bluuebunny/crossref_metadata_2025 huggingface dataset:
Comprehensive approach to identification of serovars of Mycobacterium avium complex
Authors: Denner J C , Tsang A Y , Chatterjee D , Brennan P J
Published: January 2020

Serotyping of nontuberculous mycobacteria, especially those of the Mycobacterium avium complex, provides important epidemiological information, particularly in tracing origins of infections. Seroagglutination with whole cells and polyclonal rabbit antibodies was the original way of identifying serovars and is still commonly used. The discovery of the glycolipid nature of the typing antigens allows differentiation of serovars on the basis of thin-layer chromatography of whole antigens and gas chromatography-mass spectrometry of the characteristic sugars of the oligosaccharide haptens of these antigens. In particular, the generation of monoclonal antibodies to the glycolipid antigens allows facile differentiation of serovars through enzyme-linked immunosorbent assay. All of these protocols were applied in developing a comprehensive approach to the typing of members of the M. avium complex.
Evidence for two serotype G3 subtypes among equine rotaviruses
Authors: Browning G F , Chalmers R M , Fitzgerald T A , Snodgrass D R
Published: January 2020

Ten cultivable equine rotavirus isolates, two of North American, six of British, and two of Irish origin, were compared with standard rotavirus strains and with each other by cross neutralization, neutralization with a panel of monoclonal antibodies (MAbs), hybridization to a simian rotavirus (SA-11) VP7 gene probe, and reaction with rotavirus subgrouping and serotyping MAbs in enzyme-linked immunosorbent assays. Six isolates, two of which had previously been serotyped as G3 by other workers, were found to be serotype G3; one was confirmed to be G5, and three were not related to serotypes G1 to G10. The serotype G3 strains were divisible into two subtypes, G3A and G3B, on the basis of cross neutralization. This division was also apparent in reactions with neutralizing VP7-specific MAbs and in the liquid hybridization assay. Two of the isolates were not bound by either subgroup MAb, six were bound by both subgroup I and II MAbs, and two were bound by only the subgroup I MAb. The assays used in this characterization provide a range of epidemiological information for use in future field investigations.
Sporothrix schenckii fungemia without disseminated sporotrichosis
Authors: Kosinski R M , Axelrod P , Rex J H , Burday M , Sivaprasad R , Wreiole A
Published: January 2020

Fungemia is a rare complication of Sporothrix schenckii infection and has always been associated with disseminated sporotrichosis. We describe an immunocompetent patient with localized lymphocutaneous sporotrichosis from whose blood the fungus was isolated. A lysis-centrifugation blood culture system may have improved our ability to detect low-level S. schenckii fungemia.
Local immune responses to the Campylobacter flagellin in acute Campylobacter gastrointestinal infection
Authors: Nachamkin I , Yang X H
Published: January 2020

Intestinal immunoglobulin A (IgA) anti-campylobacter flagellin antibodies were measured by an enzyme-linked immunosorbent assay in patients with and without campylobacter infections. The level of fecal IgA antiflagellin antibodies was significantly higher in patients than in controls, but only 12 of 29 patients (41.4%) with a campylobacter infection had detectable IgA antibodies against flagellin (optical density, greater than or equal to 0.100). Further testing of fecal IgA antibodies from 10 patients with homologous and heterologous purified flagellins showed strain specificity in 1 patient. Cross-reactivity of fecal IgA antibodies with heterologous flagellins did not correlate with the Lior serotype of the isolate.
A cluster of cases of Escherichia coli O157 infection in a day-care center in a communal settlement (Kibbutz) in Israel
Authors: Lerman Y , Cohen D , Gluck A , Ohad E , Sechter I
Published: January 2020

This is the first report in Israel of a cluster of cases of diarrhea associated with the isolation of Escherichia coli O157 in stool samples. E. coli O157:NM (nonmotile), producing verotoxins 1 and 2 was isolated from the stool samples of three infants less than 1 year of age with diarrhea and from a fourth asymptomatic infant staying in the same day-care class in a communal settlement (kibbutz). The clinical presentation included nonbloody diarrhea without other symptoms. Surveillance efforts should be enacted in order to define the relative importance of E. coli O157 infections in Israel.
Modification of the methodology of stool culture for Salmonella detection
Authors: Ruiz J , Sempere M A , Varela M C , Gomez J
Published: January 2020

