bluuebunny/crossref_metadata_2025 huggingface dataset:
Chemical and cultural characterization of CDC group WO-1, a weakly oxidative gram-negative group of organisms isolated from clinical sources
Authors: Hollis D G , Weaver R E , Moss C W , Daneshvar M I , Wallace P L
Published: January 2020

Ninety-six strains of weakly oxidative gram-negative rods isolated primarily from clinical specimens form a distinct group that has been designated Centers for Disease Control (CDC) group WO-1 (WO stands for weak oxidizer). The phenotypic characteristics of CDC group WO-1 were most similar to those of Comamonas acidovorans, Pseudomonas mallei, and CDC pink coccoid group III. The WO-1 group can be differentiated from C. acidovorans by the oxidation of glucose (often weak and sometimes delayed), motility by means of one or two polar flagella, and, when positive, the complete reduction of nitrate and nitrite. Motility and usually the failure to produce arginine dihydrolase distinguish this group from P. mallei. The WO-1 strains differ from the pink coccoid group III by the absence of pink growth pigment, the lack of predominantly coccoid cellular morphology, and usually the inability to produce acid from xylose. The cellular fatty acid compositions of 29 group WO-1 strains were characterized by large amounts of C16:0 and C16:1w7c; smaller amounts of C18:1w7c, C14:0, C12:0, and 3-OH-C10:0; and trace to small amounts of C15:1w6 and C17:0 acids. The fatty acid profile of WO-1, compared with the profiles of other bacteria we have tested previously, was most similar to the profiles of two phenotypically different organisms, Comamonas terrigena (a nonoxidative, multipolar gram-negative rod) and Chromobacterium violaceum (a fermentative gram-negative rod). Ubiquinone-8 was the major quinone in the five WO-1 strains examined. Eighty-five percent of the WO-1 strains were isolated from human specimens. Thirty-three percent were from blood, and 10% were from cerebrospinal fluid.
Microbiologic assessment of tissue biopsy samples from ileal pouch patients
Authors: Onderdonk A B , Dvorak A M , Cisneros R L , McLeod R S , Antionoli D , Silen W , Blair J E , Monahan-Earley R A , Cullen J , Cohen Z
Published: January 2020

Tissue biopsy samples from patients with and without ileal pouches were examined by electron microscopic and microbiologic culture techniques to determine the numbers and types of microorganisms closely associated with or within the tissue biopsy samples. The disease status of each patient was determined by endoscopic and histopathologic methods. Of the 78 biopsy samples included in this study, 64 (82%) yielded obligately anaerobic and/or facultative bacteria when they were cultured. Fourteen of the 78 samples (17.9%) were negative by culture. Of the positive samples, 54 contained facultatively anaerobic bacterial species and 50 yielded obligately anaerobic species. The total counts for facultatively anaerobic bacteria for samples from patients with pouchitis were significantly greater than for samples from patients in control groups. In addition, the number of samples from patients with normal pouches that did not contain obligate anaerobes was significantly less than that from patients with pouchitis; 4 of 23 and 6 of 12 samples, respectively (P less than 0.043). For samples in which organisms were detected, there was agreement with electron microscopic detection of bacteria in 23 of 27 samples, for an overall sensitivity of electron microscopy compared with that of culture of 85%. The qualitative studies resulted in the characterization of 273 isolates comprising 77 different phenotypes. The specificity of these findings in patients with ileal pouchitis is discussed.
Controlled comparison of the BacT/Alert and BACTEC 660/730 nonradiometric blood culture systems
Authors: Wilson M L , Weinstein M P , Reimer L G , Mirrett S , Reller L B
Published: January 2020

In a collaborative study at three university hospitals, the recovery of microorganisms and the speed of detection of microbial growth by the BacT/Alert (Organon Teknika Corporation, Durham, N.C.) and BACTEC 660/730 (Becton-Dickinson Diagnostic Instrument Systems, Sparks, Md.) nonradiometric blood culture systems were compared. A total of 5,918 comparisons were made between BacT/Alert aerobic and BACTEC NR 6A bottles and 5,992 comparisons were made between BacT/Alert anaerobic and BACTEC NR 7A bottles. Each bottle was inoculated with 5 ml of blood. The overall recoveries of microorganisms from the two aerobic bottles were comparable; members of the family Enterobacteriaceae were recovered more often from BacT/Alert aerobic bottles alone (P less than 0.001). The overall recoveries of microorganisms from the anaerobic bottles were not significantly different. Growth of Staphylococcus aureus (P less than 0.001), coagulase-negative staphylococci (P less than 0.01), streptococci (P less than 0.001), Escherichia coli (P less than 0.01), other members of the family Enterobacteriaceae (P less than 0.02), and Pseudomonas aeruginosa (P less than 0.05) was detected earlier in BacT/Alert aerobic bottles. Growth of S. aureus (P less than 0.001), coagulase-negative staphylococci (P less than 0.05), enterococci (P less than 0.01), Streptococcus pneumoniae (P less than 0.02), viridans group streptococci (P less than 0.05), E. coli (P less than 0.001), Klebsiella pneumoniae (P less than 0.01), and other members of the family Enterobacteriaceae (P less than 0.001) was detected earlier in BacT/Alert anaerobic bottles. In a system-versus-system comparison, more gram-positive cocci were recovered from the BACTEC system alone (P < 0.05), and more members or the family Enterobacteriaceae were recovered from the BacT/Alert system alone (P < 0.001). As a system, the BacT/Alert system detected growth of S. aureus (P < 0.001), coagulase-negative staphylococci (P < 0.01), streptococci (P < 0.001), E. coli (P < 0.001), other members of the familyEnterobacteriaceae (P < 0.001), and P. aeruginosa (P < 0.05) earlier than the BACTEC system did.Significantly fewer (40 versus 1,183) false-positive results occurred with the BacT/Alert system. We conclude that the BacT/Alert and BACTEC 660/730 nonradiometric systems are comparable for recovering clinically significant microorganisms form adult patients with bacteremia or fungemia, but that the BacT/Alert system detects microbial growth earlier than the BACTEC system does, with significantly fewer false-positive results.
Recombinant Leishmania Hsp90 and Hsp70 are recognized by sera from visceral leishmaniasis patients but not Chagas' disease patients
Authors: de Andrade C R , Kirchhoff L V , Donelson J E , Otsu K
Published: January 2020

