bluuebunny/crossref_metadata_2025 huggingface dataset:
Long-lasting cell-mediated immunity induced by a live Francisella tularensis vaccine
Authors: Tärnvik A , Löfgren M L , Löfgren S , Sandström G , Wolf-Watz H
Published: January 2020

The lymphocyte stimulation test was used to estimate specific cell-mediated immunity after vaccination with the live vaccine strain of Francisella tularensis. Nonvaccinated individuals and individuals vaccinated 1, 5 to 6, 7 to 8, and 9 years previously were tested. Lymphocytes from most vaccinees responded to an antigen preparation of the vaccine strain, and those vaccinated 9 years before testing responded to a similar extent as did vaccines in the other groups. A new technique was developed to study the presence of T lymphocytes among the stimulated cells. Stimulated cells were allowed to incorporate [14C]thymidine and were then fractionated into T and non-T lymphocytes. Most of the incorporation appeared in the fraction containing T lymphocytes. The data indicate that cell-mediated immunity endures for at least 9 years after vaccination with the live F. tularensis vaccine.
Use of lysis-centrifugation (isolator) and radiometric (BACTEC) blood culture systems for the detection of mycobacteremia
Authors: Gill V J , Park C H , Stock F , Gosey L L , Witebsky F G , Masur H
Published: January 2020

Patients with acquired immune deficiency syndrome may develop infection with mycobacteria, particularly Mycobacterium avium-M. intracellulare (MAI). These infections can frequently be associated with demonstrable mycobacteremia with the organism. In this study, we compared the sensitivity of a radiometric (BACTEC system; Johnston Laboratories, Inc., Towson, Md.) liquid medium culture system with that of conventional solid mycobacterial culture media for cultures of blood from these patients. Both systems were inoculated with blood concentrate prepared by lysis-centrifugation (Isolator; Du Pont Co., Wilmington, Del.). Of 46 acquired immune deficiency syndrome patients whose blood was cultured, 28% had cultures positive for MAI. Patients had from less than 1 to more than 100 MAI colonies per ml of blood. Lowenstein-Jensen and Middlebrook 7H11 agars were comparable in recovery of MAI. BACTEC 12A vials containing double the standard volume of medium (4 ml) were more sensitive and were positive slightly earlier than vials containing the standard volume (2 ml). Conventional media detected 98% of positive cultures; BACTEC vials containing double volumes of medium detected 94% of positive cultures, whereas single-volume vials detected 77%. BACTEC vials were positive approximately 5 to 6 days sooner than slants or plates containing conventional media. For a few cultures, the use of unconcentrated blood was compared with the use of Isolator-concentrated blood by using each of these as inocula for BACTEC vials. Results for these cultures suggested that, although the use of Isolator-concentrated blood resulted in greater sensitivity than the use of unconcentrated blood would, the use of unconcentrated blood would still result in the detection of at least 78% of positive cultures.
Recent vesicular stomatitis virus infection detected by immunoglobulin M antibody capture enzyme-linked immunosorbent assay
Authors: Vernon S D , Webb P A
Published: January 2020

We developed an enzyme-linked immunosorbent assay (ELISA) that was capable of detecting immunoglobulin M (IgM) antibody to vesicular stomatitis virus (VSV) in the sera of experimentally and naturally infected cattle and horses. The detection of IgM in the sera of these animals permitted an estimate of the recency of infection by VSV serotype New Jersey. A VSV serotype New Jersey epizootic strain isolated from a horse and passed once in an Aedes albopictus cell line was used to infect a horse and a calf. Sera from these animals were used to standardize the ELISA. This assay was used to test sera from cattle and horses involved in the 1982 VSV epizootic. Comparative antibody titrations were performed by three systems: the serum-dilution plaque-reduction neutralization, complement fixation, and indirect immunofluorescent tests. The antibody titers by neutralization and the ELISA were comparable for the period that IgM was present; when IgM ELISA titers diminished, the neutralization titers remained high. The complement fixation and indirect immunofluorescent antibody titers followed closely the IgM pattern determined by the ELISA. The capture IgM ELISA is applicable for the rapid detection of IgM antibody to VSV in cattle and horses and is a useful assay of recent infection.
Development of a DNA probe to detect Salmonella typhi
Authors: Rubin F A , Kopecko D J , Noon K F , Baron L S
Published: January 2020

This study was undertaken to identify a DNA sequence that could be used to facilitate the diagnostic identification of Salmonella typhi, the causative agent of typhoid fever. All virulent S. typhi strains encode a relatively unique capsular antigen termed the virulence (Vi) antigen. Two distinct genetic loci, viaA and viaB, are involved in the synthesis of this antigen. The structural genes, located at viaB, were considered as a possible specific DNA probe. The viaB locus, contained in a recombinant cosmid, was subcloned to various plasmid vectors for this purpose. Selected viaB-region DNA fragments were then analyzed for specificity in DNA colony hybridization reactions with more than 170 strains representing a variety of enteric bacteria. An 8.6-kilobase EcoRI fragment was highly specific for the viaB gene region and was considered a good hybridization probe. This DNA probe should prove useful in rapid diagnostic assays set up to detect S. typhi in mixed bacterial samples (e.g., stools) within a few hours of specimen collection.
Rapid immunotyping of Chlamydia trachomatis with monoclonal antibodies in a solid-phase enzyme immunoassay
Authors: Barnes R C , Wang S P , Kuo C C , Stamm W E
Published: January 2020

