bluuebunny/crossref_metadata_2025 huggingface dataset:
Reliable confirmation of antibodies to human immunodeficiency virus type 1 (HIV-1) with an enzyme-linked immunoassay using recombinant antigens derived from the HIV-1 gag, pol, and env genes
Authors: Ng V L , Chiang C S , Debouck C , McGrath M S , Grove T H , Mills J
Published: January 2020

An enzyme-linked immunoassay (ELISA) using six recombinant proteins corresponding to large segments of the human immunodeficiency virus type 1 (HIV-1) gag, pol, and env gene products (HIVAGEN; SmithKline Bio-Science Laboratories, Van Nuys, Calif.) was developed to confirm the presence of antibodies to HIV-1 in sera reactive in the whole-cell-derived virion screening ELISAs. Serum samples for testing were obtained from healthy seronegative blood donors and from the different categories of HIV-infected individuals (asymptomatic, acquired immunodeficiency syndrome [AIDS]-related complex, and AIDS). A positive reaction was defined as reactivity against an env and at least one other (either gag or pol) HIV-1 gene product; negative was defined as no reaction with any antigen; and indeterminate was defined as reactivity with gag or pol (or both) or with env alone. None of the 1,180 serum samples from healthy seronegative blood donors gave a positive result, and only 49 of these samples (4%) gave indeterminate results. The recombinant HIV-1 antigen ELISA panel identified seropositive individuals with a high degree of accuracy, as a positive reaction was seen with 99.3% of asymptomatic healthy seropositive individuals, 98.1% of patients with AIDS-related complex, and 90.4% of patients with AIDS. None of the 725 HIV-1-seropositive subjects had a negative test result. Reactivity with the Kp41 antigen, corresponding to an amino-terminal portion of the gp41 envelope glycoprotein, by itself demonstrated 100% sensitivity and specificity in distinguishing seronegative from seropositive sera. A subset of seronegative and seropositive samples were tested both with the recombinant HIV-1 antigen ELISA panel and by Western blot (Du Pont Co.). The recombinant HIV-1 antigen ELISA panel accurately identified more seropositive and seronegative samples and had fewer indeterminate results than did Western blot (interpreted by Du Pont criteria).
Immunochemical identification and biological characterization of cytotoxic necrotizing factor from Escherichia coli
Authors: De Rycke J , Phan-Thanh L , Bernard S
Published: January 2020

The aim of this study was to identify Escherichia coli cytotoxic necrotizing factor (CNF) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis and to investigate the possible dissociation of CNF from hemolysin (Hly), which is often produced by CNF-producing strains. CNF was purified from cell lysates of a CNF-producing strain by using ammonium sulfate precipitation, ion-exchange chromatography, gel filtration, and preparative nondenaturing PAGE. All eluates from successive longitudinal slices of a preparative polyacrylamide gel were tested for cytoxicity and analyzed by SDS-PAGE; CNF activity was quantitatively correlated with a protein of 115 kilodaltons (kDa). This procedure increased both cytotoxicity and lethal activity (about 300-fold). We then compared SDS-PAGE protein patterns of enriched lysates from eight field and mutant E. coli strains producing both CNF and Hly, Hly alone, or neither; the 115-kDa band was present solely in CNF-producing strains, irrespective of Hly production. A neutralizing antiserum was produced against unpurified CNF from strain BM2-1 and then extensively adsorbed with cells and extracts of a CNF-defective mutant from BM2-1. The adsorbed antiserum possessed antitoxin activity and neutralized both lethal and necrotic effects of cell lysates from all the CNF-producing strains tested. In an immunoblot of enriched extract from BM2-1, the adsorbed antiserum recognized, besides the 115-kDa protein, another protein of 59 kDa, which was present in the CNF-defective mutant from BM2-1 and was not associated with cytotoxicity. We can conclude from these findings that CNF is a protein of 115 kDa associated with both cytotoxicity and in vivo toxicity, distinct from Hly, and present in all presumed CNF-producing strains tested.
Identification and partial characterization of Mycobacterium avium and Mycobacterium intracellulare by using DNA probes
Authors: Saito H , Tomioka H , Sato K , Tasaka H , Tsukamura M , Kuze F , Asano K
Published: January 2020

We attempted to identify the Mycobacterium avium complex (MAC) isolated in Japan by using DNA probes specific for M. avium or Mycobacterium intracellulare (Gen-Probe Rapid Diagnostic System for MAC; Gen-Probe, Inc., San Diego, Calif.). The source and drug susceptibility distributions were examined. This assay system proved to be rapid, sensitive, specific, and reliable for identification of MAC and of the species as either M. avium or M. intracellulare. The DNA probe test showed that of the generally accepted MAC serovars, serovars 1 to 6, 8 to 11, and 21 belonged to M. avium and 7 and 12 to 20 belonged to M. intracellulare. Moreover, with the DNA probe test we found that the distribution patterns of M. avium and M. intracellulare isolates in Japan differed depending on the district in which MAC was isolated. The ratio of M. avium was much higher in eastern Japan. In Tokai and Shimane districts, the ratio of M. avium and M. intracellulare isolates significant in human disease was related to that of isolates from soil and house dust (natural sources). In M. avium, human disease-associated isolates were more resistant to rifampin, streptomycin, and kanamycin than were isolates from natural sources. However, this source dependence was not evident for M. intracellulare. In human disease-associated MAC, M. avium isolates were more resistant to most agents, except for quinolones, than were M. intracellulare isolates.
Clinical experience with cytomegalovirus isolation using conventional cell cultures and early antigen detection in centrifugation-enhanced shell vial cultures
Authors: Leland D S , Hansing R L , French M L
Published: January 2020