A total of 4,284 H2S-positive colonies isolated on salmonella-shigella agar and 4,350 isolated on Hektoen agar were flooded with 5 microliters of a reagent (MUCAP test; Biolife Italiana S.r.l., Milan, Italy) and observed after 3 to 5 min for the development of fluorescence produced in the presence of the C8 esterase enzyme. All of the 794 colonies isolated on salmonella-shigella agar and the 752 isolated on Hektoen agar and identified as positive for Salmonella spp. with conventional biochemical tests were found positive with the MUCAP test (the sensitivity was 100% and the negative predictive value was 100% for both media). Moreover, only six isolates identified by conventional biochemical tests as Proteus vulgaris were MUCAP test positive (the specificity was 99.8% and the positive predictive value was 99.2% for both media). On the basis of these results, we propose the use of the MUCAP test as a method for the screening of H2S-positive colonies and only subculturing on Kliger agar of those colonies which are MUCAP test positive. The MUCAP test is a rapid method for the presumptive detection of Salmonella spp. and reduces the work and material involved in testing.
Development and clinical significance of a diagnostic assay based on the polymerase chain reaction for detection of human cytomegalovirus DNA in blood samples from immunocompromised patients
Authors: Zipeto D , Revello M G , Silini E , Parea M , Percivalle E , Zavattoni M , Milanesi G , Gerna G
Published: January 2020

The presence of human cytomegalovirus (HCMV) DNA in blood was investigated by the polymerase chain reaction (PCR) in 293 blood samples from 86 immunocompromised patients. Of the 86 patients, 23 underwent clinical and virologic follow-up for HCMV infection. In parallel, blood samples were examined for viremia and antigenemia. Concordant results between PCR and assays for viremia and antigenemia were obtained on 124 positive and 110 negative samples, with an overall concordance of 79.8%, while 59 samples (most from patients with HCMV infection) were positive by PCR alone. PCR is a new powerful tool for detection of HCMV infections in blood samples from immunocompromised patients. However, its clinical significance appears to be restricted to the indication of a risk of reactivation of HCMV infection.
Double sugar-tyrosine medium improves O-1 phage Salmonella screening
Authors: Thaller M C , Dainelli B , Berlutti F , Schippa S , Fontana C , Pezzi R
Published: January 2020

A modification of the procedure for O-1 phage Salmonella screening is presented. The novel method is based on the use of two media, i.e., a new medium (double sugar-tyrosine [DST]), which permits the combination of adonitol and sucrose fermentation and tyrosine clearing tests, and the previously described o-nitrophenyl-beta-D-galactopyranoside urease indole medium. In comparative trials, the new procedure and the conventional one were used to screen for Salmonella isolates from 553 lactose-negative strains of members of the family Enterobacteriaceae. The O-1 phage test, performed on DST medium, recognized the same number of phage-susceptible Salmonella strains as did the standardized method; however, it permitted the correct identification of a greater number of phage-resistant strains for discard (95.6 versus 85.3%). In particular, DST medium presented a higher efficacy than triple sugar iron agar (which is the corresponding medium in the reference procedure) in correctly identifying phage-negative cultures for discard (69.1 versus 28.5%).
Cefamandole-susceptible strains of Legionella pneumophila serogroup 1: implications for diagnosis and utility as an epidemiological marker
Authors: Vickers R M , Stout J E , Tompkins L S , Troup N J , Yu V L
Published: January 2020

The standard selective Legionella medium that contains cefamandole failed to grow legionella pneumophila serogroup 1, subtype Bellingham, from a sputum sample from a patient with nosocomial Legionnaires' disease; the isolate did grow on a similar selective medium that substitutes vancomycin for cefamandole. Two Bellingham isolates from this patient's hospital environment also failed to grow when tested on the cefamandole medium. We tested 106 additional L. pneumophila serogroup 1 isolates that belonged to nine different monoclonal antibody subtypes and demonstrated that susceptibility to cefamandole was rare (10%) and limited to the Bellingham subtype. The diagnosis of Legionnaires' disease may be missed unless the culture protocol includes both a nonselective medium and a selective medium that does not contain cefamandole. In vitro susceptibility to cefamandole also provided an epidemiologic marker that linked a water source for a patient to nosocomial Legionnaires' disease.
Serological markers of posttransfusion hepatitis C viral infection
Authors: Vallari D S , Jett B W , Alter H J , Mimms L T , Holzman R , Shih J W
Published: January 2020