The technical complexity of determining the serovar of Chlamydia trachomatis strains has limited the use of serotyping in clinical and epidemiologic studies. We developed a simple method for rapidly serotyping isolates of C. trachomatis by using monoclonal antibodies in a dot-enzyme-linked immunosorbent assay (ELISA) system. Isolates were passaged three to six times in shell vial cultures to greater than 50% monolayer infection, and chlamydial elementary bodies were isolated by sonication and microcentrifugation. Chlamydial antigen was spotted onto a series of replicate nitrocellulose membrane patches and reacted with C. trachomatis-specific monoclonal antibodies. Bound antibody was detected visually by a color reaction by using peroxidase-conjugated anti-mouse immunoglobulins. This method can be routinely applied to 60 or more specimens concurrently. We compared dot-ELISA serotyping with monoclonal antibody microimmunofluorescence serotyping of 124 clinical C. trachomatis isolates and found that dot-ELISA has sensitivity and serotyping accuracy comparable to that of monoclonal antibody microimmunofluorescence.
Evaluation of antisera used for detecting enterotoxigenic Escherichia coli in Sao Paulo
Authors: Guth B E , Trabulsi L R
Published: January 2020

The usefulness of antisera in detecting enterotoxigenic Escherichia coli (ETEC) strains in Sao Paulo was evaluated. Polyvalent antisera detected 49% of ETEC isolates and were more effective in identifying E. coli that produced heat-labile and heat-stable enterotoxins and in strains that produced only heat-stable enterotoxin. ETEC strains not detected by the antisera belonged to different serogroups not isolated in Sao Paulo before; 34% of these strains had undetermined O antigens, and most of them produced only heat-labile toxin. A variation of serogroups over time was especially observed among strains that produced heat-stable toxin. The importance of H-antigen determinations in the effectiveness of ETEC diagnosis by serological methods became evident, as non-ETEC strains were also detected by polyvalent antisera, but their serotypes were different from those of ETEC strains. Although antisera can be used to identify O:H types of ETEC strains with accuracy, serotyping cannot be recommended for routine diagnosis. However, such a procedure may be useful for studying outbreaks of ETEC diarrhea if the involved serotypes are already known.
Improvement of the indirect hemagglutination assay for Salmonella typhi Vi antibodies by use of glutaraldehyde-fixed erythrocytes
Authors: Barrett T J
Published: January 2020

Glutaraldehyde-fixed sheep erythrocytes sensitized with Vi antigen were shown to be usable for at least 1 year in the indirect hemagglutination assay for Vi antibodies. The use of fixed sensitized erythrocytes improves the practicality of the indirect hemagglutination assay by (i) eliminating the need to sensitize cells each time the test is performed; (ii) reducing waste of the purified Vi antigen; and (iii) reducing test-to-test variation.
Inhibitory effect of glucocorticoid hormones on the growth of Inoue-Melnick virus
Authors: Inoue Y K , Nishibe Y , Melnick J L , Seidel E , Wang S S
Published: January 2020

Glucocorticoid hormones inhibited the early stage of propagation of Inoue-Melnick virus in a dose-dependent manner in human diploid cell cultures. Prednisolone and hydrocortisone achieved complete inhibition of replication of the three Inoue-Melnick virus serotypes at concentrations of 0.2 X 10(-4) and 10(-5) M, respectively; strong inhibition was found even at 10(-5) and 10(-6) M, respectively. The steroid hormones did not show a direct effect on the virus.
Effect of polybrene on isolation of Chlamydia trachomatis from clinical specimens
Authors: Sankar-Mistry P , Albota V , Knelsen B
Published: January 2020

Polybrene treatment of McCoy cells was used to increase the infectivity of Chlamydia trachomatis in vitro. The isolation rate for C. trachomatis in 500 specimens from males and females attending a sexually transmitted disease clinic and other clinics increased by 6.8% with the Polybrene treatment. The data also suggested that this treatment facilitated the diagnosis of low grade or latent infections.
Rapid detection of bacteremia in children with a modified lysis direct plating method
Authors: Campos J M , Spainhour J R
Published: January 2020

A modified lysis direct plating method was developed for culturing blood specimens from pediatric patients. The method, which used the commercially available Isolator 1.5 Microbial Tube, consisted of direct inoculation of lysed blood to agar media and into a tube containing modified eugonic broth. During a 4,478-specimen study, 195 of 324 (60.2%) clinically significant isolates were detected within 24 h of specimen arrival in the laboratory. This method offers a rapid means to obtain quantitative blood culture results from pediatric patients and is an alternative to conventional and radiometric broth culture procedures.
Evaluation of a new method for routine in vitro cultivation of Giardia lamblia from human duodenal fluid
Authors: Gordts B , Hemelhof W , Van Tilborgh K , Retoré P , Cadranel S , Butzler J P
Published: January 2020

Although a method for in vitro cultivation of Giardia lamblia was described as early as 1927, only a few clinical isolates were cultured in vitro due to the complexity of the techniques. We developed a method which allows for the routine isolation of Giardia trophozoites from human duodenal fluid and maintenance of the organisms in axenic culture. This study evaluates the method in 198 patients. Seventeen strains of Giardia were isolated and cultivated axenically. The method was more sensitive than the microscopic examination of aspirated fluid and examination of an impression of mucosal biopsy. Five patients, however, excreted cysts in the stool, although no trophozoites could be demonstrated in the duodenal fluid. G. lamblia were cultivated from one patient who did not excrete cysts. The method will enable the collection of G. lamblia strains from clinical material in large numbers and can offer an important advance in epidemiological, biochemical, immunological, and therapeutic investigations of giardiasis.
Comparison of outer membrane protein and biochemical profiles of Haemophilus aegyptius and Haemophilus influenzae biotype III
Authors: Carlone G M , Sottnek F O , Plikaytis B D
Published: January 2020