A total of 1,915 clinical samples was inoculated by low-speed centrifugation into shell vials (Bartels Immunodiagnostics, Bellvue, Wash.) containing cover slip monolayers of MRC-5 fibroblasts. At 1 and 2 days postinoculation, one cover slip was stained by an indirect immunofluorescence technique using a monoclonal antibody (Biotech Research Laboratories for Dupont, Billerica, Mass.) to cytomegalovirus (CMV) early antigen (EA). Clinical samples were also inoculated into three MRC-5 or MRHF cell cultures which were observed for 30 days for the appearance of a cytopathic effect (CPE). Of 157 CMV-positive samples, 92 (59%) were identified by centrifugation-enhanced EA (CE-EA) and 131 (83%) produced a CPE. CE-EA was less sensitive than CPE for all types of samples, although 17% of CMV-positive samples were detected by CE-EA alone. Evaluation of the CMV status of patients with CE-EA-positive-CPE-negative samples indicated that these samples likely represented true CMV-positive results. The average elapsed time between culture inoculation and identification of CMV decreased as follows when both CE-EA and CPE, rather than CPE alone, were used: urines, 15 to 7 days; buffy coats, 18 to 9 days; lung samples, 13 to 8 days; throat samples, 18 to 7 days. Although CE-EA was less sensitive than 30-day cell culture, both CE-EA and CPE were identified as valuable in CMV detection, and neither could be discontinued without a decrease in the CMV isolation rate or an increase in the turnaround time.
Development of an enzyme-linked immunosorbent assay for equine infectious anemia virus detection using recombinant Pr55gag
Authors: Archambault D , Wang Z M , Lacal J C , Gazit A , Yaniv A , Dahlberg J E , Tronick S R
Published: January 2020

To provide more sensitive and convenient methods for the detection of equine infectious anemia virus (EIAV), we developed an enzyme-linked immunosorbent assay (ELISA) employing the EIAV gag precursor (Pr55gag) produced by using recombinant DNA techniques. The antigenic reactivity of the recombinant EIAV Pr55gag was found to be equivalent to that of the virion p24gag and elicited high-titered antiserum in rabbits. When a large number of horse sera were analyzed for the presence of antibodies to EIAV by this ELISA, a radioimmunoassay for EIAV p15gag, or the standard agar gel immunodiffusion test, there was 98.7% concordance among the assays. By using the ELISA it was possible to specifically detect antibodies earlier after experimental infection of horses with EIAV than with the other two tests. A competition ELISA developed in order to detect EIAV gag antigens was found to be approximately 15 times more sensitive than the radioimmunoassay for EIAV p15gag. Antigens of other animal lentiviruses as well as those of the prototype oncovirus failed to compete in this assay.
Serotype-related differences in production and type of heat-labile hemolysin and heat-labile cytotoxin of Actinobacillus (Haemophilus) pleuropneumoniae
Authors: Kamp E M , van Leengoed L A
Published: January 2020

Reference strains of serotypes 1 to 12 of Actinobacillus (Haemophilus) pleuropneumoniae were cultured in Eagle minimal essential medium with 10% Serum Plus. Culture supernatants were examined for cytotoxicity to alveolar macrophages and for the ability to hemolyze sheep erythrocytes. All strains except the reference strain of serotype 6 produced cytotoxin, whereas only serotypes 1, 5, 9, 10, and 11 produced hemolysin. Both cytotoxin and hemolysin appeared to be heat labile. Antisera were raised against cytotoxin- and hemolysin-containing culture supernatants of serotypes 1 to 11. Cross-neutralization studies revealed that the hemolysins were serologically homogeneous. In contrast, four serologically different cytotoxins were distinguished. One cytotoxin was produced by serotypes 1, 5, 9, and 11, and a second was produced by serotypes 2, 3, 4, and 8. A third cytotoxin was produced by serotypes 7 and 12; this cytotoxin was related to the cytotoxins of serotypes 1, 2, 4, 5, 9, and 11. A fourth cytotoxin, produced by serotype 10, was related to the cytotoxin of serotypes 1, 5, 9, and 11. Seventy field strains belonging to serotypes 2, 3, 7, 8, 9, and 11 were also tested for production of cytotoxin and hemolysin. All strains belonging to serotypes 9 and 11 produced hemolysin and cytotoxin, whereas all strains of serotypes 2, 3, 7, and 8 produced only cytotoxin. Hemolysins and cytotoxins of both the field strains and the corresponding serotype reference strains were comparably neutralized. These findings strongly suggest that the observed differences in production and type of hemolysin and cytotoxin were related to serotype and not to strain.
Etiology of childhood diarrhea in Korea
Authors: Kim K H , Suh I S , Kim J M , Kim C W , Cho Y J
Published: January 2020

