bluuebunny/crossref_metadata_2025 huggingface dataset:
Emergence of a new opportunistic pathogen, Candida lusitaniae
Authors: Blinkhorn R J , Adelstein D , Spagnuolo P J
Published: January 2020

Candida lusitaniae has been an infrequently reported opportunistic pathogen. Most previously reported cases of serious infection caused by this organism have proven fatal and were associated with amphotericin resistance of the organism. We report two patients with hematologic malignancies undergoing cytotoxic chemotherapy who developed fungemia with this organism while they were granulocytopenic. The organisms isolated from each patient were fully susceptible and were treated successfully with amphotericin B. When isolated from an immunocompromised host, C. lusitaniae should be considered an opportunistic pathogen and undergo antifungal susceptibility testing. Amphotericin B should be considered the drug of choice, but a poor clinical response may be indicative of a resistant isolate.
Elevated soluble CD8 levels in sera of human immunodeficiency virus-infected populations
Authors: Reddy M M , Lange M , Grieco M H
Published: January 2020

Soluble CD8 levels in sera were quantitated in asymptomatic intravenous drug abusers, homosexuals, and patients with lymphadenopathy or acquired immunodeficiency syndrome. Soluble CD8 levels were elevated in human immunodeficiency virus-seronegative intravenous drug abusers and homosexuals, probably reflecting infections like cytomegalovirus. The sera of human immunodeficiency virus-seropositive groups of patients with human immunodeficiency virus infection also had elevated levels of soluble CD8, reflecting infections like cytomegalovirus and human immunodeficiency virus infection.
Detection of enterotoxigenic Escherichia coli after polymerase chain reaction amplification with a thermostable DNA polymerase
Authors: Olive D M
Published: January 2020

The direct identification of enterotoxigenic Escherichia coli from clinical specimens was examined by using the polymerase chain reaction (PCR) for amplifying the heat-labile toxin (LT) gene. Two synthetic primers, each of which was 20 bases in length, were used with the thermostable DNA polymerase from Thermus aquaticus to amplify the LT gene. The amplified PCR products were detected by either gel electrophoresis or hybridization to a 24-base synthetic oligonucleotide probe conjugated to alkaline phosphatase. The PCR method detected LT-positive bacteria but did not react with E. coli producing the heat-stable toxin, enteroinvasive E. coli, Salmonella typhi, Salmonella typhimurium, or Shigella dysenteriae. By the PCR method, a single bacterium could be detected following 30 cycles of amplification. The T. aquaticus DNA polymerase was inhibited by more than 10(3) organisms in the amplification reaction mixture. A group of 40 clinical specimens consisting of 16 LT bioassay-positive and 24 LT bioassay-negative stool specimens were tested by PCR for the presence of toxigenic E. coli. The total DNA from 100 microliters of stool specimen was extracted and partially purified with a commercially available ion-exchange column. All 16 of the bioassay-positive stool specimens were positive by PCR. In addition, one stool specimen which was bioassay negative for LT but positive for LT in a previous hybridization assay with a different LT probe was also positive by PCR. This may indicate that the LT gene is present but either is not expressed or is expressed below detectable levels. Amplification of specific DNA sequences by PCR provides a highly sensitive and specific tool for the detection of pathogenic microorganisms directly from clinical specimens without the need for prior isolation. This technique may find wide application in the detection of other organisms in addition to enterotoxigenic E.coli.
Recovery of Pseudomonas gladioli from respiratory tract specimens of patients with cystic fibrosis
Authors: Christenson J C , Welch D F , Mukwaya G , Muszynski M J , Weaver R E , Brenner D J
Published: January 2020

Pseudomonas gladioli was isolated from 11 patients with cystic fibrosis. It resembled Pseudomonas cepacia on the selective and differential medium OFPBL, producing yellow colonies after 48 to 72 h of incubation. Isolates were characterized biochemically, by DNA hybridization, and by cellular fatty acid analysis. A review of the clinical status of selected patients colonized by P. gladioli did not reveal any apparent association of this organism with infectious complications of cystic fibrosis. Thus, the clinical implications may differ depending on which of these two closely related species is reported by laboratories. Determination of the fatty acid profile of isolates by gas chromatography may be a useful adjunct to biochemical characterization as a means of identification. In contrast to P. cepacia, most isolates of P. gladioli contained 3-OH C10:0 fatty acid under the growth conditions used.
Immunoblot analysis of antibody responses to Sporothrix schenckii
Authors: Scott E N , Muchmore H G
Published: January 2020

The serologic response to Sporothrix schenckii was investigated in patients with sporotrichosis by solid-phase enzyme-linked immunosorbent assays (ELISAs) and Western immunoblot techniques. A soluble antigen preparation derived from an S. schenckii isolate contained 15 protein staining components ranging in molecular size from 22 to 70 kilodaltons (kDa) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Sera from 40 patients with sporotrichosis demonstrated Sporothrix immunoglobulin G antibody by ELISA with titers between 128 and 65,200. No sera from 300 healthy individuals or 100 patients with various systemic mycoses other than sporotrichosis had ELISA titers greater than 64. By Western immunoblotting of the antigens separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, sera from 10 patients with cutaneous sporotrichosis reacted with 8 to 10 antigen components (range, 40 to 70 kDa), while sera from 15 patients with extracutaneous sporotrichosis reacted with a greater number of antigen components (15 to 20 bands) over a wider range of molecular sizes (22 to 70 kDa). Antibody to 40- and 70-kDa antigen components was detected by immunoblots in all sera tested from patients with sporotrichosis. Antibody to 22- to 36-kDa antigen components was present in sera from 13 of 15 patients with extracutaneous sporotrichosis, but these lower-molecular-weight components were not detected by sera from patients with cutaneous sporotrichosis. Antibody to these components was not detected by Western blotting in sera from 19 of 20 patients with other fungal diseases or from 30 healthy individuals. Purification of these specific antigen fractions could provide the basis of a sensitive and specific serodiagnostic test to indicate the presence and activity of extracutaneous sporotrichosis.
Bacteremia with Streptococcus bovis and Streptococcus salivarius: clinical correlates of more accurate identification of isolates
Authors: Ruoff K L , Miller S I , Garner C V , Ferraro M J , Calderwood S B
Published: January 2020