Haemophilus aegyptius and Haemophilus influenzae biotype III are morphologically and biochemically similar; however, their outer membrane protein (Sarkosyl insoluble) profiles are distinct. Of 18 strains of H. aegyptius examined, 15 had a type 1 protein profile, and 3 had a type 2 profile, whereas the 5 strains of H. influenzae biotype III examined had three other protein profile types. All Haemophilus strains examined had 31- and 76-kilodalton (kDa) proteins and minor proteins with molecular masses between 20 and 100 kDa. H. aegyptius, with a type 1 protein profile, had major outer membrane proteins with apparent molecular masses of 27, 35.5, and 41.5 kDa, and H. aegyptius, with a type 2 protein profile, had 26-, 29-, 39.5-, and 41-kDa proteins. The type strain of H. influenzae biotype III had three major outer membrane proteins with apparent molecular masses of 29, 38.5 and 40 kDa. Four other strains designated as H. influenzae biotype III had major outer membrane proteins between 27 and 41.5 kDa representing two additional protein profiles.
Attenuation of wild-type human influenza A virus by acquisition of the PA polymerase and matrix protein genes of influenza A/Ann Arbor/6/60 cold-adapted donor virus
Authors: Snyder M H , Clements M L , De Borde D , Maassab H F , Murphy B R
Published: January 2020

Wild-type influenza A viruses can be attenuated for humans by the acquisition of genes from the A/Ann Arbor/6/60 cold-adapted (ca) donor virus. Six-gene reassortants, that is, viruses containing the hemagglutinin and neuraminidase surface glycoprotein genes of the wild-type virus and the six remaining RNA segments of the ca donor virus, are consistently attenuated for humans. During the production of a six-gene reassortant virus containing the surface glycoproteins of the A/Washington/897/80 (H3N2) wild-type virus, a reassortant virus was isolated that contained RNA segments 3 (coding for the polymerase PA protein) and 7 (coding for matrix [M] proteins) from the ca parent and all other genes from the wild-type virus. This reassortant virus is referred to as a two-gene reassortant. Because the gene or set of genes responsible for the attenuation of ca reassortant viruses has not been defined, we evaluated the two-gene reassortant for level of replication and level of virulence in ferrets and in humans, and we compared its characteristics to those of a six-gene reassortant virus derived from the same two parents. The two-gene reassortant virus infected each of 14 adult seronegative (serum hemagglutination inhibition titer of less than or equal to 1:8) volunteers when administered intranasally at a dose of 10(7) 50% tissue culture infectious doses, yet it did not produce illness. The level of replication of the two-gene reassortant virus in the upper respiratory tract was equivalent to that of the six-gene reassortant virus. This demonstrates that transfer of the A/Ann Arbor/6/60 ca PA polymerase and M genes is sufficient to confer the attenuation phenotype on wild-type influenza A viruses. In the context of previous observations, these results suggest that the A/Ann Arbor/6/60 ca donor virus PA polymerase gene plays a major role in the attenuation of ca reassortant viruses.
Enzyme immunoassay inhibition assay for the detection of rat rotavirus-like agent in intestinal and fecal specimens obtained from diarrheic rats and humans
Authors: Vonderfecht S L , Miskuff R L , Eiden J J , Yolken R H
Published: January 2020

An enzyme immunoassay inhibition assay was developed to detect rat rotavirus-like agent (RVLA) or antigenically related viruses in intestinal washings and homogenates obtained from diarrheic rats and in fecal specimens obtained from diarrheic humans. In this assay, RVLA antigens in a test sample inhibited the binding of a biotin-labeled anti-rat RVLA antibody preparation to rat RVLA antigens bound to the solid phase. Intestinal washings and homogenates obtained 1 day after RVLA infection of suckling rats inhibited the binding of the biotin-labeled antibody to the solid-phase rat RVLA antigens by 76 to 100%. The inhibition was blocked by RVLA immune rat serum but not by nonimmune rat serum. Of 27 fecal specimens obtained from diarrheic humans, 6 produced disease characteristic of rat RVLA infection when inoculated into suckling rats. Four of these six specimens produced greater than 50% inhibition in the enzyme immunoassay. Fecal specimens obtained from diarrheic humans that were determined to be negative for RVLA produced an average inhibition of 9.2%. This enzyme immunoassay appears to be a useful diagnostic and research tool for the study of infections with at least one of the antigenically distinct rotaviruses.
Rapid and sensitive identification of Mycobacterium tuberculosis
Authors: Knisley C V , Damato J J , McClatchy J K , Brennan P J
Published: January 2020