Serological markers for hepatitis C virus (HCV) infection were measured in serial samples from 14 posttransfusion chronic non-A, non-B hepatitis patients by a semiquantitative dot blot immunoassay. The assay detected antibodies to HCV by use of recombinant proteins that represent putative HCV capsid (core), nonstructural protein 3 (NS3) (33c), and NS4 (c100) epitopes. Seroconversion to anti-HCV antibodies (anti-HCV) was detected in all patients. The average time to active antibody production detected by any of the recombinant proteins was 13.8 (range, 3.6 to 22.0) weeks posttransfusion or 4.6 (range, -4.5 to 13.4) weeks after the first biochemical marker of illness. Anti-HCV were detected earliest by the core antigen in most cases; however, the patterns of anti-HCV responses varied significantly among individuals. Overall, the addition of the core and NS3 antigens to the assay enabled the detection of the antibody response 4 to 5 weeks earlier than did the addition of the c100 antigen, the sole antigen used in current screening tests in the United States. Passively transferred antibodies were detected by at least one antigen in early posttransfusion samples from 12 patients and decayed below detectable levels for all antigens in only 2 patients. Antibodies to all three gene products were evident in the last sample from all five patients monitored for greater than 3 years from transfusion indicating the persistence of antibodies in patients with chronic illness. Our data show that the period following the onset of hepatitis during which anti-HCV are not detected by current screening assays can be greatly shortened by the detection of anti-HCV responses by a combination of core, NS3, and c100 antigens.
Performance and reliability of the Enzygnost measles enzyme-linked immuno-sorbent assay for detection of measles virus-specific immunoglobulin M antibody during a large measles epidemic
Authors: Ozanne G , d'Halewyn M A
Published: January 2020

Evaluation of the Enzygnost Measles Enzyme-Linked Immuno-Sorbent Assay kit (Behring) performance to detect specific immunoglobulin M (IgM) was carried out with 3,297 single serum samples and 898 paired serum samples collected during a measles epidemic (10,184 reported cases) in Quebec, Canada. Anti-measles IgM and IgG were detected by using the Enzygnost kit with the appropriate conjugates. Complement-fixing (CF) antibody (Ab) titers were assessed by the laboratory branch complement fixation micromethod. The Centers for Disease Control's clinical measles case definition was used. A modification of the manufacturer's optical density interpretation algorithm was introduced to allow for equivocal results, in addition to positive and negative ones. These three categories differed as to their association with a significant increase in CF Ab titer and the time between the onset of symptoms and phlebotomy. The IgM positivity rate for complement fixation-confirmed measles cases was 96.6% for vaccinated subjects and 100% for nonvaccinated subjects. The daily percentage of IgM seropositivity that was detected for subjects who became IgM positive within 30 days increased gradually from 40 to 90% for sera taken 1 to 7 days after the onset of symptoms, and it plateaued at 100% for sera taken 16 to 30 days after the onset of symptoms. IgM seropositivity was strongly associated with IgG seroconversion, CF Ab titer increase, and clinical measles (P less than 0.0001). Reproducibility was 100% for nonreactive sera and 99.1% for reactive sera. In conclusion, the Enzygnost Measles Enzyme-Linked Immuno-Sorbent Assay kit performed adequately to confirm measles virus infection during this epidemic. A second serum sample should be tested when an early-acute-phase serum sample is IgM negative.
Detection of herpes simplex virus by a nonradiometric spin-amplified in situ hybridization assay
Authors: Forman M S , Merz C S , Charache P
Published: January 2020