Two biotypes of Streptococcus bovis can be identified by laboratory testing and can be distinguished from the phenotypically similar organism Streptococcus salivarius. We assessed the clinical relevance of careful identification of these organisms in 68 patients with streptococcal bacteremia caused by these similar species. S. bovis was more likely to be clinically significant when isolated from blood (89%) than was S. salivarius (23%). There was a striking association between S. bovis I bacteremia and underlying endocarditis (94%) compared with that of S. bovis II bacteremia (18%). Bacteremia with S. bovis I was also highly correlated with an underlying colonic neoplasm (71% of patients overall, 100% of those with thorough colonic examinations) compared with bacteremia due to S. bovis II or S. salivarius (17% overall, 25% of patients with thorough colonic examinations). We conclude that careful identification of streptococcal bacteremic isolates as S. bovis biotype I provides clinically important information and should be more widely applied.
Unexpected isolation of Bordetella pertussis from patients with acquired immunodeficiency syndrome
Authors: Ng V L , York M , Hadley W K
Published: January 2020

We isolated Bordetella pertussis from three patients with acquired immunodeficiency syndrome who underwent diagnostic bronchoscopy for evaluation of respiratory symptoms. The B. pertussis isolates were recovered from medium (charcoal-yeast extract agar) formulated to enhance recovery of Legionella spp., and one of the isolates stained positively with antisera directed against Legionella spp.
Recovery of Mycobacterium avium-M. intracellulare from blood specimens by using the routine BACTEC 6B blood culture system
Authors: Horn R , Laszlo A , Robson H G
Published: January 2020

We describe a simple technique for recovery of Mycobacterium avium-M. intracellulare from blood culture specimens by using the BACTEC 460, a system used for routine blood cultures in many hospital laboratories. A total of 26 of 215 blood specimens (12%) from 11 of 48 patients (23%) with acquired immunodeficiency syndrome (AIDS) or AIDS-related complex yielded isolates of M. avium-M. intracellulare. Acid-fast staining of prepared specimens was positive for 16 of 26 cultures that ultimately yielded the organism.
D-lactic acid measurements in the diagnosis of bacterial infections
Authors: Smith S M , Eng R H , Campos J M , Chmel H
Published: January 2020

Body fluids suspected of bacterial infection were cultured and examined for the presence of D-lactic acid, a specific bacterial metabolite. We examined 206 patients and 264 specimens. D-Lactic acid was found in concentrations of greater than or equal to 0.15 mM in 11 of 11 infected and 6 of 40 noninfected ascitic fluids, 6 of 6 infected and 4 of 33 noninfected pleural fluids, 4 of 4 infected and 0 of 13 noninfected synovial fluids, and 26 of 27 infected and 2 of 130 noninfected cerebrospinal fluids. The overall sensitivity was 79.7%, and the specificity was 99.5% when the D-lactic acid concentration was at least 0.15 mM. The most important clinical utility of the D-lactic acid measurement appears to be for patients with bacterial infection in various body compartments and in patients who have already received antimicrobial therapy. An elevation in D-lactic acid may indicate the presence of bacterial infection even when cultures are negative.
T-lymphocyte clones responsive to Shigella flexneri
Authors: Zwillich S H , Duby A D , Lipsky P E
Published: January 2020

T lymphocytes from a patient with Shigella flexneri dysentery and postdysenteric reactive arthritis were cloned by limiting dilution with recombinant interleukin-2 and a strain of S. flexneri different from that which had infected her. Five of eight clones produced proliferated in response to the shigellae used to generate the clones. The response required irradiated syngeneic blood mononuclear cells as antigen-presenting cells. One such clone, MC12, proliferated in response to both the shigellae used to generate the clones and the infecting shigellae but not to other shigellae, Salmonella heidelberg, or control Escherichia coli. MC12 was CD3+, CD4+, CD8-, and human histocompatibility leukocyte antigen (HLA)-DR+. The proliferative response to the shigellae was blocked by antibody to HLA-DR but not by antibody to HLA-A,B,C. The response required antigen-presenting cells that shared HLA-DR antigens with the clone and appeared to be restricted by HLA-DR2. The epitope recognized by MC12 was associated with the bacterial membranes. Thus, T-lymphocyte clones that proliferate in response to some shigellae can be isolated from patients with shigellosis.
Detection of group B rotavirus in fecal specimens by dot hybridization with a cloned cDNA probe
Authors: Eiden J J , Firoozmand F , Sato S , Vonderfecht S L , Yin F Z , Yolken R H
Published: January 2020