The fatty acid constituents of 14 species of Mycobacterium (14 isolates) and one isolate each of Corynebacterium xerosis, Nocardia asteroides, and Streptomyces albus were examined with the purpose of distinguishing Mycobacterium tuberculosis from other acid-fast bacilli. Combined thin-layer chromatography (TLC) of methyl mycolates and gas-liquid chromatography (GLC) of shorter-chain fatty acid esters provided an unequivocal identification of M. tuberculosis in a matter of 2 to 3 days. The methodology included rapid and simplified procedures for methanolysis and extraction of bacterial lipids with equally facilitated GLC and TLC analyses. These studies were performed with 0.5 to 1.0 mg of dry bacterial cells (approximately 2.5 X 10(7) CFU). When applied to 100 unknown cultures, the methodology with combined TLC-GLC correctly identified all 49 of the M. tuberculosis-Mycobacterium bovis cultures and a variety of other mycobacterium taxa. It was also interesting to note that 28 of 39 (72%) of the nontuberculous mycobacteria were correctly identified. An additional five species were tentatively identified as belonging to either of two species (Mycobacterium malmoense, Mycobacterium terrae), but in all cases, the two species belonged to the same Runyon group. All six nonmycobacterial species were differentiated from the mycobacteria studied.
Cytomegalovirus antigenic heterogeneity can cause false-negative results in indirect hemagglutination and complement fixation antibody assays
Authors: Faix R G
Published: January 2020

Cord sera and antepartum maternal sera from three congenitally cytomegalovirus (CMV)-infected infants and their mothers were CMV seronegative (titer, less than 8) in a complement fixation (CF) assay with a glycine-extracted CMV AD169 antigen; sera from two of the infants and mothers were also seronegative in a commercial indirect hemagglutination (IHA) assay with AD169 antigen. In tests with their own CMV isolates propagated and made into glycine-extracted CF antigen, all were seropositive. When 108 random cord sera were assayed for CF antibody with AD169, Davis, and A antigens (A is a locally derived antigen from one of the above infants), 44 were seropositive and 54 were seronegative for all three antigens. Of the remaining 10 sera, 4 were positive for A only, 3 were positive for A and Davis only, 2 were positive for Davis and AD169 only, and 1 was positive for AD169 only. All 10 were positive when a mixture of all three antigens was used. The IHA assay with AD169 antigen was positive with only 4 of these 10 sera. These results suggest that up to 6% of sera may be misclassified as seronegative in the CF and IHA assays if only a single antigen is used.
Advantages of a modified toluidine blue O stain and bronchoalveolar lavage for the diagnosis of Pneumocystis carinii pneumonia
Authors: Gosey L L , Howard R M , Witebsky F G , Ognibene F P , Wu T C , Gill V J , MacLowry J D
Published: January 2020

A modified toluidine blue O staining technique for Pneumocystis carinii is described. An easily prepared sulfation reagent made with sulfuric and acetic acids was used. The stain can be employed for bronchoalveolar lavages and lung tissue touch preparations. Most background material was removed by the sulfation reagent, slides were generally easy to read, and time from receipt of a specimen to reporting of results was approximately 1 h. P. carinii cysts were more easily visualized with this stain than in slides stained with modified methylene blue, Gram, Gram-Weigert, and standard toluidine blue O procedures. The importance of certain procedural aspects of subsegmental bronchoalveolar lavage, by which most of the specimens were obtained, is also emphasized.
Determination of antibodies to pneumococcal C polysaccharide in patients with community-acquired pneumonia
Authors: Holmberg H , Krook A , Sjögren A M
Published: January 2020

The pneumococcal C polysaccharide (PnC) is species specific and believed to be a cell wall component of all capsular types. Antibodies against PnC in human sera have been demonstrated previously, but the question of whether a rise in these antibodies occurs during pneumococcal infections has not been investigated. We used an indirect enzyme-linked immunosorbent assay (ELISA) for the estimation of PnC antibodies in 124 hospital-treated patients with pneumonia. In 3 of 6 patients with pneumococcal bacteremia and in 17 of 44 patients with S. pneumoniae isolated in the blood, sputum, or nasopharynx, a significant rise in antibody levels was recorded, accounting for a sensitivity of 38.6%. Of 35 patients with pneumonia of other known or suspected etiology, 1 gave a positive result, corresponding to a specificity of 97.1%. In addition, 3 of 8 patients with PnC antigen in the sputum as the only etiological finding and 5 of 37 patients with unknown etiology gave positive results. The PnC antibodies did not seem to have any protective capacity against pneumonia caused by pneumococci. The ELISA, in which only one antigen preparation was used, was more simple than other tests in which traditional capsular antigen preparations are used. It might therefore be used as a supplemental method in the diagnosis of pneumococcal pneumonia. The problems involved in expressing serum titers obtained with the ELISA are discussed.
Autobac susceptibility testing of methicillin-resistant Staphylococcus aureus isolated in an Australian hospital
Authors: Putland R A , Guinness M D
Published: January 2020

Semiautomated rapid broth elution (Autobac Multi-Test System; General Diagnostics, Div. Warner-Lambert Co., Morris Plains, N.J.) and disk diffusion tests were compared with an agar dilution breakpoint method to determine the antibiotic susceptibility of 147 methicillin-resistant Staphylococcus aureus isolates from our hospital. Although the disk diffusion method, in general, correlated well with the agar dilution tests, the overall agreement of the Autobac tests with agar dilution tests was only 79%, with many very major discrepancies occurring with clindamycin (88%), gentamicin (33%), and methicillin (15%). When we used a 10-fold higher inoculum for the Autobac tests, all isolates were shown to be resistant to methicillin, but significant numbers of major and minor discrepancies occurred with chloramphenicol, fusidic acid, and neomycin. The majority of isolates were shown to belong to three biotypes, distinguishable by lactose fermentation, lipolysis, hemolysis, and pigment production. The antibiotic susceptibility profile of one biotype was found to be markedly different from those of the other biotypes and contained a high incidence of clindamycin susceptibility and neomycin, gentamicin, and kanamycin resistance. In contrast, the other two biotypes had a high incidence of clindamycin, gentamicin, and kanamycin resistance and neomycin susceptibility and accounted for most of the very major discrepancies in the clindamycin and aminoglycoside tests. In these methicillin-resistant S. aureus strains, discrepancies possibly may arise from partial expression of methicillin resistance, dissociated or inducible clindamycin resistance, and instability of gentamicin resistance.
New germ tube induction medium for the identification of Candida albicans
Authors: Berardinelli S , Opheim D J
Published: January 2020