An in situ hybridization kit (Diagnostic Hybrids, Inc., Athens, Ohio) was evaluated for use in the detection and identification of herpes simplex virus (HSV) from clinical specimens. For in situ hybridization, a 10-min spin amplification onto monolayers of African green monkey kidney cells (CV-1) in 24-well polystyrene dishes, 24-h culture amplification, and hybridization with an alkaline phosphatase-labeled DNA probe were used. A total of 648 specimens were tested, including 275 specimens from patients with symptomatic diseases sent specifically for HSV detection and 373 specimens from asymptomatic immunocompromised patients sent for detection of HSV shedding. Overall, the sensitivity of the hybridization assay was 97.8% (131 of 134 specimens), with 105 of 105 (100%) specimens from symptomatic patients and 26 of 29 (89.9%) specimens from asymptomatic patients being detected. The three specimens that were false negative by in situ hybridization had low virus titers, as determined by tissue culture. The specificity was 99.6% (512 of 514 specimens). The rapid, accurate results suggest that the in situ hybridization kit may be used as an alternative to conventional tissue culture for the detection of HSV.
Revision of standards for adjusting the cation content of Mueller-Hinton broth for testing susceptibility of Pseudomonas aeruginosa to aminoglycosides
Authors: Barry A L , Reller L B , Miller G H , Washington J A , Schoenknect F D , Peterson L R , Hare R S , Knapp C
Published: January 2020

A multilaboratory study was undertaken to reassess the amount of calcium and magnesium that should be added to Mueller-Hinton broth when testing Pseudomonas aeruginosa against amikacin, gentamicin, isepamicin, netilmicin, and tobramycin. To achieve parity with agar dilution tests, cation-adjusted broth should contain 20 to 25 mg of calcium and 10 to 12.5 mg of magnesium per liter rather than the 50- and 25-mg/liter supplements recommended previously. For quality control of tests with contemporary media, MIC control limits should be adjusted by lowering the current MIC limits by at least 1 doubling-dilution interval.
Bacteriophage lytic patterns for identification of salmonellae, shigellae, Escherichia coli, Citrobacter freundii, and Enterobacter cloacae
Authors: He X Q , Pan R N
Published: January 2020

A series of bacteriophages specific for Escherichia coli (E-1, E-2, E-3, and E-4), Citrobacter freundii (phi I, phi II, and phi III), Enterobacter cloacae (Ent), and Shigella spp. (Sh) have been isolated from hospital sewage. These bacteriophages, in combination with Felix Salmonella phage O-I, were used as a diagnostic phage typing set which included seven phage preparations: O-I, C (phi I and phi III), Sh, E (E-1 and E-2), CE (phi II and E-3), E-4, and Ent. After 20,280 cultures of 27 species and 9 biogroups of 15 genera of the family Enterobacteriaceae and 276 cultures of 8 species of 6 genera outside the Enterobacteriaceae were tested, it was shown that most strains of salmonellae, E. coli, C. freundii, and E. cloacae can be identified accurately. The sensitivities of identification were 83.6% for E. cloacae, 88.8% for C. freundii, 90.3% for E. coli, and 95.76% for salmonellae. The specificities were 99.78% for salmonellae, 99.84% for E. cloacae, 99.89% for E. coli, and 99.97% for C. freundii. The results of bacteriophage lytic patterns were highly correlated with Shigella serotypes. Therefore, such a phage typing set may be used routinely in public hygiene and clinical laboratories.
Diagnostic accuracy of a Mycobacterium phlei-absorbed serum enzyme-linked immunosorbent assay for diagnosis of bovine paratuberculosis in dairy cows
Authors: Bech-Nielsen S , Jorgensen J B , Ahrens P , Feld N C
Published: January 2020

The purpose of this study was to describe the responses of sera from five groups of cattle to an enzyme-linked immunosorbent assay (ELISA) for paratuberculosis by using serum absorbed with Mycobacterium phlei at a single working dilution. The infection status of the cattle was determined by fecal culture. Cattle with different levels of exposure (high versus low prevalence and test negative) and disease manifestation (clinically suspect infection versus subclinical infection) were examined, as follows: (i) two paratuberculosis-negative herds; (ii) a fecal culture-confirmed, clinically suspect cases of paratuberculosis; (iii) cows from a paratuberculosis-infected herd with a high infection rate, as determined by fecal culture, but with no clinical cases at the time of sampling; (iv) cows from three paratuberculosis-infected herds known to have paratuberculosis diagnosed on the farm (low infection rate determined by fecal culture); and (v) one fecal culture-negative herd with known serologically positive cattle. Results generally showed a decreased ELISA response when absorbed rather than nonabsorbed serum from each animal was used. The results of the fecal culture confirmed clinically suspect cases, which were analyzed in relation to the amount of colonies isolated from the animals on fecal culture (0, +, ++,+++ , ++++, and above). There was a significant increase in the ELISA response for animals with heavy Mycobacterium paratuberculosis shedding ( ++++ or above), when both unabsorbed and absorbed sera were used, compared with the response in animals that were fecal culture negative or that shed M. paratuberculosis at lower levels (less than +) (P less than 0.05). The effects on sensitivity and specificity by using different cutoff points for the five groups of cattle with different levels of exposure is described, since sera were not discretely segregated into distinct groups of positive and negative samples. The specificity of the ELISA in the two fecal culture-negative herds was 100% at an ELISA cutoff of an optical density (OD) of 0.1 and above for absorbed serum. For unabsorbed serum the specificity was 62.9% at a similar cutoff value. Similarly, the specificity of the fecal culture-negative, serologically positive herd increased from 37.5 to 72.2 at an ELISA cutoff value of 0.1 to 0.2 (OD) by using absorbed versus unabsorbed serum from 75.0 to 94.4 at an ELISA cutoff value of 0.2 to 0.3 (OD).
Siderophore production and DNA hybridization groups of Aeromonas spp
Authors: Zywno S R , Arceneaux J E , Altwegg M , Byers B R
Published: January 2020