Approximately 70% of the cDNA clones identified by immunoscreening Leishmania donovani expression libraries with serum from a patient with visceral leishmaniasis (kala-azar) were found to encode the highly conserved Hsp90 and Hsp70 members of the heat shock protein family. Recombinant fusion proteins containing the C-terminal portions of L. donovani Hsp90 and Hsp70 were used as target antigens in enzyme-linked immunosorbent assays of various sera. Sera from four patients with visceral leishmaniasis recognized recombinant Leishmania Hsp90 and Hsp70, while sera from seven patients with Chagas' disease did not, despite the fact that Trypanosoma cruzi Hsp90 and Hsp70 share more than 80% amino acid identity with their counterparts in Leishmania spp. Thus, Leishmania Hsp90 and Hsp70 elicit strong humoral responses and are potential candidates for specific serodiagnostic assays capable of distinguishing between L. donovani and T. cruzi infections.
Characterization of pseudorabies virus antibody responses in young swine after infection and vaccination by using an immunoglobulin M antibody capture enzyme-linked immunosorbent assay
Authors: McCaw M B , Molitor T W , Joo H S
Published: January 2020

An immunoglobulin M (IgM) antibody capture enzyme-linked immunosorbent assay (MACELISA) was developed for the detection of pseudorabies virus (PRV)-specific IgM antibody in swine sera because false-positive reactions frequently occurred when sera from older swine were tested with an indirect IgM enzyme-linked immunosorbent assay. Monoclonal mouse anti-swine IgM was used as the capturing antibody, and rabbit anti-PRV hyperimmune gamma globulin was used as the indicating antibody. Sera from non-PRV-infected, experimentally infected, vaccinated and challenged, passively immune and challenged, and naturally infected swine were evaluated. The PRV MACELISA had a specificity of 95% and was as sensitive and reproducible as previously reported in direct assays. An antibody response was still detectable with the MACELISA 21 days after inoculation. The PRV MACELISA did not detect a consistent antibody response in sera from swine vaccinated with either killed-PRV or modified live-virus vaccines but did detect an antibody response in sera from passively immune pigs after challenge with virulent PRV. These results indicated that the PRV MACELISA may be useful for the rapid serodiagnosis of recent PRV infection in swine.
Cultivation of Borrelia burgdorferi from erythema migrans lesions and perilesional skin
Authors: Berger B W , Johnson R C , Kodner C , Coleman L
Published: January 2020

Skin biopsy specimens from the peripheral aspect of erythema migrans lesions (site 1) and from clinically normal perilesional areas (site 2) were compared as sources of Borrelia burgdorferi. This spirochete was isolated from the skin of 18 of 21 (86%) patients with untreated early Lyme disease at one or both biopsy sites. Site 1 specimens were superior to site 2 specimens for the isolation of B. burgdorferi. Site 1 specimens from 18 (86%) patients were culture positive, and site 2 specimens from 12 (57%) patients were culture positive. For patients whose site 2 specimens were culture positive, site 1 specimens were also found to be culture positive. B. burgdorferi was isolated from two patients with atypical lesions and from two patients with erythema migrans lesions that were less than 5 cm in diameter. This study demonstrates that the cultivation of B. burgdorferi from skin biopsy specimens from cutaneous lesions thought to be erythema migrans can be an efficacious procedure for confirming the diagnosis of Lyme disease and that the spirochete is present in clinically normal appearing perilesional skin.
Epidemiologic typing of Staphylococcus aureus by DNA restriction fragment length polymorphisms of rRNA genes: elucidation of the clonal nature of a group of bacteriophage-nontypeable, ciprofloxacin-resistant, methicillin-susceptible S. aureus isolates
Authors: Blumberg H M , Rimland D , Kiehlbauch J A , Terry P M , Wachsmuth I K
Published: January 2020

Analysis of DNA restriction fragment length polymorphisms of rRNA genes (ribotyping) was employed to assist in the epidemiologic investigation of the emergence and spread of ciprofloxacin-resistant Staphylococcus aureus at the Atlanta VA Medical Center because many isolates of interest were nontypeable by phages and harbored few plasmids useful as strain markers. Chromosomal DNAs of selected S. aureus isolates were digested initially with 20 different restriction enzymes. EcoRI appeared to give the best discrimination of hybridization banding patterns (ribotypes) and was used with all study isolates. Overall, 15 different ribotypes were seen among the 50 S. aureus isolates studied (7 ribotypes among 13 methicillin-susceptible S. aureus [MSSA] isolates and 9 ribotypes among 37 methicillin-resistant S. aureus [MRSA] isolates). Seven of eight ciprofloxacin-resistant MSSA (CR-MSSA) patient isolates had identical antibiograms, were nontypeable by phages, and had a single 22-MDa plasmid. Six of these seven CR-MSSA isolates had an identical ribotype pattern. Ribotyping distinguished this CR-MSSA strain or clone from MRSA and other MSSA isolates, including nontypeable isolates that contained a 22-MDa plasmid. Five ciprofloxacin-susceptible MSSA isolates studied had five ribotypes; one pattern was identical to the CR-MSSA clone. Twenty-three CR-MRSA isolates recovered from the Atlanta VA Medical Center had four different ribotypes. Ribotyping proved to be a useful molecular epidemiologic tool in the study of S. aureus because it differentiated isolates which were indistinguishable by more traditional methods. In addition, this technique demonstrated that at our institution, ciprofloxacin resistance emerged in multiple strains of MRSA, as opposed to primarily a single strain or clone of MSSA.
Clinical and laboratory analyses of cytospin-prepared Gram stains for recovery and diagnosis of bacteria from sterile body fluids
Authors: Chapin-Robertson K , Dahlberg S E , Edberg S C
Published: January 2020