A new germ tube induction medium, composed of three parts Rabbit Coagulase Plasma with EDTA (BBL Microbiology Systems) and two parts Tryp-Soy broth (Scott Laboratories, Inc.), was effective for the presumptive identification of Candida albicans. This medium was safer to use, more accurate, and less expensive than other commercial germ tube induction media.
Serum bactericidal activity against Legionella pneumophila
Authors: Plouffe J F , Para M F , Fuller K A
Published: January 2020

Two strains of Legionella pneumophila serogroup 1 (UH1 and RH1) were incubated in fresh human serum. The UH1 strain was serum resistant, whereas the RH1 strain was serum susceptible. The bactericidal activity of fresh serum was abrogated by heating. Serum resistance of L. pneumophila strains may correlate with increased virulence.
Effect of swab type and storage temperature on the isolation of Chlamydia trachomatis from clinical specimens
Authors: Mahony J B , Chernesky M A
Published: January 2020

We evaluated various swabs for the recovery of Chlamydia trachomatis and found that only 7 of 14 swabs tested resulted in a greater than 50% recovery of organisms. Storage of chlamydial specimens at 4 degrees C for more than 1 day resulted in a rapid loss of viability, whereas freezing of specimens at -70 degrees C resulted in only a marginal decrease in the isolation rate.
Association of slime with pathogenicity of coagulase-negative staphylococci causing nosocomial septicemia
Authors: Ishak M A , Gröschel D H , Mandell G L , Wenzel R P
Published: January 2020

To assess the role of slime in the pathogenesis of nosocomial bloodstream infections caused by coagulase-negative staphylococci, we compared the characteristics of 27 nosocomial bloodstream isolates with those of 27 skin isolates from non-hospital personnel. Of 27 bloodstream isolates, 14 were judged to be significant by a clinical index, and 13 were contaminants. Slime production was observed in 13 of 14 significant isolates but in only 3 of 13 contaminants (P = 0.0003) and 4 of 27 skin isolates (P = 0.0001). The 14 pathogens were identified as Staphylococcus epidermidis. Only 7 of 13 contaminants and 9 of 27 skin isolates belonged to the same species (P less than 0.006). Slime-producing strains of S. epidermidis represented 13 of 14 pathogens but only 2 of 13 contaminants (P less than 0.0003). Neither adherence to Teflon catheters nor phagocytosis and killing of coagulase-negative staphylococci by polymorphonuclear leukocytes was significantly influenced by slime production. Nevertheless, the identity of the organism and the slime production test predicted the clinical significance of blood isolates of coagulase-negative staphylococci with an overall accuracy of 89%.
Method for the isolation of highly purified Salmonella flagellins
Authors: Ibrahim G F , Fleet G H , Lyons M J , Walker R A
Published: January 2020

Ten different Salmonella serotypes were grown in a chemically defined medium supplemented with 0.01% yeast extract. After sedimentation of the cells by centrifugation, flagella were detached by exposure to pH 2 for 30 min at room temperature. The flagellaless cells were removed by centrifugation, and the flagellin in the supernatant was further purified by high-speed centrifugation, ammonium sulfate precipitation, and dialysis in 50,000-molecular-weight-cutoff tubing. The 10 flagellin preparations were of a high degree of purity, as demonstrated by electron microscopy, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and measurement of salmonella H and O agglutination titers of antisera raised in rabbits with the flagellin preparations as immunogens.
Nosocomial Pseudomonas pickettii colonization associated with a contaminated respiratory therapy solution in a special care nursery
Authors: McNeil M M , Solomon S L , Anderson R L , Davis B J , Spengler R F , Reisberg B E , Thornsberry C , Martone W J
Published: January 2020

Pseudomonas pickettii caused respiratory tract colonization in five infants in the special care nursery of a Chicago hospital. All organisms had the same antimicrobial susceptibilities. Endotracheal suctioning with saline from 5-ml unit-dose vials was identified by epidemiologic investigation as a risk factor for colonization. The vials of saline were contaminated with a strain of P. pickettii having the same antimicrobial susceptibility pattern as the isolates from patients. As part of an investigation of the manufacturing plant where the saline solution was produced, P. pickettii was recovered from deionized water used to make the product and from several sites in the processing line. Bypassing of a 180 degrees F (ca. 82 degrees C) water-holding tank appeared to be temporally related to product contamination. The ability of P. pickettii to survive and grow in this solution has been demonstrated in the laboratory. This outbreak demonstrates that, despite pertinent Food and Drug Administration regulations and company programs for identifying such contamination, intrinsically contaminated solutions can occasionally reach the bedside of the patient.
Effect of heat and fresh human serum on the infectivity of human T-cell lymphotropic virus type III evaluated with new bioassay systems
Authors: Harada S , Yoshiyama H , Yamamoto N
Published: January 2020