A correlation between the genospecies (DNA-DNA hybridization group) and the type of siderophore produced by 118 isolates of the genus Aeromonas was established. Organisms in hybridization groups 1 through 5 (including 5A, 5B, and 5AB) and group 12 predominantly produced the siderophore amonabactin, while an enterobactinlike siderophore was prevalent in groups 8/10 and 9. The siderophore produced by strains in group 6 may be an as-yet-unidentified nonphenolate, nonhydroxamate compound, and group 7 isolates synthesized no siderophores. Determination of the indigeneous siderophore (or the absence of one) produced by an isolate of the genus Aeromonas may assist in identification of the organism's genetic species and may suggest the presence of certain virulence properties.
Clonal analysis of the Actinobacillus pleuropneumoniae population in a geographically restricted area by multilocus enzyme electrophoresis
Authors: Møller K , Nielsen R , Andersen L V , Kilian M
Published: January 2020

The genetic diversity among 250 isolates of Actinobacillus pleuropneumoniae from lungs of pigs with pleuropneumonia and from tonsils of apparently healthy pigs at slaughter was estimated by multilocus enzyme electrophoresis. The Danish strains were derived from both specific-pathogen-free and conventional herds. Sixty-six percent of the isolates belonged to three electrophoretic types (ETs) of a total of 37 ETs detected. While five biotype 2 isolates constituted a separate ET closely related to biotype 1 isolates, the type strain of the species (Shope 4074) belonged to its own ET, with a genetic distance of 0.30 from its nearest neighbor. Isolates of serotypes traditionally considered to have less pathogenic potential (serotypes 6, 10, and 12) from herds with acute outbreaks of pleuropneumonia belonged to the same ETs as isolates from apparently healthy pigs, suggesting that factors such as cross immunity and management may lead to divergent clinical results. Isolates from four herds harboring more than one serotype showed distinct profiles between the serotypes, indicating no or only limited chromosomal recombination among clones. Isolates from tonsils belonged to the same ET as isolates from lungs. The same ET was isolated from widely different parts of the world. Evidence from this study indicates that multilocus enzyme electrophoresis may be a valuable tool for the epidemiological analysis of A. pleuropneumoniae.
Rapid presumptive identification and further characterization of Bacteroides forsythus
Authors: Braham P H , Moncla B J
Published: January 2020

Bacteroides forsythus is a fastidious anaerobic gram-negative organism associated with various forms of periodontal disease. It is dependent on N-acetylmuramic acid for growth. A method for rapid presumptive identification of human-derived strains of B. forsythus is presented, based on the following eight criteria: (i) positive activity for alpha-glucosidase, (ii) positive activity for beta-glucosidase, (iii) positive activity for sialidase, (iv) positive activity for trypsinlike enzyme, (v) negative indole production, (vi) requirement for N-acetylmuramic acid, (vii) colonial morphology, and (viii) gram stain morphology from blood agar medium deficient in N-acetylmuramic acid. Enzymes were assayed with rapid filter paper spot tests based on fluorogenic substrates (4-methylumbelliferone derivatives and N alpha-carbobenzoxy-L-arginine-7-amino-4-methylcoumarin hydrochloride). Gas-liquid chromatography analysis of the metabolic products of B. forsythus grown in peptone yeast extract broth supplemented with N-acetylmuramic acid and heat-inactivated horse serum revealed predominant amounts of acetate, propionate, butyrate, isovalerate, and phenyl acetate, with minor amounts of isobutyrate and succinate. The described presumptive identification scheme facilitated recognition of four strains of B. forsythus which were isolated from subgingival plaque samples from monkeys (Macaca fascicularis). With the exception of indole production, these organisms were essentially identical to the human strains of B. forsythus for all phenotypic and genotypic characteristics examined.
Recurrent Staphylococcus aureus bacteremia
Authors: Hartstein A I , Mulligan M E , Morthland V H , Kwok R Y
Published: January 2020