The smear of a clinical specimen provides essential laboratory information that is used to make therapeutic decisions. For this study, smears were made by centrifugation in a Beckman Microfuge 11 (Beckman Instruments, Palo Alto, Calif.) and in parallel by using a Cytospin 2 apparatus (Shandon Inc., Pittsburgh, Pa.). Of 350 consecutive body fluid specimens examined, 50 (14.0%) grew bacteria. Both methods were culture and smear positive for 24 (6.9%) specimens; 18 (5.1%) specimens were cytocentrifuge smear positive, culture positive, and high-speed centrifugation (HSC) negative; 3 (0.8%) were culture negative and positive by both smear methods; and 1 (0.2%) was HSC smear positive, culture positive, and cytocentrifuge negative. Seven (2.0%) specimens were culture positive and negative by both smear methods. Clinically, cytocentrifuge preparations showed greater sensitivity for culture-positive specimens and a closer correlation with the CFU per milliliter than HSC did, resulting in a greater ability to treat patients with specific therapies. In addition, analysts needed to examine only a 6-mm-diameter area on the slide, cells and microbes were somewhat larger and more regular in appearance, and smears stained more uniformly. Because of the increased clinical and laboratory utility of the cytocentrifuge, its use is recommended in clinical microbiology laboratories for all sterile body fluid specimens.
Clinical illnesses associated with isolation of dysgonic fermenter 3 from stool samples
Authors: Blum R N , Berry C D , Phillips M G , Hamilos D L , Koneman E W
Published: January 2020

The clinical significance of the fastidious organism DF-3 isolated from stool cultures is unclear. We sought to improve our understanding of this organism and to further define its association with human disease. Stool cultures for DF-3 were obtained from three sources: an ongoing study of enteric pathogens in patients infected with the human immunodeficiency virus, a screening procedure in which all stool samples submitted for Clostridium difficile toxin assay were cultured for DF-3, and stool samples submitted specifically for DF-3 culture. Retrospective clinical data were obtained from chart reviews of patients with positive cultures. Antimicrobial susceptibility testing and cell wall fatty acid analysis were performed for each DF-3 isolated. Eight isolates of DF-3 were obtained over a period of 8 months. All patients either had severe underlying disease or were immunocompromised, including three patients coinfected with human immunodeficiency virus and two patients with inflammatory bowel disease. The spectrum of clinical disease ranged from chronic diarrhea with a well-defined response to therapy for DF-3 to an asymptomatic carrier state. Cell wall fatty acid analysis of these isolates demonstrated a consistent pattern with a large peak of 12-methyltetradecanoate. DF-3, a fastidious gram-negative coccobacillus, can be recovered from stool cultures of immunocompromised patients by using selective media. The presence of 12-methyltetradecanoate in cell wall fatty acid analysis assists in identification. The increased use of a selective medium-(cefoperazone-vancomycin-amphotericin B) in the evaluation of diarrhea in immunocompromised hosts, including persons with inflammatory bowel disease, may better define the association of DF-3 with human gastrointestinal disease.
Clinical and epidemiological importance of typing of Mycobacterium avium complex isolates
Authors: Tsang A Y , Denner J C , Brennan P J , McClatchy J K
Published: January 2020

The results of the application of a range of typing procedures to the identification and classification of 6,264 cultures of nontuberculous mycobacteria from human sources and the environment are reported. Seroagglutination, an enzyme-linked immunosorbent assay applied to whole bacteria or the glycolipid typing antigens and based on serovar-specific polyclonal or monoclonal antibodies, thin-layer chromatography of these antigens, and gas chromatography of their specific sugar determinants were used to arrive at identifications. As a result of this comprehensive approach, 4,452 (71%) of all cultures and 88% of those of samples from patients with AIDS proved to be typeable. The rank order of frequency of occurrence of individual organisms within the entire group of isolates was Mycobacterium avium complex serovar 4 greater than serovar 8 greater than serovar 1 greater than serovar 9 greater than serovar 6 greater than serovar 14 greater than serovar 2 greater than M. fortuitum greater than M. kansasii greater than M. xenopi greater than an apparent mixture of serovar 4 and M. xenopi greater than a mixture of serovar 4 and serovar 8. These results were similar but not identical to the pattern observed for isolates obtained from patients with AIDS; the order was M. avium complex serovar 4 greater than serovar 8 greater than serovar 1 greater than a mixture of serovar 4 and M. xenopi, a mixture of serovar 4 and serovar 8 greater than serovar 9 greater than serovar 2 greater than serovar 6. Serotyping was also used to demonstrate the possible clinical significance of nontuberculous mycobacteria recovered from different body sites. Other information on the distribution of M. avium serovars in patients from different geographical environments is provided.
Liquid medium for rapid presumptive identification of group B streptococci
Authors: Teixeira L A , Figueiredo A M , Benchetrit L C
Published: January 2020