MT-4 cells, which are a human T-cell lymphotropic virus type I (HTLV-I)-positive cell line highly permissive to HTLV-III infection, were used to detect the biologically active virus. For quantitation of the virus, induction of HTLV-III-specific antigen(s) and inhibition of DNA synthesis in infected MT-4 cells were assessed by indirect immunofluorescence and by a proliferation assay measuring [3H]thymidine uptake, respectively. HTLV-III was fully inactivated by treatment at 56 degrees C for 30 min. It was not inactivated by treatment with fresh anti-HTLV-III-negative serum. Thus, these assay systems with MT-4 cells would be useful in further studies on acquired immune deficiency syndrome.
Mode of growth of bacterial pathogens in chronic polymicrobial human osteomyelitis
Authors: Marrie T J , Costerton J W
Published: January 2020

Direct examination of material from two cases of persistent (2 and 60 years) osteomyelitis by morphological and culture techniques showed that the pathogens comprised several bacterial species whose cells grew predominantly in discrete exopolysaccharide-enclosed microcolonies made up of a single bacterial morphotype. Bacterial microcolonies were seen between tissue elements in infected connective tissue, and the microcolonies adherent to bone surfaces coalesced to form extensive biofilms that occluded the surfaces of dead bone in sequestrae. Decalcification techniques were required to examine the interior of infected bone, but recognizable remnants were associated with very large amounts of fibrous, ruthenium red-stained material. All bacterial growth in these persistent infections occurred within an intercellular matrix, and some elements of this matrix, which was fibrous in transmission electron microscopy and amorphous in scanning electron microscopy, were associated with the surfaces of bacterial cells in a manner that suggested their production by these organisms. All of the implications of this microcolony mode of bacterial growth in osteomyelitis, and in other chronic bacterial diseases, have yet to be determined.
Characterization of Haemophilus spp. isolated from healthy swine and evaluation of cross-reactivity of complement-fixing antibodies to Haemophilus pleuropneumoniae and Haemophilus taxon "minor group"
Authors: Rapp V J , Ross R F , Young T F
Published: January 2020

Of 30 sows from a herd believed to be free of Haemophilus pleuropneumoniae infection, 2 had complement-fixing antibodies to H. pleuropneumoniae serotype 5. Necropsy and microbiological examination of the two sows revealed no evidence of H. pleuropneumoniae infection; however, Haemophilus taxon "minor group" and a urease-negative, indole-positive Haemophilus sp. were isolated from numerous respiratory tract sites in both sows. Isolation of these Haemophilus spp. was facilitated by serially diluting specimens in two broth media. Pigs from a closed, respiratory disease-free herd were inoculated with four strains of Haemophilus taxon "minor group" to determine whether the organism induces antibodies which cross-react with H. pleuropneumoniae in the complement fixation test. Antigenic heterogeneity among the taxon "minor group" strains was apparent; however, antibodies cross-reacting between these strains and H. pleuropneumoniae serotypes 1 through 5 and 7 were not detected.
Simple visual assay for determination of Pasteurella haemolytica cytotoxin neutralizing antibody titers in cattle sera
Authors: Gentry M J , Confer A W , Kreps J A
Published: January 2020

A simple visual assay is described for determining the capacity of bovine serum to neutralize the cytotoxin produced by Pasteurella haemolytica serotype 1. The test was reproducible from day to day with different target cell populations and cytotoxin preparations. Cytotoxin neutralization titers obtained by the visual assay were comparable to those determined by the trypan blue exclusion and 51Cr-release methods. The visual assay was used to measure neutralization titers of bovine sera obtained from vaccination experiments and fractions of purified serum obtained by gel filtration. The major advantages of the visual assay over other assays are that it is rapid, inexpensive, and does not use radioisotopes. It also does not require specialized equipment, making it adaptable to most laboratories.
Evaluation of the RapID-ANA system for identification of anaerobic bacteria of veterinary origin
Authors: Adney W S , Jones R L
Published: January 2020

This study evaluated the ability of the RapID-ANA system (Innovative Diagnostic Systems, Inc., Atlanta, Ga.) to accurately identify a spectrum of freshly isolated veterinary anaerobes. A total of 183 isolates were tested and included 7 Actinomyces spp., 53 Bacteroides spp., 32 Clostridium spp., 2 Eubacterium spp., 65 Fusobacterium spp., 1 Peptococcus spp., 22 Peptostreptococcus spp., and 1 Propionibacterium spp. All isolates were initially identified by conventional biochemical testing and gas-liquid chromatography of short-chain fatty acid metabolites. Additional tests were performed as required by the RapID-ANA system. Of these isolates, 81.4% were correctly identified to the genus level, including 59.6% to the species level, 14.2% were incorrectly identified at the genus level, and 4.4% were not identified. Initially, 20.2% of the strains were not identified because the microcodes were not in the code book. The majority of the incorrect identifications were caused by the misidentification of Fusobacterium spp. as Bacteroides spp. Errors also occurred when veterinary anaerobes not included in the data base were assigned an identification from the existing data base. The RapID-ANA system appears to be a promising new method for rapid identification of veterinary anaerobes; however, further evaluation with an extended data base is needed before the system can accurately identify all clinically significant anaerobes.
Rapid dot-immunobinding assay on nitrocellulose for viral antibodies
Authors: Heberling R L , Kalter S S
Published: January 2020