Cloned cDNA copies were synthesized from the genomic RNA of the IDIR strain of group B rotavirus (GBR) isolated in Baltimore, Md. These clones were screened for hybridization with heterologous GBR to identify cDNA for use in dot hybridization experiments. In multiple screening experiments, cDNA clones derived from gene segment 3 provided the most intense hybridization signals. 32P-labeled probes were produced from one of the gene 3 clones, and these were employed in dot hybridization assays. Purified preparations of bovine GBR were detected in concentrations of greater than or equal to 0.5 ng, and GBR was detected in fecal specimens obtained from five of six infected calves. Four of six human fecal specimens containing the Baltimore strain of GBR were also positive in the hybridization assay, while GBR was identified in only one of the six specimens by means of immunoelectron microscopy. A fecal specimen obtained from a patient infected with the adult diarrhea rotavirus strain of GBR was also positive in the dot hybridization assay. Fecal specimens from uninfected humans, calves, and rats, as well as specimens containing group A rotaviruses, did not hybridize with the cloned cDNA probe.
Diagnosis of Giardia lamblia infections by detection of parasite-specific antigens
Authors: Janoff E N , Craft J C , Pickering L K , Novotny T , Blaser M J , Knisley C V , Reller L B
Published: January 2020

Antigen detection methods may facilitate diagnosis of Giardia lamblia in stool specimens. As determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and immunoblotting, G. lamblia cysts and trophozoites share several antigens, especially in the 65-kilodalton and 30- to 34-kilodalton regions. By using blind methods, we compared results obtained by counterimmunoelectrophoresis using cyst-immune rabbit serum and by enzyme-linked immunosorbent assay (ELISA) using trophozoite-immune rabbit serum with results obtained by microscopic examination of a preserved, concentrated, and permanently stained stool specimen. Results were similar when these three methods were used to examine 118 stool specimens from clinical microbiology laboratories (53 specimens with G. lamblia) and specimens from 239 day-care-center toddlers (39 specimens with G. lamblia). Compared with microscopy, we found, for counterimmunoelectrophoresis and ELISA, respectively: sensitivity, 88 versus 94%; specificity, 97 versus 95%; positive predictive value, 86 versus 76%; negative predictive value, 98 versus 97%; and concordance, 89%. The false-positive rate by ELISA was 24% (10 of 42) in day-care-center toddlers but only 3% (1 of 32) in healthy adults (P less than 0.04) as corroborated by microscopy. This discrepancy suggests that the ELISA may be more sensitive than microscopy, which is considered the reference standard, and that results may be dependent, in part, on the epidemiology of the infection in the study subjects.
Ultrastructure and chemical analysis of Campylobacter pylori flagella
Authors: Geis G , Leying H , Suerbaum S , Mai U , Opferkuch W
Published: January 2020

Flagella of Campylobacter pylori were analyzed by electron microscopy and purified, and the molecular weight of the flagellin was determined. Isolation of flagella was performed by mechanical shearing from the cell surface, sucrose density gradient centrifugation, and Sepharose CL-4B gel chromatography. The flagella of C. pylori differ from those of other Campylobacter species and of most other bacteria by the presence of a flagellar sheath. The sheath narrows at the end and is linked to a club-shaped terminal structure. The molecular weight of C. pylori flagellin was 51,000.
Dot blot hybridization assay of B19 virus DNA in clinical specimens
Authors: Mori J , Field A M , Clewley J P , Cohen B J
Published: January 2020

A nonradioactive dot blot hybridization assay for human parvovirus B19 DNA was set up by using a biotin-labeled DNA probe and streptavidin-alkaline phosphatase conjugate. The assay was used to examine 4,895 specimens referred for B19 virus diagnosis during 1987. Of 48 specimens that gave positive reactions for B19 DNA, 41 were confirmed virus positive by electron microscopy (n = 36), radioimmunoassay (n = 26), or counterimmunoelectrophoresis (n = 20). In 7 samples which were not confirmed and in 11 samples giving weak reactions for B19 DNA, there was serological or epidemiological evidence of recent B19 infection. A further 70 specimens gave weak, apparently false-positive reactions. By electron microscopy, 13 of 16 were contaminated by bacteria, and plasmid DNA was demonstrated in one specimen. Of 55 specimens tested, 52 reacted with streptavidin-alkaline phosphatase conjugate alone. These were probable sources of nonspecificity in an otherwise practical and economic screening method for B19 virus.
Cellular fatty acid compositions and isoprenoid quinone contents of 23 Legionella species
Authors: Lambert M A , Moss C W
Published: January 2020

The cellular fatty acid compositions and ubiquinone contents of 182 Legionella strains representing 23 species were determined by capillary gas-liquid chromatography and reverse-phase high-performance liquid chromatography, respectively. Except for the type strain of Legionella erythra (ATCC 35303T), all Legionella species contained large (40 to 90%) amounts of branched-chain fatty acids and only trace to small (less than 0.5 to 5%) amounts of ester-linked hydroxy acids. The 23 species were placed in three major fatty acid groups on the basis of differences in the relative amounts of 14-methylpentadecanoic (Ci16:0), hexadecanoic (C16:1), and 12-methyltetradecanoic (Ca15:0) acids. All Legionella species contained ubiquinones with 9 to 14 isoprene units in the side chains and were divided into five different ubiquinone groups. The species were further differentiated into 16 groups on the basis of qualitative and quantitative differences in their fatty acid compositions and ubiquinone contents. Both of these chemical characteristics can be used to distinguish Legionella species from other gram-negative bacteria and rapidly and accurately identify suspected isolates before serologic and other tests are done.
Comparative evaluation of mitogenicity and basement-membrane-degrading activity of Pseudomonas aeruginosa slime glycolipoprotein and alginate
Authors: Anastassiou E D , Karakiulakis G , Missirlis E , Maragoudakis M E , Dimitracopoulos G
Published: January 2020