To assess the role of recently recognized enteropathogens in childhood diarrhea in Korea, 231 children with diarrhea admitted to and 104 children without diarrhea seen at the well-baby clinic or the outpatient department of Hanyang University Hospital in Seoul, Korea, were evaluated during a 14-month period. Stools were cultured for bacterial pathogens, including enterotoxigenic (heat-labile and heat-stable enterotoxin-producing) and enteroadherent organisms. Only those stools obtained from patients with diarrhea were examined for rotavirus. All Escherichia coli isolates were screened for Shiga-like toxin (SLT) I, SLT-II, enterohemorrhagic E. coli fimbriae, and enteroinvasiveness by colony hybridization. One or more pathogens were identified in 75.8% of the children with diarrhea. Rotavirus was the most frequently identified pathogen, accounting for 47% of the cases. Other major enteropathogens were enterotoxigenic E. coli (22%), Clostridium difficile (16%), enteroadherent E. coli (15%), and enteropathogenic E. coli (6%). Shigella spp., Campylobacter jejuni, Salmonella spp., SLT-I-and enterohemorrhagic-E. coli-fimbria-probe-positive E. coli serotype O26:H11 and enteroinvasive E. coli were isolated from only a few patients. Aeromonas hydrophila and E. coli O157 were not isolated. Compared with those of the controls, the isolation rates of heat-stable-enterotoxin-producing E. coli (P less than 0.05), C. difficile (P less than 0.025), and enteroadherent E. coli (P less than 0.05) were significantly higher in the patients with diarrhea. The greatest number of rotavirus, enterotoxigenic E. coli, and C. difficile cases were identified during the cool, dry months of October and November.
Cell surface protein antigen from Wolinella recta ATCC 33238T
Authors: Kokeguchi S , Kato K , Kurihara H , Murayama Y
Published: January 2020

A high-molecular-weight (approximately 150,000) protein was selectively isolated by acid extraction from the cell surface of Wolinella recta and purified by negative adsorption on DEAE-cellulose and gel filtration. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis showed that this protein was found in W. recta but not in other Wolinella species, such as W. curva and W. succinogenes. Sera from patients with periodontitis reacted strongly with this protein antigen, whereas sera from healthy donors showed little or no reactivity, as determined by immunoblotting analysis. In serum, titers of immunoglobulin G antibodies to the protein antigen were significantly higher in patients with periodontitis than in periodontally healthy donors, as detected by an enzyme-linked immunosorbent assay.
Rapid flow cytometric bacterial detection and determination of susceptibility to amikacin in body fluids and exudates
Authors: Cohen C Y , Sahar E
Published: January 2020

A flow cytometry-based method for rapid and quantitative detection of bacteria in various clinical specimens and for rapid determination of antibiotic effect is described. Achieving such a measurement with high sensitivity required discrimination between bacteria and other particles which were often present in clinical samples in high concentrations. This discrimination was facilitated by detecting the bacterial characteristic light scatter and fluorescence signals following staining, e.g., with the fluorescent nucleic acid-binding dye ethidium bromide, as well as by measuring bacterial proliferation during short time intervals. Antibiotic susceptibility was measured by observing the inhibition of such proliferation. The method was applied to 43 clinical specimens from various sources, such as wound exudates, bile, serous cavity fluids, and bronchial lavage. Bacterial detection, achieved in less than 2 h, agreed with results of conventional methods with a sensitivity of 74% and a specificity of 88%. Susceptibility to amikacin was detected in 1 h in 92% of 13 positive specimens.
Typing of Aeromonas strains by DNA restriction endonuclease analysis and polyacrylamide gel electrophoresis of cell envelopes
Authors: Kuijper E J , van Alphen L , Leenders E , Zanen H C
Published: January 2020

The outer membrane protein (OMP) composition (OMP typing) of 46 fecal Aeromonas strains from hybridization groups (HGs) 1 (A. hydrophila; n = 10), 4 (A. caviae; n = 16), and 8 (A. veronii; n = 20) were examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis as a phenotypic typing method. Almost every isolate of HG-1 and HG-8 had a unique OMP profile, in contrast to isolates of HG-4, which were separated into five different OMP types. It was possible to recognize HGs 1, 4, and 8 by OMP profiles. Twenty-three Aeromonas strains from HGs 1 (n = 5), 4 (n = 10), and 8 (n = 8) were tested by whole-cell DNA restriction endonuclease analysis (REA) as a genetic typing method. All strains tested by REA (with SmaI) had different DNA digestion patterns. Although additional DNA-rRNA hybridization analyses with SmaI and 16S and 23S rRNAs from Escherichia coli showed a reduction in the number of restriction bands to 8 to 13 hybridized fragments, the discriminative value was less when compared with that obtained by REA. The individual differences found by REA were used to analyze whether patients remained colonized by the same Aeromonas strain. Of 11 patients with diarrhea, 2 had a different isolate on repeat culture. In addition, one of nine tested fecal samples contained two Aeromonas isolates with different REA patterns. These results indicate that during diarrheal disease the intestinal tract may be colonized simultaneously with different Aeromonas isolates.
Development of quality control procedures for the human immunodeficiency virus type 1 antibody enzyme-linked immunosorbent assay
Authors: Kudlac J , Hanan S , McKee G L
Published: January 2020

A standardized pool of human sera that was positive for human immunodeficiency virus type 1 (HIV-1) antibody was developed. This positive control serum was used to analyze test differences among eight laboratories, among the HIV-1 antibody test kits of three different manufacturers, among different lots of the same test kit, and among pipetting devices and techniques. The standardized pool of human sera was tested 327 times by the different laboratories. In terms of positive tests, a reproducibility of 99.69% was achieved; however, significant test variance among laboratories, among test kit lots, and among pipetting devices and techniques could be demonstrated if the tests were compared on the basis of the net positive optical density (OD) value. This value was calculated by subtracting the cutoff OD value (i.e., the value below which an OD value was considered negative for HIV-1 antibody) from the observed OD value of the standardized pool of human sera. The results obtained suggest that this strategy can be used for proficiency testing, for monitoring the quality of HIV-1 antibody enzyme-linked immunosorbent assay reagents, and for evaluating pipetting devices and techniques.
Enteropathogenic Escherichia coli serotype O111:HNT isolated from preterm neonates in Nairobi, Kenya
Authors: Senerwa D , Olsvik O , Mutanda L N , Lindqvist K J , Gathuma J M , Fossum K , Wachsmuth K
Published: January 2020