A procedure is described for the routine laboratory diagnosis of viral serum antibodies. Antigens are dotted on nitrocellulose strips or sheets, and sera are applied on absorbent paper strips. Antigen-antibody complexes are detected with enzyme-conjugated antiglobulin and development of a colored, insoluble substrate product. The test allows processing of multiple sera in one 3- to 5-h operation and is equal to or more sensitive than serum neutralization, hemagglutination inhibition, and fluorescent antibody assays. Highly infectious viruses inactivated with a psoralen derivative and long-wavelength UV light irradiation can be used as antigens, allowing the study of human pathogens. Although the test detects cross-reacting, group-specific herpesvirus antigens, the intensity of the antibody reaction is greatest with type-specific antigens. Preliminary data suggest that the technique will be useful for the rapid typing of viruses from clinical specimens.
Comparison of the MicroScan system with the API Staph-Ident system for species identification of coagulase-negative staphylococci
Authors: Hussain Z , Stoakes L , Stevens D L , Schieven B C , Lannigan R , Jones C
Published: January 2020

To evaluate the accuracy of the MicroScan System (American Hospital Supply Corp., Sacramento, Calif.) for identification of coagulase-negative staphylococci, we tested 175 clinical isolates of coagulase-negative staphylococci. The results obtained by the MicroScan system were compared with those of the API Staph-Ident system (Analytab Products, Plainview, N.Y.). Forty-three discrepancies between the two systems were resolved by the conventional method of Kloos and Schleifer (W.E. Kloos and K.H. Schleifer, J. Clin. Microbiol. 1:82-88, 1975). The MicroScan and the Staph-Ident systems correctly identified 146 (86.4%) and 154 (88%) of 175 strains, respectively. The API system failed to identify phosphatase-negative Staphylococcus epidermidis. The MicroScan system demonstrated the greatest accuracy in the identification of S. epidermidis and S. saprophyticus, whereas lesser accuracy was achieved with S. hominis, S. warneri, and S. sciuri.
Evaluation of the MicroScan antimicrobial susceptibility system with the autoSCAN-4 automated reader
Authors: Baker C N , Stocker S A , Rhoden D L , Thornsberry C
Published: January 2020

The American MicroScan (American MicroScan, Mahwah, N.J.) identification and antimicrobial susceptibility system consists in part of an automated reading system (autoSCAN-4) with data management capabilities. We evaluated the system with 404 gram-negative and 170 gram-positive facultative anaerobic and aerobic bacteria. We compared MicroScan results read automatically and visually with each other and with the results obtained by the reference method (read visually). The overall agreement within +/- 1 log2 dilution was 94.3% when the MicroScan system (read automatically) was compared with the reference method (read visually), 96.4% when MicroScan panels (read visually) were compared with reference panels, and 97.4% when the autoSCAN-4 automated reading was compared with the visual reading of the MicroScan panels. Total discrepancies (susceptibility interpretation category changes) for the MicroScan system compared with the reference method were 7%, with 6.2% considered a minor discrepancy. The autoSCAN-4 and the complete MicroScan system yielded accurate results compared with the reference method.
Ureaplasma urealyticum serotypes in urinary tract disease
Authors: Hewish M J , Birch D F , Fairley K F
Published: January 2020

Ureaplasma urealyticum cultures from 124 patients with urinary tract disease were serotyped by indirect immunofluorescence, using antisera to serotypes I to VIII. A similar range of serotypes was recovered from first-voided, midstream, and bladder-aspiration (SPA) urine, upper urinary tract samples, and vaginal swabs. Serotype VI was predominant (44/124) among the samples, whereas serotypes V (1/124 samples) and VII (0/124 samples) were uncommon. Twenty of 124 cultures contained more than one serotype, and three cultures were untypeable. Serotypes cultured from bladder urine were also present in vaginal and urethral samples, although these samples often carried additional serotypes. Consecutive SPA samples from the same patient invariably contained the same serotype, whereas some consecutive midstream urine samples showed a loss or gain of serotypes with time. One patient carried the same serotype in SPA urine over a period of 13 months. The pattern of serotypes recovered from the urinary tract was similar irrespective of the sampling site, the site of infection, the clinical diagnosis and renal function of the patient, and the presence or absence of other microorganisms. Colonization above the urethra and association with urinary tract disease appeared to be serotype independent.
Inability of the Chemstrip LN compared with quantitative urine culture to predict significant bacteriuria
Authors: Jones C , MacPherson D W , Stevens D L
Published: January 2020

The Chemstrip LN (Boehringer Mannheim Biochemicals, Indianapolis, Ind.), designed to detect pyuria and bacteriuria, was compared with culture of 1,020 unselected, consecutive midstream urine specimens and evaluated on its ability to predict colony counts at three levels. At the level of greater than or equal to 10(5) CFU/ml, the combined test (detection of leukocyte esterase and nitrite) had sensitivity of 82.3%, specificity of 67.9%, positive predictive value of 41.3%, and negative predictive value of 93.3% at prevalence rate of 21.6%. The test would have rejected 9.4% of the specimens with significant bacteriuria if the Chemstrip alone had been used.
Growth of Neisseria gonorrhoeae on Campylobacter medium at 40 degrees C
Authors: Johnson P
Published: January 2020