Alginate from a heavily mucoid Pseudomonas aeruginosa strain and slime glycolipoprotein obtained from the revertant nonmucoid variant of the mucoid strain were tested for mitogenic activity on human peripheral lymphocytes and for degradation of 3H-labeled basement membranes of the anterior lens capsule of bovine eyes. Slime glycolipoprotein exerted mitogenic activity in concentrations from 50 to 200 micrograms/ml, whereas alginate was not mitogenic as shown by [3H]thymidine uptake. Alginate did not show any basement membrane degradation, whereas slime glycolipoprotein exhibited basement-membrane-degrading activity from 35 to 450 micrograms/ml in a dose-related manner. This activity was inhibited by metal chelators but not thiol protease inhibitors. The results suggest that alginate lacks the mitogenic and biodegrading activities of slime glycolipoprotein; these activities nevertheless need further investigation.
Synthetic exfoliative toxin A and B DNA probes for detection of toxigenic Staphylococcus aureus strains
Authors: Rifai S , Barbancon V , Prevost G , Piemont Y
Published: January 2020

Two methods for the detection of exfoliative toxin (ET) from Staphylococcus aureus were compared: (i) a phenotypic assay, electrosyneresis, and (ii) a genotypic assay, staphylococcal DNA hybridization with oligodeoxynucleotide probes. The probes were chosen from the previously determined sequences of serotype A and B of ET, one probe for serotype A and another for serotype B. Strains exhibiting ET production in electrosyneresis always possessed the ET gene(s). Conversely, some strains not exhibiting ET production in electrosyneresis harbored the ET gene(s). The latter strains produced levels of ET. ET-negative phage group 2 strains of S. aureus as well as tested coagulase-negative staphylococci did not possess the ET gene(s). The sensitivity of the DNA hybridization technique was 10(6) bacteria or 100 ng of genomic DNA.
Indirect enzyme-linked immunosorbent assay for immunoglobulin G and four immunoassays for immunoglobulin M to Toxoplasma gondii in a series of heart transplant recipients
Authors: Sluiters J F , Balk A H , Essed C E , Mochtar B , Weimar W , Simoons M L , Ijzerman E P
Published: January 2020

Toxoplasma gondii infections in heart transplant recipients were monitored by indirect enzyme-linked immunosorbent assay for immunoglobulin G (ELISA-IgG), indirect ELISA-IgM in serum IgM fractions, antibody capture ELISA-IgM, IgM-immunosorbent agglutination assay (ISAGA), and IgM immunoblotting. Basic immunosuppression consisted of cyclosporine and low-dose steroids. Before transplantation, 26 of 43 recipients showed serological evidence of infection. In serum samples from 15 (35%) recipients, specific antibodies were not detected. Approximately 50% of the heart donors, were toxoplasma seropositive. Eight of the fifteen seronegative recipients received hearts from toxoplasma-seropositive donors. In four of the eight recipients, seroconversion could be demonstrated with all tests used. In three of these four patients, clinical disease developed. One patient with strong serological evidence of toxoplasmosis died, but toxoplasma parasites and antigens were not detected at autopsy. In two patients, toxoplasma cysts were found in cardiac biopsies. Seroconversion was not prevented by the use of spiramycin prophylaxis in two recipients. Reactivations of latent infections or reinfections were detected by indirect ELISA in six (23%) seropositive recipients, but symptoms and signs of active T. gondii infection were not seen. Seroconversion and reactivation of infection were readily found by a combined use of indirect ELISA-IgG and ELISA-IgM and antibody capture ELISA-IgM. Discrepancies in results could be examined by immunoblotting. IgM-ISAGA retained stable positive values longer than IgM-ELISAs did. Cyclosporine treatment did not hamper detection of seroconversion but could cause antibody levels to remain relatively low in primary infections. Seronegative recipients should receive antitoxoplasma treatment on seroconversion.
Taxonomic investigation of Legionella pneumophila using monoclonal antibodies
Authors: Brindle R J , Bryant T N , Draper P W
Published: January 2020

A panel of 19 monoclonal antibodies was used to produce patterns of immunofluorescent staining of 468 isolates of Legionella pneumophila. Twelve monoclonal antibodies were selected that divided L. pneumophila into 17 phenons which, in the majority of cases, conform to serogroup divisions. These phenons are more easily defined than the present serogroups, and isolates can be placed in them with little ambiguity. The standardized set of monoclonal antibodies was also used to define the subgroups of serogroup 1.
Serodiagnosis of erythema migrans and acrodermatitis chronica atrophicans by the Borrelia burgdorferi flagellum enzyme-linked immunosorbent assay
Authors: Hansen K , Asbrink E
Published: January 2020

The diagnostic performance of an enzyme-linked immunosorbent assay (ELISA) using purified Borrelia burgdorferi flagella as test antigen was compared with that of a B. burgdorferi sonic extract ELISA. We tested sera from 200 healthy controls, 107 patients with erythema migrans (EM), 50 patients with acrodermatitis chronica atrophicans (ACA), and 98 patients with various dermatological disorders without clinical evidence of active Lyme borreliosis. The flagellum ELISA was significantly more sensitive than the sonic extract ELISA. With sera from patients with EM, the diagnostic sensitivity for immunoglobulin G (IgG) antibody detection increased from 11.2 to 35.5% (P less than 0.001) and for IgM antibody detection it increased from 16.6 to 44.8% (P less than 0.001). In the flagellum ELISA, the number of positive tests increased significantly (P less than 0.005) when the duration of EM exceeded 1 month, but still only about 50% of patients with longstanding (1 to 12 months) untreated EM were IgG seropositive. Concomitant general symptoms did not affect the antibody level, whereas patients with multiple erythema were more frequently seropositive. All sera from patients with EM which were positive in the sonic extract ELISA were also positive in the flagellum ELISA. Not only did the overall number of positive tests increase, but the flagellum ELISA yielded a significantly better quantitative discrimination between seropositive patients and controls (P less than 0.002). IgG antibodies to the B. burgdorferi flagellum were found in all sera from patients with ACA, indicating persistence of an antiflagellum immune response in late stages of Lyme borreliosis. IgM reactivity in sera from patients with ACA was shown to be unspecific and the result IgM rheumatoid factor. A rheumatoid factor was detected in sera from 32% of patients with ACA, compared with 7.5% of patients with EM. The improved diagnostic performance, the ease of standardization of the flagellum antigen, and the lack of strain variation make the B. burgdorferi flagellum a needed reference antigen for growing routine serology in Lyme borreliosis.
Characterization of virulence marker antigen of Shigella spp. and enteroinvasive Escherichia coli
Authors: Pál T , Formal S B , Hale T L
Published: January 2020