This investigation was initiated as a consequence of several cases of diarrhea in a nursery ward for preterm babies in Nairobi, Kenya. Ten lactose-positive colonies were isolated from the stools of each of 30 neonates, regardless of whether they had diarrhea; 229 strains were identified as Escherichia coli and 65 strains were identified as Klebsiella pneumoniae. Six strains were lost during laboratory handling. No other bacterial, viral, or parasitic enteropathogens were identified. Using synthetic alkaline phosphatase-labeled probes, the bacterial isolates were found to be negative for the presence of genes coding for heat-stable and heat-labile enterotoxins. Seventy-eight E. coli strains isolated from a total of 13 neonates possessed the E. coli enteropathogenic adhesion factor (EAF) gene, as demonstrated by the use of a cloned radiolabeled DNA fragment probe. These strains possessed similar plasmid profiles constituting a core plasmid profile, and while all adhered to HeLa cells, none produced Vero cell cytotoxins. The EAF gene was located on a 65-megadalton plasmid. Serotyping showed the strains to be of serogroup O111 and serotype H nontypable, a well known enteropathogenic type. Five neonates died during the outbreak, and the fatality rate was 30.7% (4 of 13) for neonates infected with EAF-positive E. coli strains compared with 7.7% (1 of 13) for neonates from whom only EAF-negative E. coli strains were isolated. K. pneumoniae only was isolated from five neonates.
Antibody response to low-molecular-weight antigens of Aspergillus fumigatus in allergic bronchopulmonary aspergillosis
Authors: Kurup V P , Greenberger P A , Fink J N
Published: January 2020

Sera from patients with allergic bronchopulmonary aspergillosis (ABPA) or aspergilloma and normal sera were analyzed for specific antibodies by Western (immuno-) blotting with Aspergillus fumigatus antigens transferred electrophoretically onto polyvinylidene difluoride membranes. Western blot analysis demonstrated consistent reactivity of low-molecular-weight A. fumigatus antigens against ABPA sera but not against uncomplicated aspergilloma or normal sera. None of these low-molecular-weight components had any lectin-binding activity. Sera from patients with aspergilloma, however, frequently reacted with high-molecular-weight components of A. fumigatus. The majority of these high-molecular-weight antigenic components demonstrated concanavalin A-binding activity. The low-molecular-weight bands were discernible in Western blots with sera from all ABPA patients irrespective of disease activities, such as relapse, flare, or treatment. Antibodies detected by methods such as immunodiffusion or enzyme-linked immunosorbent assays demonstrated total antibody responses to most or all antigenic components, while Western blots demonstrated the reactivities of the individual components with the specific antibodies. Western blot analysis thus provided more information for immunodiagnosis of ABPA than other methods, especially when only crude antigens were available.
Incidence of catheter-associated gram-negative bacteremia in children with short bowel syndrome
Authors: Piedra P A , Dryja D M , LaScolea L J
Published: January 2020

Children with catheter-associated bacteremia were evaluated for the type of bacteria recovered and the relationship of the bacteria to the predisposing disease. A previously unrecognized observation was that gram-negative isolates, namely, Escherichia coli and Klebsiella sp., were almost exclusively recovered (11 of 12 isolates [92%]) from children with short bowel syndrome (SBS) compared with those from children with other underlying diseases, such as inflammatory bowel disease, malignancies, and other disorders (P less than 0.001). Furthermore, children with SBS had a higher frequency of repeated infection (3.1 catheter-associated infections compared with 1.3 catheter-associated infections in children with other disorders during the same period). Only gram-positive bacteria were isolated from children with malignancies and other predisposing disorders. The very high frequency of catheter-associated gram-negative bacteremia in children with SBS compared with that in children with other bowel disorders, malignancies, and other predisposing diseases requires attention by the clinician in the management of patients in this group.
Molecular epidemiology and restriction site mapping of adenovirus type 3 genome types
Authors: Adrian T , Best B , Hierholzer J C , Wigand R
Published: January 2020

One hundred thirty-eight strains of adenovirus type 3 isolated from patients in the United States, West Germany, and other regions between 1961 and 1984 were analyzed with six restriction endonucleases; 18 genome types were found. BglII was the most discriminative enzyme. Mapping of altered restriction sites was also performed for all six enzymes. The genome types D1 (like the prototype) and D3 prevailed among 45 and 47 strains, respectively. All genome types could be divided into two groups related to D1 or D3. Several clusters of infections by strains with the same genome type were observed. Only hints of differences were found in the pathogenicities of individual genome types. D1 strains were present in the United States and in Europe; group D3 prevailed almost exclusively in the United States.
Differential antibody responses to Plasmodium falciparum and P. vivax circumsporozoite proteins in a human population
Authors: Burkot T R , Graves P M , Wirtz R A , Brabin B J , Battistutta D , Cattani J A , Maizels R M , Alpers M P
Published: January 2020