A suitable test was developed for distinguishing group B streptococci from other beta-hemolytic streptococci during growth in liquid medium. One hundred and sixty of 161 human group B strains tested yielded positive reactions within a 5 h incubation. The dye medium tested is a reliable substitute for more expensive serological procedures.
Minimum number of pneumococci required for capsular antigen to be detectable by latex agglutination
Authors: Holloway Y , Boersma W G , Kuttschrütter H , Snijder J A
Published: January 2020

Forty-eight strains of Streptococcus pneumoniae were tested in vitro to determine the minimum number required for pneumococcal capsular antigen to be detectable by latex agglutination. It was found that 10(6) to 10(7) microorganisms per ml were needed and that antigen remained detectable even when viable pneumococci could no longer be demonstrated.
Comparison of four techniques for detection of antibodies to cytomegalovirus
Authors: Kraat Y J , Hendrix R M , Landini M P , Bruggeman C A
Published: January 2020

Four serological methods were compared and evaluated for use in detecting cytomegalovirus antibody in blood and organ donors. Western blotting (immunoblotting), latex agglutination, enzyme-linked immunosorbent assay, and a recent available microparticle enzyme immunosorbent assay were used. The microparticle enzyme immunoassay appears to compare favorably with each of the other three assays tested for screening blood and organ donors for a previous cytomegalovirus infection.
Evaluation of an enzyme-linked immunoassay employing a covalently bound capture antibody for direct detection of herpes simplex virus
Authors: Needham C A , Hurlbert P
Published: January 2020

The FDL enzyme-linked immunosorbent assay (ELISA; Fairleigh Dickinson Laboratories, Inc., Abilene, Tex.) for the detection of herpes simplex virus (HSV) makes use of a covalently attached antibody. This assay was compared with viral isolation and with the Ortho HSV Antigen Detection ELISA (Ortho Diagnostic Systems, Inc., Raritan, N.J.). One hundred forty-eight specimens were tested. The FDL ELISA identified 66 of 104 specimens from which HSV was isolated, yielding a sensitivity of 63% and a specificity of 95%. These results compared favorably with those obtained by using the Ortho ELISA. The total test time was shorter and the washing step was simpler than that with the Ortho assay, making the FDL assay an attractive alternative to similar methodologies.
Rapid method for epidemiological evaluation of gram-positive cocci by field inversion gel electrophoresis
Authors: Goering R V , Winters M A
Published: January 2020

We report a rapid method for the isolation of intact chromosomal DNA from gram-positive cocci that is suitable for in situ restriction endonuclease digestion in agarose blocks. When combined with a rapid field inversion gel electrophoresis protocol, this approach allows the preparation and electrophoretic analysis of chromosomal restriction fragments produced by rare-cutting enzymes in a total time period of 2 days from start to finish. The utility of the method is demonstrated in the epidemiological evaluation of Staphylococcus epidermidis clusters from two hospitals as well as of additional representative staphylococci and enterococci.
Differentiation between active and inactive human brucellosis by measuring antiprotein humoral immune responses
Authors: Goldbaum F A , Rubbi C P , Wallach J C , Miguel S E , Baldi P C , Fossati C A
Published: January 2020

A preparation of Brucella abortus cytoplasmic proteins was depleted of lipopolysaccharide (LPS) by immunoadsorption with a monoclonal antibody (MAb), BC68, specific for the O antigen of B. abortus smooth LPS. Two enzyme-linked immunosorbent assay (ELISA) systems were developed and used in this study. The first system includes an LPS-free cytoplasmic protein preparation; the second one was based on antigen capture on MAb BC68. By using these systems, we have demonstrated that 94% (33 of 35) of the brucellosis patients studied showed immunoglobulin G antiprotein response and also that all of the patients showed a strong anti-LPS reactivity. Thirty-six serologically positive individuals with no active infection at the time of examination (SPI) were also included. No immunoglobulin G antibodies against proteins were detected in 34 of them (92%), whereas 31 SPI (86%) showed various degrees of anti-LPS reactivity. The use of the LPS-free protein extract in ELISAs made it possible to establish differential reactivity patterns between active and inactive brucellosis.
Use of single-gene reassortant viruses to study the role of avian influenza A virus genes in attenuation of wild-type human influenza A virus for squirrel monkeys and adult human volunteers
Authors: Clements M L , Subbarao E K , Fries L F , Karron R A , London W T , Murphy B R
Published: January 2020

The transfer of six internal RNA segments from the avian influenza A/Mallard/New York/6750/78 (H2N2) virus reproducibly attenuates human influenza A viruses for squirrel monkeys and adult humans. To identify the avian influenza A virus genes that specify the attenuation and host range restriction of avian-human (ah) influenza A reassortant viruses (referred to as ah reassortants), we isolated six single-gene reassortant viruses (SGRs), each having a single internal RNA segment of the influenza A/Mallard/New York/6750/78 virus and seven RNA segments from the human influenza A/Los Angeles/2/87 (H3N2) wild-type virus. To assess the level of attenuation, we compared each SGR with the A/Los Angeles/2/87 wild-type virus and a 6-2 gene ah reassortant (having six internal RNA segments from the avian influenza A virus parent and two genes encoding the hemagglutinin and neuraminidase glycoproteins from the wild-type human influenza A virus) for the ability to replicate in seronegative squirrel monkeys and adult human volunteers. In monkeys and humans, replication of the 6-2 gene ah reassortant was highly restricted. In humans, the NS, M, PB2, and PB1 SGRs each replicated significantly less efficiently (P less than 0.05) than the wild-type human influenza A virus parent, suggesting that each of these genes contributes to the attenuation phenotype. In monkeys, only the NP, PB2, and possibly the M genes contributed to the attenuation phenotype. These discordant observations, particularly with regard to the NP SGR, indicate that not all genetic determinants of attenuation of influenza A viruses for humans can be identified during studies of SGRs conducted with monkeys. The PB2 and M SGRs that were attenuated in humans each exhibited a new phenotype that was not observed for either parental virus. Thus, it was not possible to determine whether avian influenza virus PB2 or M gene itself or a specific constellation of avian and human influenza A virus specified restriction of virus replication in humans.
Sialidases (neuraminidases) in bacterial vaginosis and bacterial vaginosis-associated microflora
Authors: Briselden A M , Moncla B J , Stevens C E , Hillier S L
Published: January 2020