Antisera produced in rabbits immunized with an enteroinvasive O143 strain of Escherichia coli were absorbed with an avirulent derivative of the same strain. The resulting sera have been previously shown to recognize enteroinvasive pathogens when used in an enzyme-linked immunosorbent assay. In the current study, Western blots (immunoblots) showed that such an absorbed rabbit antiserum recognized two proteins (IpaB and IpaC) which are encoded by a large, virulence-associated plasmid. These proteins are the apparent constituents of the virulence marker antigen which is expressed by shigellae and enteroinvasive E. coli.
Synthesis and secretion of phospholipase C by oral spirochetes
Authors: Siboo R , al-Joburi W , Gornitsky M , Chan E C
Published: January 2020

Four strains of oral treponemes and Treponema phagedenis Reiter synthesize and secrete phospholipase C (PLC), which was detected by the hydrolysis of p-nitrophenylphosphorylcholine. PLC was detected in gingival crevicular fluid from diseased but not from healthy sulci. The initiation and progression of periodontal lesions may begin with the hydrolysis of membrane phospholipids by PLC.
Detection of antibodies to caprine arthritis-encephalitis virus by protein G enzyme-linked immunosorbent assay and immunoblotting
Authors: Zanoni R , Krieg A , Peterhans E
Published: January 2020

Sera from goats suffering from caprine arthritis-encephalitis contained antibodies to virus proteins of 15, 17, 28, 40, and 130 kilodaltons in immunoblots of maedi-visna virus. We propose to use immunoblotting as a validation test for enzyme-linked immunosorbent assay and demonstrate that the specificity of indirect enzyme-linked immunosorbent assay can be improved by replacing second antibody by a protein G-avidin-biotin conjugate.
Rapid detection of Bordetella pertussis by a monoclonal antibody-based colony blot assay
Authors: Gustafsson B , Askelöf P
Published: January 2020

Monoclonal antibodies to Bordetella pertussis filamentous hemagglutinin (FHA) and lipopolysaccharide (LPS) were used in a colony blot enzyme-linked immunosorbent assay designed for rapid detection of B. pertussis. Bacterial colonies from Bordet-Gengou agar plates were blotted onto nitrocellulose filter disks, lysed by immersion in chloroform, and reacted with monoclonal antibodies. Following reaction with peroxidase-conjugated rabbit anti-mouse immunoglobulin antisera and 4-chloro-1-naphthol, blue dots representing single colonies appeared on the filters. Blotting of single B. pertussis colonies could be performed after incubation for 40 h, i.e., before the colonies were visible by eye on the agar surface. Ten of ten B. pertussis strains showed positive blotting reactions with antibodies specific for B. pertussis FHA and LPS. Fourteen of fourteen B. parapertussis strains reacted with two of the FHA-specific antibodies but not with two of the LPS-specific antibodies. Strains of B. bronchiseptica showed a variable reaction pattern. No cross-reactions were observed with strains of Streptococcus mitis, S. pyogenes, S. pneumoniae, Staphylococcus aureus, Branhamella catarrhalis, or Klebsiella pneumoniae. This assay may be useful for identification of B. pertussis and B. parapertussis in suspected cases of whooping cough.
Evaluation of a prototype DNA probe test for the noncultural diagnosis of gonorrhea
Authors: Granato P A , Franz M R
Published: January 2020

A prototype, nonisotopic, chemiluminescent DNA probe test called the Gen-Probe PACE (Probe Assay-Chemiluminescence Enhanced) system for Neisseria gonorrhoeae (Gen-Probe, San Diego, Calif.) was compared with conventional Martin-Lewis culture medium in JEMBEC plates for the laboratory diagnosis of gonorrhea. This 2-h noncultural assay is based upon the use of an acridinium ester-labeled DNA probe. The rRNA-directed DNA probe hybridizes with the target rRNA, and the hybridized probe is separated from the unhybridized probe through the use of magnetic microparticles. The esterified acridinium is hydrolyzed from the hybridized probe by the addition of an alkaline hydrogen peroxide solution, resulting in the production of visible light which is measured in a luminometer. The amount of light generated is directly proportional to the amount of gonococcal target rRNA present in the sample. A total of 407 clinical specimens (203 urethral and 204 endocervical) were collected from high-risk walk-in patients attending a sexually transmitted disease clinic. Separate patient specimens were collected for culture on Martin-Lewis medium in JEMBEC plates and for DNA probe assay. Statistical analysis of the overall comparative results showed that the DNA probe assay had a sensitivity, specificity, and positive and negative predictive values of 93, 99, 97, and 99%, respectively, in a patient population with a gonococcal disease prevalence of 21%. The results of this comparative study showed that the prototype chemiluminescent DNA probe assay is a rapid and reliable noncultural alternative for the laboratory diagnosis of gonorrhea.
Poor performance of BACTEC NR 730 blood culture system in early detection of Neisseria meningitidis
Authors: Schnur E R , Azimi P H , Belchis D A
Published: January 2020