Papua New Guineans exposed to hyperendemic malaria in the Madang area showed different antibody responses to Plasmodium falciparum and Plasmodium vivax sporozoites despite comparable entomological inoculation rates. Although there was a significant trend of increasing prevalence of anti-P. falciparum circumsporozoite (CS) protein immunoglobulin G (IgG) with age, there was no significant increase in the antibody units of IgG recognizing P. falciparum CS proteins. Antibodies recognizing P. vivax CS proteins steadily increased in prevalence and antibody units with age. Significant trends of increasing prevalence of antibody responders (both IgG and IgM) with increasing splenic enlargement were found in the younger age groups for P. falciparum CS proteins but not for P. vivax CS proteins. When antibody responders were analyzed by quartiles, there was a trend of increasing antibody response with age against P. vivax CS peptide, but not for P. falciparum CS protein. There was no evidence for increasing protection against blood-stage infections with increasing antibody levels for either P. falciparum or P. vivax. Neither were any significant relationships found between entomological inoculation rates and either CS antibody prevalence or concentration among the villages studied.
Evaluation of blood collected on filter paper for detection of antibodies to human immunodeficiency virus type 1
Authors: Fortes P , Menitove J , Ross A , Steece R , Cabrian K , Ferrera C , Perkins P A , Sturge J , Lealos R , Krieger M S
Published: January 2020

This study was undertaken to evaluate the sensitivity and specificity of a commercial enzyme immunoassay in detecting antibody to human immunodeficiency virus type 1 using whole-blood specimens collected onto filter paper. Results obtained with specimens collected onto filter paper were comparable with those obtained with the corresponding serum or plasma specimens.
Modifications of a Candida albicans biotyping system
Authors: Childress C M , Holder I A , Neely A N
Published: January 2020

Methods of preparing and plating inocula onto media for the Odds and Abbott Candida albicans biotyping system (F. C. Odds and A. B. Abbott, Sabouraudia 18:301-317, 1980; Odds and Abbott, Sabouraudia 21:79-81, 1983) were altered to utilize inexpensive and commercially available supplies and equipment. The modified system correlates well with the reference system.
Bacteremia and infection of a hip prosthesis caused by Bacillus alvei
Authors: Reboli A C , Bryan C S , Farrar W E
Published: January 2020

Of the 34 Bacillus species described, 10 have been reported to cause infection in humans and 6 are insect pathogens. We report a case of an infected prosthetic hip caused by Bacillus alvei, only the third documented case of human infection with this organism.
Comparison of the agar dilution, tube dilution, and broth microdilution susceptibility tests for determination of teicoplanin MICs
Authors: Kenny M T , Dulworth J K , Brackman M A
Published: January 2020

The purpose of this study was to evaluate the National Committee for Clinical Laboratory Standards agar dilution, tube dilution, and broth microdilution susceptibility tests for the measurement of teicoplanin MICs. The three standardized tests gave equivalent (within a twofold dilution) results with 98.8 to 99.0% of the 508 gram-positive clinical isolates tested, indicating that either method may be used for teicoplanin MIC determination.
Quality control slide for potassium hydroxide and cellufluor fungal preparations
Authors: Perry J L , Miller G R
Published: January 2020

An opaque, water-insoluble quality control material with a skinlike microscopic appearance was prepared by inoculating melted xanthine (0.4%) agar with filamentous fungi and dispensing drops onto glass slides. After solidification of the agar, the material was rapidly cleared by 10% KOH, revealing fungal elements stained by Cellufluor reagent.
Serum immunoglobulin A response to Norwalk virus infection
Authors: Erdman D D , Gary G W , Anderson L J
Published: January 2020

We describe the serum immunoglobulin A (IgA) antibody response to Norwalk virus infection in human volunteers and compare it with previously described IgM and total antibody responses. Whereas specific IgA and IgM peak within 2 weeks after onset of symptoms, titers of total blocking antibody continue to rise, implying mediation by IgG antibody.
Expression of gamma interferon on circulating lymphocytes in viral infections
Authors: Caruso A , Stellini R , Scalzini A , Pollara P , Foresti I , Bonfanti C , Scura G , Gonzales R , Manca N , Turano A
Published: January 2020

Peripheral blood lymphocytes from a total of 111 patients and 40 healthy individuals were studied for gamma interferon (IFN-gamma) expression on their surfaces by indirect immunofluorescence assay and flow cytometry, with a new anti-IFN-gamma monoclonal antibody (IGMB-14) as a specific reagent. Of 64 patients with proven acute viral infections, 59 had a significantly higher percentage of lymphocytes expressing IFN-gamma on their membranes than healthy individuals did. On the other hand, only 3 (8.9%) of 34 patients with proven bacterial infections had an increased percentage of IFN-gamma-expressing lymphocytes. None of the eight patients with other infections and none of the five with systemic lupus erythematosus showed an increased percentage of IFN-gamma-positive lymphocytes. The percentage of IFN-gamma-expressing lymphocytes during a viral infection was found to be related to different stages of the disease. Finally, some applications of this rapid IFN-gamma assay method in viral diseases are discussed.
Evidence for a 6.5-day minimum exoerythrocytic cycle for Plasmodium falciparum in humans and confirmation that immunization with a synthetic peptide representative of a region of the circumsporozoite protein retards infection
Authors: Murphy J R , Baqar S , Davis J R , Herrington D A , Clyde D F
Published: January 2020

Immunization with a synthetic peptide which is representative of part of the repeating region of Plasmodium falciparum circumsporozoite protein resulted in an immunity which allowed vaccinees to retard the development of patent malaria as compared to nonimmunized controls. Analysis of infection dynamics showed that immunity could be attributed to either neutralization of about 92% of inoculated sporozoites, delayed development of the majority of parasites, or a combination of neutralization and delayed development. In spite of this impressive antiplasmodial capacity, all volunteers after being bitten by infected mosquitoes developed malaria, and seven of eight developed parasitemia between 6.5 and 7.0 days after infective mosquito bites.
Routine viral culture for pediatric respiratory specimens submitted for direct immunofluorescence testing
Authors: Blanding J G , Hoshiko M G , Stutman H R
Published: January 2020