Bacterial vaginosis, Prevotella species, and Bacteroides species have been associated with prematurity and upper genital tract infection. Prevotella (Bacteroides) species and Bacteroides fragilis have also been associated with preterm birth. However, the mechanism by which lower genital tract infection causes upper genital tract disease remains poorly understood. Sialidases (neuraminidases) are enzymes which enhance the ability of microorganisms to invade and destroy tissue. Elevated levels of sialidase activity were detected in 42 (84%) of 50 vaginal fluid specimens from women with bacterial vaginosis and none of 19 vaginal fluids from women without bacterial vaginosis (P less than 0.001). Vaginal fluid from women with bacterial vaginosis had a median specific activity of 9.8 U compared to 2.5 U of sialidase in women without bacterial vaginosis (P less than 0.001). In order to determine the probable source of sialidases in vaginal fluid, the microorganisms recovered from women with bacterial vaginosis before and after treatment were assayed. Of 28 specimens from women with bacterial vaginosis, 27 (96%) yielded sialidase-positive bacteria, at a median concentration of 10(6.5) CFU/ml of vaginal fluid. Prevotella and Bacteroides species accounted for the sialidase activity in 26 of the vaginal fluids, and Gardnerella vaginalis accounted for the sialidase activity in the remaining fluid. After treatment, sialidase was detected in the vaginal fluid of 1 (5%) of 22 women who responded to therapy and in all of 6 women for whom therapy failed. These data suggest that vaginal fluid sialidase is highly correlated with bacterial vaginosis and that the probable sources for this enzyme activity are the Bacteroides and Prevotella species present in the vagina.
Detection of cytomegalovirus in urine samples by an enzyme-linked immunosorbent assay using a monoclonal antibody against the viral 150-kilodalton protein
Authors: Yamanaka T , Kiyotani K , Sakaguchi T , Fukuda Y , Dohi K , Yamada M , Yoshida M , Nii S , Yoshida T
Published: January 2020

McKeating et al. (J.A. McKeating, P.D. Griffiths, and J.E. Grundy, J. Gen. Virol. 68:785-792, 1987; J. A. McKeating, J. E. Grundy, Z. Varghese, and P. D. Griffiths, J. Med. Virol. 18:341-348, 1986; J. A. McKeating, S. Stagno, P. R. Stirk, and P. D. Griffiths, J. Med. Virol. 16:367-373, 1985) reported previously that beta 2 microglobulin inhibits the detection of human cytomegalovirus (CMV) in urine specimens by an enzyme-linked immunosorbent assay (ELISA) with a monoclonal antibody against the glycoprotein of CMV. They postulated that beta 2 microglobulin binds to the viral glycoproteins and masks the antigenic determinants. We developed here an ELISA method for the detection of CMV in urine by using a monoclonal antibody against the viral 150-kDa protein to capture the viral antigen. This assay detected CMV both in culture medium and in urine specifically at concentrations higher than 10(3) PFU/ml and quantitatively at concentrations higher than 10(4) PFU/ml. The sensitivity of the ELISA increased about 10-fold when peroxidase-labeled F(ab')2 from goat anti-human immunoglobulin G was used as a secondary detecting antibody in combination with concentration of the virus in urine samples by ultracentrifugation. The inhibition of ELISA by beta 2 microglobulin was not observed in this ELISA system. When 56 urine specimens from renal transplant recipients were examined for CMV antigens, the ELISA system had a sensitivity of 78% and a specificity of 97%. The positive and negative predictive values of the assay were 95 and 86%, respectively. Furthermore, CMV antigens in urine were quantitated by the assay during the course of typical CMV disease of renal transplant recipient. These results suggest strongly that the measurement of CMV antigens in urine by our rapid and quantitative ELISA system provides very useful data for the monitoring of CMV infections in renal transplant recipients and making decisions about therapy.
Comparisons of rotavirus VP7-typing monoclonal antibodies by competition binding assay
Authors: Raj P , Matson D O , Coulson B S , Bishop R F , Taniguchi K , Urasawa S , Greenberg H B , Estes M K
Published: January 2020