During an 8-month period at Children's Hospital, Oakland, Calif., a 9% rate for positive blood culture for children with Neisseria meningitidis meningitis was identified. The blood culture system used in each case was the BACTEC NR 730. This rate seemed significantly lower than previous rates (33 to 55%) (P.R. Dodge and M.N. Swartz, N. Engl. J. Med. 272:1003-1010, 1965; A.L. Hoyne and R.H. Brown, Ann. Intern. Med. 28:248-259, 1948; S. Levin and M.B. Painter, Ann. Intern. Med. 64:1049-1057, 1966). The low rate prompted our study. With 14 test strains, anaerobic and aerobic BACTEC bottles were evaluated for their ability to support and detect the growth of N. meningitidis. Sodium polyanetholesufonate (SPS) and inoculum size, two factors thought to affect the growth of N. meningitidis, were controlled for by use of bottles with and without SPS and by inoculum sizes simulating the magnitudes of bacteremia previously described for children infected with N. meningitidis (L.J. La Scolea, Jr., D. Dryja, T.D. Sullivan, L. Mosovich, N. Ellerstein, and E. Neter, J. Clin. Microbiol. 13:478-482, 1981). BACTEC failed to detect growth in aerobic bottles after 6 h of incubation, while 76 of 80 bottles (95%) showed growth when subcultured. At 24 h, BACTEC detected growth in only 29 of 80 bottles (36%); when subcultured, all 80 cultures grew confluently. At 48 h, BACTEC detected growth in the remaining 53 bottles. BACTEC failed to detect growth in anaerobic bottles at 6 h and at 1, 2, 4, and 5 days of incubation despite growth in subculture. Subcultures from bottles with tryptic soy broth with and without SPS showed growth in 63 to 76 bottles in 6 h and in all bottles after 24 h. The presence of SPS in BACTEC bottles had no effect on growth detection. On the basis of these studies and our clinical experience, we find the NR 730 system to be insensitive and unsuitable for detection of N.meningitidis in </= 24h. However, all test strains were detected after 48h.
Radiometric cytolysis inhibition assay, a new rapid test for neutralizing antibodies to intact and trypsin-cleaved poliovirus
Authors: Hovi T , Roivainen M
Published: January 2020

We have developed a new rapid test, the radiometric cytolysis inhibition assay (RACINA), for the determination of neutralizing poliovirus antibodies. HeLa cells prelabeled with 51Cr, [3H]leucine, or, preferentially, with [3H]uridine are used as sensitive quantitative indicators of residual infectious virus. Both suspensions and monolayer cultures of the indicator cells can be used. Neutralization of a fraction of a high-titer virus preparation can be scored after the first replication cycle at 8 to 10 h. By lowering the incubation temperature to 30 degrees C, the completion of the cytolysis due to the first replication cycle of poliovirus was delayed beyond 21 h. This makes it possible to use the RACINA, unlike the standard microneutralization assay, for measuring antibodies to trypsin-cleaved polioviruses. The RACINA was found to be as sensitive as and more reproducible than the standard microneutralization assay in the measurement of neutralizing poliovirus antibodies. The RACINA is a rapid and reliable test for neutralizing antibodies and in principle it may be applicable for quantitation of neutralizing antibodies to other cytolytic agents as well.
Assessment of the BACTEC NR660 blood culture system for the detection of bacteremia in young children
Authors: Anderson J D , Trombley C , Cimolai N
Published: January 2020

The sensitivity of the BACTEC NR660 blood culture system was assessed by using paired bottles of anaerobic (NR7A) and resin-containing aerobic (NR16A) media and conditions and organisms which simulated those found in pediatric practice. Corresponding media (7D and 16B) of the established BACTEC 460 radiometric method served as controls. The performances of the two systems were similar with 50 isolates of 10 aerobic organisms (aerobic medium) and with 21 isolates of 15 strict anaerobes (anaerobic medium).
Identification of gram-positive coccal and coccobacillary vancomycin-resistant bacteria
Authors: Facklam R , Hollis D , Collins M D
Published: January 2020

A total of 84 of 150 vancomycin-resistant (defined as no inhibition of bacterial growth around a 30-micrograms vancomycin disk placed on 5% sheep blood-Trypticase soy agar [BBL Microbiology Systems, Cockeysville, Md.]) bacteria were definitively identified by determining the phenotypic criteria. The identity of representatives was also confirmed by DNA-DNA hybridizations. The following strains were identified: 1 Enterococcus faecium, 18 Leuconostoc mesenteroides, 15 Leuconostoc citreum, 9 Leuconostoc pseudomesenteroides, 2 Leuconostoc lactis, 20 Pediococcus acidilactici, 5 Pediococcus pentosaceus, and 14 Lactobacillus confusus. The remaining vancomycin-resistant strains were identified as probable Leuconostoc (6 strains), probable Pediococcus (1 strain), and probable lactobacilli (28 strains). A total of 32 strains of gram-positive coccobacillary bacteria remained unidentified. Tests used for the phenotypic identification of strains to the genus level included a Gram stain of bacteria grown in thioglycolate broth, gas production in Lactobacillus Mann, Rogosa, and Sharpe broth, hydrolysis of pyrrolidonyl-beta-naphthylamide, bile-esculin reaction, demonstration of streptococcal group D antigen, and growth at 10 and 45 degrees C and in 6.5% NaCl broth. Strains were identified to the species level by hydrolysis of esculin, reactions in litmus milk, slime production on 5% sucrose agar, acidification of maltose, melibiose, and raffinose broths, deamination of arginine, and growth at 42 degrees C and in 6.5% NaCl broth.
Identification of Enterococcus species isolated from human infections by a conventional test scheme
Authors: Facklam R R , Collins M D
Published: January 2020