From 1986 to 1987, 69 (25%) of 274 specimens from children with lower respiratory tract syndromes were positive for respiratory syncytial virus antigen by direct immunofluorescence assay (DFA). Comprehensive viral culture was performed on all 205 DFA-negative specimens, and 72 specimens yielded viruses; 5 specimens yielded multiple agents. Thus, 52% of specimens yielded a specific virus, supporting the routine use of viral culture. Isolates from the DFA-negative specimens included respiratory syncytial virus (n = 7), rhinovirus (n = 34), hemadsorbing viruses (n = 13), cytomegalovirus (n = 11), adenovirus (n = 8), enteroviruses (n = 3), and herpes simplex virus (n = 2). Although serologic confirmation is needed, cytomegalovirus may be an underappreciated cause of acute lower respiratory tract infection in normal children. Further studies must be conducted to document this possibility.
Cross-sectional study of reverse transcriptase-inhibiting antibody as a marker of acquired immune deficiency syndrome
Authors: Advani M , Imagawa D T , Lee M H , Sano K , Morales F , Mitsuyasu R T , Detels R
Published: January 2020

A cross-sectional study of 128 individuals infected with human immunodeficiency virus type 1 (HIV-1) was conducted to determine the correlation of reverse transcriptase-inhibiting (RTI) antibody to clinical disease. Thirty-two individuals were studied in each of four clinical groups: asymptomatic individuals, those with persistent generalized lymphadenopathy, those with acquired immune deficiency syndrome (AIDS)-related complex, and those with AIDS. Our study showed that 78% of asymptomatic individuals, 53% of those with persistent generalized lymphadenopathy, 50% of those with AIDS-related complex, and only 25% of those with AIDS have RTI antibody. Concurrent measurement of measles antibody level was used as an indicator of the immune status of these individuals. Measles antibody did not decline in persons with clinical disease, but asymptomatic individuals had lower antibody titers, possibly due to hypergammaglobulinemia associated with advanced HIV infection. These results indicate that more HIV-infected asymptomatic individuals than symptomatic individuals have RTI antibody. This suggests either that the RTI antibody level decreases with the progression of disease in HIV infection or that symptomatic individuals do not produce RTI antibody. The presence or absence of RTI antibody can thus be used as a marker of advanced disease.
Location of double bonds in monounsaturated fatty acids of Campylobacter cryaerophila with dimethyl disulfide derivatives and combined gas chromatography-mass spectrometry
Authors: Moss C W , Lambert-Fair M A
Published: January 2020

Location of the double-bond position of monounsaturated fatty acids in Campylobacter cryaerophila was accomplished with combined gas chromatography-mass spectrometry analysis of dimethyl disulfide (DMDS) derivatives. The monoenoic fatty acids from whole bacterial cells were converted to methyl esters and then to DMDS adducts and analyzed by capillary gas chromatography-mass spectrometry. The mass spectra of DMDS adducts gave an easily recognizable molecular ion (M+) and two major diagnostic ions attributable to fragmentation between the two CH3S groups located at the original site of unsaturation. Two previously unidentified acids that distinguish C. cryaerophila from other bacteria were identified as C14:1 omega 7 and C16:1 omega 7 from their mass spectral fragmentation patterns. Resolution of cis and trans isomers by capillary column gas chromatography permitted assignment of the trans isomer to the C16:1 omega 7 acid.
Growth and endotoxin production of Yersinia enterocolitica and Enterobacter agglomerans in packed erythrocytes
Authors: Arduino M J , Bland L A , Tipple M A , Aguero S M , Favero M S , Jarvis W R
Published: January 2020

Since 1987, the Centers for Disease Control investigated six cases of transfusion-associated sepsis. All six patients developed septic shock after receiving units of packed erythrocytes (PRBCs) contaminated with Yersinia enterocolitica (five patients) and Enterobacter agglomerans (one patient); three of the blood recipients died. We studied the growth and endotoxin production of Y. enterocolitica and E. agglomerans in units of PRBCs stored at 4 degrees C for 60 days. When PRBCs were inoculated with 0.1 to 1.0 CFU of these organisms per ml, both Y. enterocolitica and E. agglomerans entered log-phase growth 2 to 3 weeks after inoculation; generation times were 15 and 22 h, respectively. Endotoxin was first detected at 3 weeks following inoculation, and the concentration paralleled the log phase of growth of the strains tested. These data show that prolonged storage of PRBCs at 4 degrees C provides conditions that allow these two organisms to grow and subsequently produce high concentrations of endotoxin.
Quantitative determination of staphylococcal enterotoxin A by an enzyme-linked immunosorbent assay using a combination of polyclonal and monoclonal antibodies and biotin-streptavidin interaction
Authors: Edwin C
Published: January 2020

A sandwich enzyme-linked immunosorbent assay to detect staphylococcal enterotoxin A (SEA) was developed by using monoclonal antibodies (MAb) to SEA as primary capture antibodies. The antigen was detected with purified rabbit anti-SEA antibody as the secondary antibody. The secondary antibody was identified by direct conjugation with biotin or via biotinylated sheep F(ab')2 fragments to rabbit antibody. The biotin was then reacted with avidin-alkaline phosphatase (AP) conjugate, avidin-biotin-AP conjugated complex, or streptavidin-AP conjugate. The enzyme was identified by using p-nitrophenylphosphate. The incorporation of the avidin-biotin-AP conjugated complex or streptavidin-AP conjugate augmented the sensitivity 32-fold over that of the enzyme-linked immunosorbent assay without these reagents. Controls were run by substitution of the anti-SEA MAb with unrelated MAb of the same isotype. Sample values were considered positive when the A405 exceeded those of the negative controls by 3 standard deviations (greater than 99% confidence interval). The toxin could be quantitated with purified SEA standards through linear regression analysis with lower detection limits of 4 ng/ml (r = 0.99) and 0.25 ng/ml (r greater than or equal to 0.98). Concentrations of protein A up to 10 micrograms/ml did not cause interference. Analyses of crude growth extracts of SEA-secreting strains of Staphylococcus aureus were reproducible and were expressed in terms of 95% confidence intervals. Lack of cross-reactivity was seen with extracts of other toxigenic and nontoxigenic strains of S. aureus. The assay can be completed in one working day, provided that MAb-coated plates are available.
Multisite evaluation of a monoclonal antibody reagent (Syva) for rapid diagnosis of cytomegalovirus in the shell vial assay
Authors: Jespersen D J , Drew W L , Gleaves C A , Meyers J D , Warford A L , Smith T F
Published: January 2020