Three sets of neutralizing monoclonal antibodies (MAbs) used to type the outer capsid protein VP7 of four group A rotavirus serotypes (1 through 4) were compared in competition immunoassays. Reciprocal competition was observed for each of the VP7 type 2-, 3-, and 4-specific MAbs. The VP7 type 1 MAbs exhibited variable competition patterns with other VP7 type 1 MAbs. MAb RV4:3, which has been used to recognize antigenic variants within VP7 type 1 strains, showed reciprocal competition with the four VP7 type 3 MAbs (RV3:1, YO-1E2, 4F8, and 159) using a VP7 type 3 virus (SA11) as antigen. MAb 2C9, also prepared against VP7 type 1, reacted with VP7 type 3 strains and competed with a VP7 type 3 MAb, 159, using RRV as antigen. Use of the different sets of VP7 type-specific MAbs in the enzyme-linked immunosorbent assay permitted the recognition of six antigenic variants within VP7 types 1, 2, and 3 among specimens whose VP7 type could not be determined previously with only one set of typing MAbs. These results demonstrate differences of typing ability among these VP7-specific MAbs and emphasize the need to improve the sensitivity of typing systems by incorporating panels of MAbs reacting with several neutralizing epitopes.
Epidemiology of Yersinia pseudotuberculosis and Y. enterocolitica infections in sheep in Australia
Authors: Slee K J , Skilbeck N W
Published: January 2020

Infections with Yersinia pseudotuberculosis serotype III and Y. enterocolitica serotype O2,3 were found to be common in Australian sheep flocks. Transmission of Y. pseudotuberculosis occurred in late winter and early spring, while Y. enterocolitica transmission occurred from midwinter to early summer. Excretion of Y. pseudotuberculosis was limited to the winter and spring period and was particularly common in 1- and 2-year-old sheep. Infection persisted for up to 14 weeks. Y. pseudotuberculosis infection did not confer immunity to natural infection with Y. enterocolitica. Y. enterocolitica excretion occurred year-round, with the greatest prevalence being in summer and autumn. Infection persisted for up to 29 weeks. Sheep less than 1 year old were most commonly infected with Y. enterocolitica. Infection with either Y. pseudotuberculosis or Y. enterocolitica was rare in aged sheep. Restriction endonuclease analysis of Y. pseudotuberculosis serotype III from sheep, cattle, deer, and pigs showed that the bacterial isolates were genetically indistinguishable. Similarly, Y. enterocolitica isolates from sheep were indistinguishable from those isolated from goats and cattle.
Evaluation of a microtiter plate fluorescent-antibody assay for rapid detection of human cytomegalovirus infection
Authors: Boppana S B , Smith R J , Stagno S , Britt W J
Published: January 2020

The use of monoclonal antibody (MAb) p63-27, which is reactive with the major immediate-early human cytomegalovirus (HCMV) protein pp72, was explored for the rapid diagnosis of HCMV viruria. The rapid assay detected all but 1 of 19 specimens identified by standard virus isolation methods from 1,676 newborn urine specimens, achieving a sensitivity of 94.5% and a specificity of 100%. The monoclonal antibody recognized 260 randomly obtained clinical isolates of HCMV, indicating the suitability of this reagent for use in screening assays. The sensitivity of the microtiter plate method declined rapidly for specimen from older infants and children with congenital CMV infection and virus-infected children attending a day-care center and was judged to be unacceptable for screening populations in this age group.
Evaluation of the Genetic Systems Corp. Integra HIV-1 Pageblot system
Authors: Starkey C A , Yen-Lieberman B , Proffitt M R
Published: January 2020

The Genetic Systems Corp. Integra HIV-1 Pageblot system was evaluated as a supplementary assay to confirm the presence of antibodies to human immunodeficiency virus type 1 in 57 specimens from individuals at high risk of infection with the virus. Forty-one specimens identified as reactive in the Genetic Systems Integra HIV-1 Pageblot system were likewise identified as reactive in a U.S. Food and Drug Administration-licensed (Biotech/Dupont) Western blot (immunoblot). Six specimens identified as indeterminate in either or both immunoblot assays were all identified as nonreactive in a U.S. Food and Drug Administration-licensed enzyme immunoassay with recombinant antigens.
Taxonomical implications of the emergence of high frequency of occurrence of 2,4-diamino-6,7-diisopropylpteridine-resistant strains of Vibrio cholerae from clinical cases of cholera in Calcutta, India
Authors: Ramamurthy T , Pal A , Pal S C , Nair G B
Published: January 2020

Of the 110 consecutive isolates of Vibrio cholerae recovered from cholera patients admitted to the Infectious Diseases Hospital, Calcutta, India, between July 1989 and October 1990, 90 and 82.7% were resistant to 10 and 150 micrograms of 2,4-diamino-6,7-diisopropylpteridine (O/129), respectively. Additionally, all O/129-resistant strains of V. cholerae were multiply resistant to antimicrobial agents. Except in the cases of four strains, resistance to O/129 was invariably linked with resistance to co-trimoxazole. Although O/129 susceptibility is still a useful test for Vibrio identification, resistance of V. cholerae to this compound in local areas might occasionally pose a problem.
Viability of fungal cultures maintained at -70 degrees C
Authors: Pasarell L , McGinnis M R
Published: January 2020

One thousand four hundred forty-seven clinical and environmental isolates of molds, yeasts, aerobic actinomycetes, and algae belonging to 164 genera (382 taxa) maintained on potato dextrose agar at -70 degrees C for periods ranging from 6 months to 13 years were subcultured and then incubated at 25 degrees C to determine their viabilities. Thirty-three isolates, Alternaria alternata (n = 1), Apophysomyces elegans (n = 1), Bipolaris spicifera (n = 1), Blastomyces dermatitidis (n = 4), Cokeromyces recurvatus (n = 1), Coremiella cubispora (n = 1), Cryptococcus ater (n = 1), Curvularia sp. (n = 1), Exserohilum monoceras (n = 1), Exserohilum pedicillatum (n = 1), Exserohilum rostratum (n = 1), Filobasidium floriforme (n = 1), Madurella mycetomatis (n = 1), Oedocephalum spp. (n = 2), Penicillium marneffei (n = 1), Pseudomicrodochium spp. (n = 4), Saksenaea vasiformis (n = 1), Sporothrix sp. (n = 1), and Mycelia Sterilia (n = 8), did not grow after repeated attempts at subculturing. Neither time in storage nor taxonomic classification was associated with a lack of viability. Storage at low temperature for either short or long periods of time is an excellent method for maintaining most medically important fungi.
Hemagglutination by a human rotavirus isolate as evidence for transmission of animal rotaviruses to humans
Authors: Nakagomi O , Mochizuki M , Aboudy Y , Shif I , Silberstein I , Nakagomi T
Published: January 2020