Streptococci (206 cultures) previously identified as enterococci were retrieved from storage and reidentified by using tests designed to identify species of the genus Enterococcus. Of these 188, 91% were correctly identified as Enterococcus species. Of the remaining strains, nine (4%) were unidentified and six (3%) and 3 (1.5%) were identified as Leuconostoc sp. and Lactococcus sp., respectively. Two new Enterococcus species were discovered: E. raffinosus and E. solitarius. DNA-DNA hybridizations were performed on selected strains to assure correct identification. Cultures representing 10 of the 12 Enterococcus species were among the 188 strains identified. An identification system based on the grouping of key reactions of 20 phenotypic characteristics of Enterococcus species is described.
Characterization of CDC group DF-3 by cellular fatty acid analysis
Authors: Wallace P L , Hollis D G , Weaver R E , Moss C W
Published: January 2020

Fourteen strains of Centers for Disease Control group DF-3 bacteria were examined for cellular fatty acid composition to evaluate their chemical relatedness to known bacterial species and groups. The fatty acids were liberated from whole cells by base hydrolysis, methylated, and analyzed by capillary gas-liquid chromatography. All group DF-3 strains possessed a distinct fatty acid profile which was characterized by large amounts (24%) of 12-methyltetradecanoate (a-C15:0), moderate amounts of saturated iso-branched-chain acids (i-C14:0 and i-C15:0), and small to moderate amounts of both branched- and straight-chain hydroxy acids (3-OH C15:0, i-3-OH C16:0, 3-OH C16:0, and i-3-OH C17:0). This fatty acid profile was unique as compared with the profiles of other bacteria we have previously tested but was most similar to the profiles of Capnocytophaga species.
Biotypes and O serogroups of Escherichia coli involved in intestinal infections of weaned rabbits: clues to diagnosis of pathogenic strains
Authors: Camguilhem R , Milon A
Published: January 2020

A total of 575 Escherichia coli strains isolated from weaned rabbits experiencing diarrhea in 119 French commercial farms were tested for O serogroups. The results showed a strong predominance of serogroup O103 strains. A sample of 126 strains were further checked for simplified biotypes by using five carbohydrate fermentation reactions. Of 72 O103 strains, 70 were shown to belong to biotypes characterized by a rhamnose-negative reaction. Four of nine serogroup O68 strains also showed this type of reaction. Thirty-nine strains, representative of the serotypes and biotypes found, were further tested for experimental pathogenicity in weaned rabbits and for antibiotic susceptibility. All the rhamnose-negative strains produced life-threatening watery or hemorrhagic diarrhea, whereas rhamnose-positive strains induced only mild diarrheic syndrome without any mortality or no clinical signs at all. Rhamnose-negative, highly pathogenic strains did not belong to related antibiotypes. We think that O serogrouping together with biotyping, or even rhamnose fermentation testing, may be an important clue in the diagnosis of enteropathogenic strains from rabbits in France, permitting rapid identification of highly pathogenic strains and leading to improved prognosis and treatment.
Evaluation of enzyme immunoassay for the detection of pathogenic Yersinia enterocolitica and Yersinia pseudotuberculosis strains
Authors: Kaneko S , Maruyama T
Published: January 2020

To determine the virulence plasmid-harboring strains of Yersinia enterocolitica, we prepared antiserum against plasmid-encoded proteins of Y. enterocolitica serotype O3 and carried out an enzyme immunoassay (EIA) against temperature-inducible released proteins. This serum reacted with proteins released from not only a Y. enterocolitica serotype O3 strain but also Y. enterocolitica serotype O5:27, O8, and O9 and Y. pseudotuberculosis serotype 1b, 2a, 2b, 2c, 3, 4a, 4b, 5a, 5b, 6, 7, and 8 strains, which all harbored plasmids. Plasmid-cured Y. enterocolitica and Y. pseudotuberculosis strains did not react in the EIA, nor did nonpathogenic Y. enterocolitica strains or Y. frederiksenii, Y. intermedia, and Y. kristensenii strains. These observations demonstrated that this EIA was useful for determining whether the isolated Yersinia strains were pathogenic or not.
Evaluation of culture, immunofluorescence, and serology for the diagnosis of pertussis
Authors: Halperin S A , Bortolussi R , Wort A J
Published: January 2020

Nasopharyngeal culture, direct immunofluorescence, and serology of acute-phase and paired serum specimens were compared for the laboratory diagnosis of infections due to Bordetella pertussis in a community-based pediatric population with both high vaccine usage and high pertussis incidence. In 77 (37%) of 210 patients evaluated, one or more tests were positive for pertussis. A clinical illness compatible with pertussis was present in 52 (71%) of 73 pertussis test-positive and 42 (35%) of 119 test-negative patients (P less than 0.001). Nasopharyngeal culture was of low sensitivity (20 [26%] of 77 positive tests) but was most commonly confirmed by another positive pertussis test (85%). Direct immunofluorescence was both insensitive and nonspecific; only 6 (30%) of 20 cases positive by culture were positive by immunofluorescence, and only 4 (33%) of 12 of the culture-negative, immunofluorescence-positive cases could be confirmed by another positive pertussis test. Although serology by enzyme immunoassay proved to be the most sensitive of the laboratory tests (87%), this sensitivity could be achieved only by assaying both acute-phase and paired serum specimens and measuring immunoglobulin G (IgG), IgA, and IgM antibodies to two pertussis antigens (pertussis toxin and filamentous hemagglutinin). Loss of sensitivity occurred with any reduction in the number of these serologic assays performed. Optimal laboratory diagnosis of endemic pertussis in a pediatric population requires both nasopharyngeal culture and serology by enzyme immunoassay.
Comparison of lipopolysaccharides from Brazilian purpuric fever isolates and conjunctivitis isolates of Haemophilus influenzae biogroup aegyptius. Brazilian Purpuric Fever Study Group
Authors: Erwin A L , Munford R S
Published: January 2020

