinically normal carriers, culture-negative cattle, or cattle with disease due to Pasteurella haemolytica or P. multocida. Little cross-reactivity between the 270-kilodalton Fc receptor antigen and antigens from other gram-negative bovine pathogens was seen. Thus, this antigen may be a useful diagnostic antigen.
Antigenically variable Borrelia burgdorferi isolated from cottontail rabbits and Ixodes dentatus in rural and urban areas
Authors: Anderson J F , Magnarelli L A , LeFebvre R B , Andreadis T G , McAninch J B , Perng G C , Johnson R C
Published: January 2020

Spirochetes were isolated from 71 subadult Ixodes dentatus removed from cottontail rabbits captured in Millbrook, N.Y., and in New York, N.Y. Spirochetes were also cultured from kidney tissues of six rabbits. While all isolates reacted with monoclonal antibody H9724, which identifies the spirochetes as borreliae, more than half did not bind with antibody H5332 and even fewer reacted with H3TS, both of which were produced to outer surface protein A of Borrelia burgdorferi. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis protein profiles of three isolates differed from one another and from all previously characterized B. burgdorferi strains from humans, ticks, and wildlife in North America. The 12 periplasmic flagella that originated subterminally from each pointed end of a rabbit Borellia isolate contrasted with the 11 or fewer flagella for B. burgdorferi reported previously from North America. Although DNA homology and restriction endonuclease analysis also revealed differences among a rabbit kidney isolate, an I. dentatus isolate, and B. burgdorferi B31, similarities were sufficient to lead us to conclude that the borreliae in rabbits and I. dentatus are B. burgdorferi. Enzyme-linked immunosorbent assay titers of sera from humans with diagnosed Lyme disease to rabbit tick B. burgdorferi were often similar to one another and to those recorded for a reference B. burgdorferi strain.