Human rotavirus strain Ro1845, which was isolated in 1985 from an Israeli child with diarrhea, has a hemagglutinin that is capable of agglutinating erythrocytes from guinea pigs, sheep, chickens, and humans (group O). Hemagglutination was inhibited after incubation with hyperimmune sera or in the presence of glycophorin, the erythrocyte receptor for animal rotaviruses. These results suggest that Ro1845 is an animal rotavirus that infected a human child.
Proposed interpretive criteria and quality control parameters for ofloxacin susceptibility testing of Neisseria gonorrhoeae
Authors: Fuchs P C , Barry A L , Baker C , Murray P R , Washington J A
Published: January 2020

A multilaboratory study designed to determine the in vitro susceptibility criteria and quality control parameters for ofloxacin against Neisseria gonorrhoeae was conducted according to the guidelines of the National Committee for Clinical Laboratory Standards. Proposed susceptibility breakpoints are MICs of less than or equal to 0.25 microgram/ml for the agar dilution test and greater than or equal to 31 mm for the disk diffusion test. A category for resistance could not be defined. Proposed acceptable quality control MICs for N. gonorrhoeae ATCC 49226 and Staphylococcus aureus ATCC 29213 range from 0.004 to 0.03 microgram/ml and 0.25 to 1.0 microgram/ml, respectively. With 5-micrograms ofloxacin disks, acceptable inhibitory zone diameters for S. aureus ATCC 25923 and the N. gonorrhoeae control strains range from 22 to 27 mm and 43 to 51 mm, respectively.
Distinguishing between respiratory syncytial virus subgroups by protein profile analysis
Authors: Walpita P , Mufson M A , Stanek R J , Pfeifer D , Connor J D
Published: January 2020

We subgrouped 75 strains of respiratory syncytial virus by a protein profile method (PPM) which relies on different mobilities of the phosphoprotein in one-dimensional polyacrylamide gel electrophoresis and does not require monoclonal antibodies. When compared with enzyme immunoassay, PPM correctly subgrouped 54 of 56 subgroup A and all 19 subgroup B strains.
Monoclonal antibodies to a specific 54-kilodalton antigen of Nocardia spp
Authors: Boiron P , Stynen D , Belkacem G , Goris A , Provost F
Published: January 2020

Two monoclonal antibodies (MAbs) of the immunoglobulin G2A isotype, reacting with a Nocardia-specific 54-kDa antigen, were generated. As determined by Western blot (immunoblot), both MAbs reacted only with the 54-kDa band. As determined by indirect immunofluorescence or enzyme immunoassay with whole microorganisms, the MAbs did not react with Nocardia cells. One of the MAbs showed weak cross-reactivity with mycobacterial antigens, while the other showed no cross-reactivity.
Chemiluminescent universal probe for bacterial ribotyping
Authors: Gustaferro C A , Persing D H
Published: January 2020

We describe a novel procedure for direct covalent coupling of horseradish peroxidase to rRNA and ribotyping by using enhanced chemiluminescence. Compared with their 32P-end-labeled counterparts, chemiluminescent rRNA probes are stable and easy to synthesize and provide results as good as or superior to those obtained with isotopic labeling. Direct chemiluminescent labeling of Escherichia coli rRNA produces a sensitive, universal probe suitable for clinical laboratory use in the investigation of nosocomial outbreaks.
Survival of hepatitis A virus on human hands and its transfer on contact with animate and inanimate surfaces
Authors: Mbithi J N , Springthorpe V S , Boulet J R , Sattar S A
Published: January 2020

The survival of hepatitis A virus (HAV; strain HM175) on the hands of five volunteers was determined by depositing 10 microliters of fecally suspended virus on each fingerpad and eluting the inoculum after 0, 20, 60, 120, 180, and 240 min. The amount of virus recovered from each fingerpad at 0 min was approximately 6.0 x 10(4) PFU. At the end of 4 h, 16 to 30% of the initially recoverable virus remained detectable on the fingerpads. HAV inocula (10 microliters; approximately 1.0 x 10(4) PFU) placed on fingerpads or 1-cm-diameter metal disks were used to determine virus transfer to clean surfaces upon a 10-s contact at a pressure of nearly 0.2 kg/cm2. When the inoculum was dried for 20 min, virus transfer from fingerpad to fingerpad, fingerpad to disk, and disk to fingerpad ranged from 2,667 to 3,484 PFU, while 0 to 50 PFU could be transferred after 4 h of drying. Elevation of the contact pressure alone from 0.2 to 1.0 kg/cm2 resulted in an approximately threefold increase in the amount of virus transferred. Incorporation of friction (10 half turns of the finger during 10 s of contact) with the low and high levels of pressure gave two- and threefold increases in the PFU of virus transferred, respectively. Pressure and friction were found to significantly affect HAV transfer (F = 33.98; P less than 0.05), irrespective of the mode of transfer used. No statistically significant interaction was observed between mode of transfer and pressure or friction. The findings of this quantitative study suggest that human hands may play an important role in the direct as well as the indirect spread of HAV.
Examination of whether persistently indeterminate human immunodeficiency virus type 1 Western immunoblot reactions are due to serological reactivity with bovine immunodeficiency-like virus
Authors: Whetstone C A , Sayre K R , Dock N L , VanDerMaaten M J , Miller J M , Lillehoj E , Alexander S S
Published: January 2020