Haemophilus influenzae biogroup aegyptius (H. aegyptius) has been identified as the etiologic agent of the recently described disease Brazilian purpuric fever (BPF). Although there is heterogeneity among the strains associated with conjunctivitis, isolates from patients with BPF appear to be derived from a single clone. The clinical presentation of BPF suggests that bacterial lipopolysaccharides (LPS) are involved in its pathogenesis. We prepared LPS from H. influenzae biogroup aegyptius and found them to be similar to H. influenzae type b LPS in apparent size (by sodium dodecyl sulfate-polyacrylamide gel electrophoresis), biological activities, and fatty acid composition. We compared LPS from BPF clone isolates with LPS from non-BPF clone isolates in tests of Limulus lysate activation, spleen cell mitogenesis, promotion of neutrophil adherence to LPS-treated endothelial cells, and the dermal Shwartzman reaction. In none of these activities were LPS from the BPF clone isolates more potent. Because LPS shed from growing bacteria may be involved in the pathogenesis of purpura, we also measured the rate at which LPS were released into culture medium during bacterial growth and found no significant difference between BPF clone and non-BPF clone isolates.
Increased detection of herpes simplex virus in MRC-5 cells treated with dimethyl sulfoxide and dexamethasone
Authors: West P G , Aldrich B , Hartwig R , Haller G J
Published: January 2020

Treatment of MRC-5 cells with dexamethasone, dimethyl sulfoxide, or a combination of the two enhanced the detection of herpes simplex virus by three- to fourfold in these cells. Fluorescent plaques were noticeably larger on cover slips treated with the enhancing agents. Low-positive clinical specimens were stored and tested in parallel in treated and untreated shell vials, and 6 to 40% of these stained positive in the treated cultures but not in the untreated controls. In standard tube cultures, cytopathic effect began earlier and was more extensive in treated tubes.
Efficacy of the urinary bladder for isolation of Borrelia burgdorferi from naturally infected, wild Peromyscus leucopus
Authors: Callister S M , Agger W A , Schell R F , Brand K M
Published: January 2020

The efficacy of culturing urinary bladder tissue for Borrelia burgdorferi from naturally infected, wild Peromyscus leucopus mice was determined. The urinary bladder cultures were as efficient as spleen, kidney, and blood tissue cultures. The rapid B. burgdorferi isolation (mean, 6 days) from mouse urinary bladders should aid in defining new Lyme disease foci.
Rapid detection of Escherichia coli in urine samples by a new chromogenic beta-glucuronidase assay
Authors: Delisle G J , Ley A
Published: January 2020

A new compound, indoxyl-beta-D-glucuronide, was assessed as a substrate for the rapid detection of Escherichia coli in urine. Incorporation of this compound into MacConkey agar allowed the direct differentiation of E. coli as deep blue colonies distinct from lactose and nonlactose fermenters. The sensitivity was 88 to 90%, and the specificity was 100%.
Passive protection against rotavirus-induced diarrhea by monoclonal antibodies to the heterotypic neutralization domain of VP7 and the VP8 fragment of VP4
Authors: Matsui S M , Offit P A , Vo P T , Mackow E R , Benfield D A , Shaw R D , Padilla-Noriega L , Greenberg H B
Published: January 2020

A murine model was used to determine whether neutralizing monoclonal antibodies (MAbs) with heterotypic specificity directed to VP7 (MAb 57-8) or to the VP8 fragment of VP4 (MAb M14) passively protect mice against challenge with various strains of rotavirus. (The gene 4 product, an outer capsid protein, has traditionally been called VP3. It has been proposed, however, that the rotavirus gene 4 product be named VP4. The gene 3 product, a core protein, has been identified recently and named VP3 [M. Liu, P. A. Offit, and M. K. Estes, Virology 163:28-32, 1988]). Suckling mice orally inoculated with MAb 57-8 did not develop diarrhea when challenged with virulent serotype 3, 4, or 6 rotaviruses, while those inoculated with MAb M14 were passively protected from challenge with serotype 3 or 6 rotaviruses, as predicted by in vitro neutralization tests. These MAbs, however, did not protect mice from infection when the mice were challenged with rotaviruses of other serotypes. We conclude that specific neutralization epitopes on each surface protein are capable of mediating protection against one or several rotavirus serotypes.
Special features of distribution of Yersinia pseudotuberculosis in Japan
Authors: Tsubokura M , Otsuki K , Sato K , Tanaka M , Hongo T , Fukushima H , Maruyama T , Inoue M
Published: January 2020

The incidence of Yersinia pseudotuberculosis isolation from humans, animals, water, and pork in Japan is presented on the basis of a review of the literature and serotyping of 633 strains referred to our laboratory. Most of the strains belonged to serogroup 4b, followed in frequency of isolation by serogroups 3, 1b, 5b, 5a, and the others, whereas serogroup 1a has not been detected. Although strains were isolated from 11 species of animals, none were isolated from birds.