As the result of finding Neisseria gonorrhoeae in a rectal culture for Campylobacter spp., a study was done to determine if this was an unusual isolate. Five gonococcal isolates grew equally well on modified Thayer-Martin medium in a microaerobic chamber whether incubated at 35 or 40 degrees C. All five grew as well as controls microaerobically on campylobacter medium containing lysed horse blood at 35 degrees C; three of the five grew on this medium at 40 degrees C.
Influence of patient hemogram on growth index values generated from a radiometric detection device
Authors: Berger S A , Dan M , Battat A , Kahn Y
Published: January 2020

Blood samples from 20 patients with widely varying concentrations of erythrocytes and leukocyte subtypes were inoculated into BACTEC 6B bottles (Johnston Laboratories, Inc., Towson, Md.). There was no relationship between growth index value and hemogram. Although sterile blood is capable of generating small amounts of 14CO2, the mechanism for this phenomenon is not related to the concentration of a specific type of blood cell.
Enhancement of Bacteroides intermedius growth by Fusobacterium necrophorum
Authors: Price S B , McCallum R E
Published: January 2020

Previous work from this laboratory has demonstrated the persistence of Bacteroides intermedius in the livers of mice receiving an intraperitoneal inoculum of B. intermedius and Fusobacterium necrophorum. This study was undertaken to determine whether F. necrophorum enhanced the in vitro growth of B. intermedius. Tryptose phosphate broth did not support the growth of B. intermedius alone, but the bacterium did survive in a tryptose phosphate broth culture of F. necrophorum. B. intermedius cultured in F. necrophorum-conditioned tryptose phosphate broth grew impressively, reaching maximal absorbance at 24 h after inoculation. The growth of B. intermedius in F. necrophorum-conditioned tryptose phosphate broth was proportional to the amount of conditioned medium present. The B. intermedius growth-stimulating factor was detectable in conditioned medium 8 h after inoculation with F. necrophorum and could be detected throughout the 96-h incubation period. Growth-factor-active fractions eluted from a Sephadex G-100 column did not absorb at 280 nm and were retained on the column until 4 column volumes were eluted. The growth factor was nondialyzable and stable to boiling, lyophilization, extraction with hot aqueous phenol, and trypsin digestion. The factor was inactivated by exposure to pH 2.0 in the pepsin digestion protocol. Significant amounts of hexose, methyl pentose, and 2-keto-3-deoxyoctonate were detected in pooled growth-factor-active fractions eluted from the Sephadex column. This pool was also active in the Limulus lysate endotoxin assay. These results suggest that the B. intermedius growth-stimulating factor produced by F. necrophorum is a lipopolysaccharide.
Fluorescent anti-immunoglobulin G assay of circulating immune complexes extracted with polyethylene glycol
Authors: Levinson S S , Goldman J
Published: January 2020

After a polyethylene glycol extraction which removes monomeric immunoglobulin G (IgG), circulating immune complexes were assayed by the FIAX fluorescence assay (Whittaker M. A. Bioproducts, Walkersville, Md.). The extraction portion of the procedure can be completed within a few hours after an overnight precipitation step, and the fluorescence assay takes about 0.5 h. The fluorescence assay is similar to the routine FIAX method for measuring IgG in cerebrospinal fluid except that a 10-microliter sample from sera is substituted for a 50-microliter sample. Good parallelism between endogenous immune complexes, monomeric IgG, and aggregated human globulin, along with good between-run precision (coefficients of variation, less than 10%), indicates that monomeric IgG calibrators from the kit could be used to standardize the assay. This standardization eliminates the need for aggregated human globulin, which is unstable and difficult to prepare. A reference range of 9 to 63 mg/liter followed a gaussian distribution. Correlation data indicate that the test provides information similar to the C1q-binding test (rho = 0.87; n = 30) but has better diagnostic sensitivity and better discrimination in the high range. Because of its simplicity, good reproducibility, and accessibility to equipment available in many laboratories, the method described here may be a preferred technique for measuring circulating immune complexes.
Immunoblot analyses of Candida albicans-associated antigens and antibodies in human sera
Authors: Manning-Zweerink M , Maloney C S , Mitchell T G , Weston H
Published: January 2020

We tested 10 patient sera for the presence of immunoglobulin G (IgG) antibodies to Candida albicans and for C. albicans antigens by immunoblot analysis (i.e., electrotransfer blot radioimmunoassay) (G. E. Smith and M. D. Summers, J. Virol. 39:125-137, 1981). We evaluated sera from two patients at risk for candidiasis, five patients with systemic candidiasis documented by culture, and two patients who had experienced transient candidemia. Both the specificity and the relative amount of IgG antibodies to C. albicans in each serum sample were readily visualized by this technique, as was the absence of antibody from serum of neonatal and immunocompromised patients. No antibody species appeared to be uniquely associated with candidiasis patients (i.e., each antibody species present in the candidiasis patient was also present in sera of normal individuals or "at-risk" patients). IgG from rabbits immunized with whole cells or with a cytoplasmic fraction of C. albicans was used to detect C. albicans antigens in patient sera. Although several antigens were detected in the sera from patients with candidiasis, the same antigens were also detected in sera from patients at risk and in normal human serum. No antigens were detected in human serum when preimmune rabbit sera were used. These results suggest that the antigens detected by the rabbit antisera were human serum proteins that cross-reacted with C. albicans antigens. These findings may have important implications in studies of both the pathobiology of C. albicans and the serodiagnosis of candidiasis.