Sequential blood isolates from eight patients with 10 episodes of recurrent Staphylococcus aureus bacteremia were typed by restriction endonuclease analysis of plasmid DNA (REAP DNA fingerprinting) and immunoblotting. There were six early recurrences (within 2 months of stopping antimicrobial therapy) and four late recurrences. All early recurrences isolates were identical to initial isolates. These recurrences were defined as possible relapses. Three of four late recurrence isolates were different from the preceding isolates recovered from four patients. This was considered indicative of new infections. There was complete concordance between REAP DNA fingerprinting and immunoblot typing results. However, four isolates lacked plasmid DNA and could be typed only by immunoblotting. All initial isolates from different patients were different types by immunoblotting and by REAP DNA fingerprinting (except for those lacking plasmid DNA). The bacterial traits detected by these methods appear to be stable in vivo for up to 3 months. Relapsing infections were associated with the presence of intravascular foreign bodies and vancomycin therapy of the preceding episodes.
Multicenter evaluation of the novel ABN Western blot (immunoblot) system in comparison with an enzyme-linked immunosorbent assay and a different Western blot
Authors: Weber B , Hess G , Enzensberger R , Harms F , Evans C J , Hamann A , Doerr H W
Published: January 2020

A new, modular Western blot (immunoblot) system for human immunodeficiency virus (HIV) antibodies (ABN WesPage; Wellcome) was compared with enzyme immunoassays (Wellcome, Behringwerke, and Abbott) and with a U.S. Food and Drug Administration (FDA)-licensed Western blot (DuPont) in a multicenter study. A total of 649 serum samples from HIV patients at different stages of the disease, as well as from high-risk patients, from patients with conditions unrelated to AIDS, and from healthy blood donors, were used in the evaluation along with nine seroconversion panels. For evaluation of Western blot reactivity, both Centers for Disease Control (CDC) and FDA criteria were used. With the DuPont Western blot as the reference assay, the overall sensitivity and specificity of the ABN WesPage were 100 and 99.1%, respectively, when indeterminate results were not taken into account and when both tests were interpreted in accordance with CDC criteria. The DuPont Western blot detected significantly more antibodies to pol and gag gene products than the ABN WesPage. The ABN WesPage showed a higher positive rate of detection of viral envelope band gp160. When both Western blots were interpreted in accordance with CDC criteria, the ABN WesPage and the DuPont Western blot yielded 9.3 and 10.4% indeterminate results, respectively. When the DuPont Western blot was interpreted in accordance with the manufacturer's instructions (FDA criteria), 25.7% of the samples tested were regarded as indeterminate. The choice of interpretation criteria is of paramount importance for the evaluation of HIV Western blot patterns.
DNA analysis of Borrelia burgdorferi NCH-1, the first northcentral U.S. human Lyme disease isolate
Authors: Hughes C A , Kodner C B , Johnson R C
Published: January 2020

The DNA of the first northcentral United States human Lyme disease isolate, Borrellia burgdorferi NCH-1, was characterized and compared with the DNAs of nine other B. burgdorferi isolates. Strain NCH-1 was isolated in August 1989 from a human skin biopsy specimen. DNA was analyzed by pulsed-field gel electrophoresis and restriction endonuclease analysis. Contour-clamped homogeneous electric field pulsed-field gel electrophoresis of in situ-lysed cells was performed to compare the plasmid profiles of the various isolates. The plasmid profile of isolate NCH-1, which included five plasmids of approximately 69, 42, 38, 32, and 23 kb, could be distinguished from those of the other isolates examined. The DNA profile of NCH-1 was most similar to those of strain 297 (human cerebrospinal fluid isolate, Connecticut) and strain PAL (human erythema migrans isolate, New York) and most dissimilar from those of strain P/Gau (human erythema migrans isolate, Germany) and strain IPF (Ixodes persulcatus tick isolate, Japan). These results indicate that genetic diversity exists among B. burgdorferi strains isolated from different geographical areas.
Latex agglutination testing directly from throat swabs for rapid detection of beta-hemolytic streptococci from Lancefield serogroup C
Authors: Hayden G F , Turner J C , Kiselica D , Dunn M , Hendley J O
Published: January 2020