A pre-cytopathic effect (CPE) monoclonal antibody reagent (Syva Co., Palo Alto, Calif.) was evaluated in four laboratories for the rapid detection of cytomegalovirus (CMV) in shell vial cell cultures at 16 to 24 h and 40 to 48 h postinoculation. Results were compared with those obtained by inoculation of the specimen into conventional tube cell cultures that were examined for the presence of typical CMV CPE and subsequently tested by reaction with the monoclonal antibody reagent in an indirect immunofluorescence test. Of 937 specimens, CMV was positive in 184 (20%). CMV was detected twice as frequently in shell vials only (n = 29) as in conventional tube cell cultures (n = 14). Pre-CPE shell vial assay was 91% sensitive (range, 84 to 98%) and 96% specific (range, 93 to 98%) compared with the detection of CPE in conventional tube cell cultures. Overall, 137 of 166 (83%) and 143 of 166 (86%) of the CMV strains were detected at 16 to 24 h and 40 to 48 h postinoculation, respectively. The Syva reagent produced sensitive and specific results for the rapid detection of CMV infection in shell vial cell cultures and reliably confirmed the presence of the virus as detected by CPE in conventional tube cell cultures.
Comparison of hemagglutinin and agglutinin schemes for the serological classification of Haemophilus paragallinarum and proposal of a new hemagglutinin serovar
Authors: Eaves L E , Rogers D G , Blackall P J
Published: January 2020

Ninety-five isolates of Haemophilus paragallinarum were classified serologically by a hemagglutinin serotyping scheme, and the results were compared with results from agglutinin serotyping of the same isolates. Of the isolates, 65 were from Australia, 5 were from Japan, 6 were from the Federal Republic of Germany, 7 were from the United States, and 12 were from South Africa. Seven hemagglutinin serovars, HA-1 to HA-7, corresponding to agglutinin serovars A (HA-1, HA-2, and HA-3), C (HA-4, HA-5, and HA-6), and B (HA-7) have been described. Only one of the seven hemagglutinin serovars was found among the Australian isolates. This was serovar HA-5, comprising 49 isolates. Fifteen Australian isolates, all from southeast Queensland and northern New South Wales, were found to belong to a new hemagglutinin serovar. This was designated HA-8 and represented a fourth subgroup of agglutinin serovar A. Of the 95 isolates examined, only 1 did not produce a hemagglutinating antigen and was nonserotypeable by the hemagglutinin system. This compared favorably with the agglutinin scheme, which serotyped only 60 of the 95 isolates, with 29 nonserotypeable and 6 autoagglutinating.
Alkaline phosphatase capture test for the rapid identification of Escherichia coli and Shigella species based on a specific monoclonal antibody
Authors: Husson M O , Mielcarek C , Izard D , Leclerc H
Published: January 2020

A specific monoclonal antibody for Escherichia coli and Shigella sp. alkaline phosphatase was used in an immunocapture assay and allowed identification of E. coli either in culture isolates or directly in clinical specimens. The assay was easy and required only four steps: (i) alkaline phosphatase was released within 10 min by using a gentle lysis procedure, (ii) cell lysates were transferred to antibody-coated tubes for 45 min, (iii) p-nitrophenyl phosphate substrate was added, and (iv) alkaline phosphatase activity was detected in a microsample spectrophotometer at 410 nm. This immunocapture assay was highly specific: only one false-positive reaction was observed with a Klebsiella pneumoniae lysate among the 205 non-E. coli strains tested. The assay was sensitive, detecting 10(7) CFU/ml from culture isolates or 10(5) CFU/ml from urine specimens which had first been grown in phosphate-limiting medium for 2 h. At these bacterial concentrations, the percentages of detected E. coli were high: 91% for blood cultures, 95.4% for culture isolates, and 96.8% for urine specimens.
Four-year study of rotavirus electropherotypes from cases of infantile diarrhea in Rome
Authors: Ruggeri F M , Marziano M L , Tinari A , Salvatori E , Donelli G
Published: January 2020

Rotavirus infections were detected in 210 of 675 children with acute diarrhea admitted to a major pediatric hospital in Rome from January 1982 through December 1985. Most of the patients with rotavirus infections were admitted during the winter season in both 1982 and 1985, whereas during the two intermediate years, cases occurred in all months. Among 84 rotavirus samples examined, 14 different electropherotypes were recognized, 2 of which largely predominated over the others. The two electropherotypes were particularly frequent in the 2 epidemic years, altogether accounting for 70.2% of the samples typed, and circulated in distinct periods. None of the viruses showed a short pattern of electrophoretic migration of the genome, indicating a minor involvement of subgroup I rotaviruses in hospitalization-requiring diarrheas occurring in the area surveyed.
Clinical and epidemiologic aspects of members of Aeromonas DNA hybridization groups isolated from human feces
Authors: Kuijper E J , Bol P , Peeters M F , Steigerwalt A G , Zanen H C , Brenner D J
Published: January 2020