The bovine lentivirus, known as bovine immunodeficiency-like virus (BIV), is genetically, structurally, and antigenically related to human immunodeficiency virus type 1 (HIV-1). It is not known whether sera from persons exposed to BIV proteins would show either positive or indeterminate reactivity on HIV-1 antibody tests. We used a BIV Western blot (immunoblot) analysis to examine human sera characterized as HIV-1 antibody positive, HIV-1 antibody negative, HIV-1 persistently indeterminate, HIV-1 p17 antibody positive only, HIV-1 p24 antibody positive only, human T-cell leukemia virus type 1 (HTLV-1) p19 antibody positive only, or HTLV-1 p24 antibody positive only. None of these sera were positive by Western blot to BIV-specific proteins. Many of these sera, however, displayed strong reactivities to bovine cell culture antigens on blots prepared from both mock-infected and BIV-infected cell cultures. The HIV-1 p17 and p24 antibody-positive and the HTLV-1 p19 and p24 antibody-positive sera were further examined by Western blot to bovine leukemia virus (BLV) and were found to be negative. We examined sera from laboratory personnel at risk for BIV exposure, including two laboratory workers who were exposed to BIV by accidental injection with BIV-infected cell culture material, and found no evidence of seroconversion to BIV-specific proteins. We tested 371 samples of fetal bovine sera, each sample representing serum pooled from one to three fetuses. All samples were negative by BIV Western blot. To date, we have not detected any human sera with antibody to BIV-specific proteins. Our data indicate that persistently indeterminate results on HIV-1 Western blot are not caused by a human antibody response to BIV proteins.
Distribution of serotypes of human rotavirus in different populations
Authors: Woods P A , Gentsch J , Gouvea V , Mata L , Santosham M , Bai Z S , Urasawa S , Glass R I
Published: January 2020

Serotyping is a useful tool to study the epidemiologic characteristics of rotaviruses in large populations and to assess the need for a vaccine to protect against all strains. By using an enzyme immunoassay with serotype-specific monoclonal antibodies to the four most common rotavirus serotypes, we analyzed 1,183 rotavirus-positive specimens from 16 stool collections in eight countries on four continents that were obtained from 1978 to 1989. Of the 926 strains (78%) that could be serotyped, 48% were serotype 1, 8% were serotype 2, 15% were serotype 3, and 7% were serotype 4. Twenty-two percent had insufficient numbers of double-shelled virus particles to react with the monoclonal antibody of the VP4 rotavirus protein and therefore could not be serotyped. Our results indicate that vaccines being developed must provide the greatest coverage against serotype 1 and that the serotype distribution cannot be predicted currently by the geographic area or prevalence in the preceding year.
Identification of Chlamydia pneumoniae by DNA amplification of the 16S rRNA gene
Authors: Gaydos C A , Quinn T C , Eiden J J
Published: January 2020

Chlamydia pneumoniae is an important cause of respiratory disease in humans, but diagnosis of C. pneumoniae is hindered by difficulties in the in vitro growth of the organism. In order to improve detection and identification, we recently developed a polymerase chain reaction (PCR) assay which uses oligonucleotide primers specific for C. pneumoniae. The nucleic acid sequence was determined for the 16S rRNA of C. pneumoniae, and regions in which C. pneumoniae differed from both Chlamydia psittaci and Chlamydia trachomatis were identified. Oligonucleotide primers corresponding to these unique regions were then synthesized and used in a PCR for the detection of C. pneumoniae. The C. pneumoniae-specific primers permitted the identification of six isolates of C. pneumoniae, but no reaction was observed with the 15 serovars of C. trachomatis or two strains of C. psittaci. PCR should prove to be valuable in confirming the identification of C. pneumoniae and in the diagnosis of C. pneumoniae infections.
Analysis of rRNA restriction fragment length polymorphisms from Bacteroides spp. and Bacteroides fragilis isolates associated with diarrhea in humans and animals
Authors: Smith C J , Callihan D R
Published: January 2020

The Escherichia coli rRNA operon rrnB was used as a 32P-labeled hybridization probe in Southern blots of genomic DNAs from representative strains of the saccharolytic, gram-negative, obligate anaerobes of the genus Bacteroides. Control experiments with the B. fragilis type strain ATCC 25285 established that nearly identical rRNA fragment patterns were produced when either the E. coli rrnB gene probe or homologous rRNA isolated from B. fragilis was used as the probe. In addition, it was shown that a specific 16S or 23S rrnB gene probe also could be used to produce fragment patterns suitable for analysis. Thirty-one strains from 8 of the 10 recognized Bacteroides species were then examined. The resulting autoradiographs revealed specific fragment patterns for all but one (B. ovatus) of the species tested. Restriction fragment length polymorphisms were observed for many of the strains tested, but these differences did not hinder species classification. The five B. ovatus strains examined did not form a distinct group, and their rRNA fragment patterns displayed a marked heterogeneity. The same approach was applied to a unique set of enterotoxin-producing B. fragilis strains isolated from animals and humans with diarrhea. The results demonstrated that these strains were in fact B. fragilis and that they produce rRNA fragment patterns closely related to those of the type strain ATCC 25285. This set of strains did not appear to form a separate subgroup or genotype within the B. fragilis species, and there were no distinguishable restriction fragment length polymorphisms that could be used to specifically separate enterotoxin-producing strains from nonenterotoxigenic strains.