Between June 1982 and May 1987 Aeromonas species were isolated from 208 of 34,311 (0.61%) fecal samples submitted to a Regional Public Health Laboratory in The Netherlands. Aeromonas isolates were found most frequently in summer and rarely in winter. Of 169 Aermonas isolates that were available for further study, 19% were isolated from patients with a mixed infection, 5% from patients with underlying diseases, and 15% from patients who used medication that could predispose the intestinal tract to colonization with Aeromonas species. Aeromonas species that produced cytotoxins to Vero cells (cytotoxigenic) were found in hybridization groups 1 (11% of all isolates), 2 (1%), 3 (2%), and 8 (25%) and were identified phenotypically as A. hydrophila or A. sobria. Aeromonas species that did not produce cytotoxins to Vero cells (noncytotoxigenic) were found in hybridization groups 4 (57%) and 5A (4%) and were identified phenotypically as A. caviae. Distribution of Aeromonas species by age showed a predominance of noncytotoxigenic strains in children under the age of 5 years (46% of all noncytotoxigenic strains), while cytotoxigenic strains were mainly cultured from patients aged 50 years or older (54% of all cytotoxigenic strains). Significant correlations were found between cytotoxigenic strains and hospitalization, foreign travel, and contact with surface water. Cytotoxigenic strains were isolated significantly more often than noncytotoxigenic strains from patients with diarrhea, but in a multivariate analysis including age, previous medication, underlying disease, and foreign travel, this association was not significant.
Isolation of a cell-detaching factor of Trichomonas vaginalis
Authors: Garber G E , Lemchuk-Favel L T , Bowie W R
Published: January 2020

The pathogenetic role of soluble products of Trichomonas vaginalis growth in culture is controversial. To evaluate this role, T. vaginalis was grown in broth and cell culture and the cell-free filtrate was applied to fresh cell culture monolayers. When adjusted to pH 6.5, filtrates obtained from 22-h culture growth totally disrupted McCoy, HEp-2, human foreskin fibroblast, and Chinese hamster ovary cell monolayers within 6 h. These detached cells remained greater than 90% viable. This cell-detaching factor (CDF) was heat and acid labile, with a pH optimum of 6.5. CDF has trypsinlike activity which disrupts monolayer cells, but cells do not die if the pH is controlled. CDF was purified by ethanol precipitation, ammonium sulfate fractionation, and ion-exchange and gel filtration column chromatography. A 200,000-molecular-weight glycoprotein which was also immunogenic by immunoblot with human sera reactive to T. vaginalis was isolated in this manner. This confirms the presence of a specific soluble CDF derived from T. vaginalis whose application may be important as a diagnostic tool and in further studies of pathogenesis.
Effects of growth temperature on the ingestion and killing of clinical isolates of Listeria monocytogenes by human neutrophils
Authors: Stecha P F , Heynen C A , Roll J T , Brown J F , Czuprynski C J
Published: January 2020

In this study, we compared three human isolates (F5380, Scott A, and Murray B) and one laboratory strain (EGD) of Listeria monocytogenes for their resistance to ingestion and killing by human neutrophils. We observed no substantial difference in killing among these strains when they were grown at 37 degrees C. Because it is likely that listerial growth occurs at lower temperatures during food-borne outbreaks of listeriosis, we also compared these strains after they were grown at 22 and 4 degrees C. A general reduction in the ability of human neutrophils to kill L. monocytogenes was observed as the temperature at which the listeriae were grown decreased. Growth at 4 degrees C significantly decreased the killing of all four strains of L. monocytogenes by human neutrophils; two strains (EGD and F5380) were more resistant to killing than were the other two strains (Scott A and Murray B). No obvious relationship was noted between the chemiluminescence response of neutrophils to opsonized listeriae and the ability of the neutrophils to kill listeriae in vitro. Growth at 4 degrees C, however, significantly increased the resistance of L. monocytogenes to killing by hydrogen peroxide.
Xenorhabdus luminescens (DNA hybridization group 5) from human clinical specimens
Authors: Farmer J J , Jorgensen J H , Grimont P A , Akhurst R J , Poinar G O , Ageron E , Pierce G V , Smith J A , Carter G P , Wilson K L
Published: January 2020

An unusual isolate from a human leg wound was identified as Xenorhabdus luminescens. This finding led to the discovery or isolation of four additional strains, two from blood and two from wounds. Three of the five strains were from patients in San Antonio, Tex. Three strains were studied by DNA-DNA hybridization (S1 nuclease-trichloroacetic acid method) and were 77 to 100% related to each other, 34% related to the type strain of X. luminescens, 35 to 40% related to three of Grimont's other DNA hybridization groups of X. luminescens, and 9% related to the type strain of Xenorhabdus nematophilus. The new group of five strains was designated X. luminescens DNA hybridization group 5. All five strains were very inactive biochemically and fermented only D-glucose and D-mannose. The key reactions for recognizing this new organism are yellow pigment production, negative test for nitrate reduction to nitrite, weak bioluminescence (10 to 15 min of dark adaptation is required to see the weak light produced), and a unique hemolytic reaction on sheep blood agar plates incubated at 25 degrees C. Two case histories of strains from wounds are given; these suggest that X. luminescens DNA hybridization group 5 may be a new bacterial agent that causes wound infections. The two cases of wound infection, along with the two blood isolates, suggest that the new organism is clinically significant.