A latex agglutination method for the rapid detection of beta-hemolytic streptococci from Lancefield serogroup C in throat swabs from 403 university students with symptomatic pharyngitis was evaluated. Compared with culture, the rapid test was poorly sensitive (34.4%) but very specific (98.4%) in detecting group C beta-hemolytic streptococci. The sensitivity of the rapid test improved with an increasing quantity of growth on culture.
Association of an important Neisseria species, Neisseria elongata subsp. nitroreducens, with bacteremia, endocarditis, and osteomyelitis
Authors: Wong J D , Janda J M
Published: January 2020

We retrospectively analyzed epidemiologic information associated with 22 cultures of Neisseria elongata subsp. nitroreducens (formerly CDC group M-6) submitted to the Microbial Diseases Laboratory, California Department of Health Services, Berkeley, over a 16-year period. The most common illnesses noted with this bacterium were endocarditis, bacteremia, and osteomyelitis. Risk factors associated with N. elongata subsp. nitroreducens infection included dental manipulations and/or a previous history of endocarditis, valve damage, or rheumatic heart disease.
Comparison of six commercial human T-cell lymphotropic virus type I (HTLV-I) enzyme immunoassay kits for detection of antibody to HTLV-I and -II
Authors: Cossen C , Hagens S , Fukuchi R , Forghani B , Gallo D , Ascher M
Published: January 2020

Three licensed and three research human T-cell lymphotropic virus type I (HTLV-I) enzyme immunoassay (EIA) kits were evaluated with a panel of 213 serum or plasma samples which were previously tested by the indirect immunofluorescence method for HTLV-I and -II antibodies. The three research kits were found to be more sensitive and specific than the three licensed kits.
Phenotypic profile and functional characteristics of human gamma and delta T cells during acute toxoplasmosis
Authors: De Paoli P , Basaglia G , Gennari D , Crovatto M , Modolo M L , Santini G
Published: January 2020

Gamma and delta (gamma delta) T-cell receptor lymphocytes are increased during acute toxoplasmosis. These cells are BB3+ CD45RO+ CD8-. Purified gamma delta T cells failed to proliferate in response to Toxoplasma gondii antigen (stimulation index, 1.4 +/- 0.6) but were responsive to phytohemagglutinin stimulation (stimulation index, 20.8 +/- 1.9). Natural-killer-like cytotoxicity was strongly acquired only after in vitro culture of purified gamma delta T cells with recombinant interleukin 2 (40% +/- 7% specific lysis). Our data show that gamma delta T-cell receptor T cells with a peculiar phenotype are increased during human acute T. gondii infection.
Use of polymerase chain reaction-amplified Helicobacter pylori urease structural genes for differentiation of isolates
Authors: Foxall P A , Hu L T , Mobley H L
Published: January 2020

Helicobacter pylori has been demonstrated as an etiologic agent of human gastritis and peptic ulcer formation. However, there is no straightforward basis to distinguish different isolates. We used the polymerase chain reaction (PCR) to amplify the urease structural subunit genes, ureA and ureB, which, when digested with appropriate restriction endonucleases, allow the differentiation of patterns on agarose gels. PCR amplification was possible with DNA rapidly extracted from H. pylori by alkaline lysis and phenol-chloroform. The 2.4-kb PCR products amplified from 22 clinical isolates and subjected to HaeII restriction endonuclease digestion produced 10 distinct patterns on agarose gels, with two patterns being shared between five and six strains. PCR amplification of the urease genes may enable the differentiation of closely related H. pylori strains by restriction digest analysis of PCR-amplified ureA and ureB genes.
Assessment of enzyme detection tests useful in identification of campylobacteria
Authors: On S L , Holmes B
Published: January 2020

Twenty-one type or other reference strains, each representing a different Campylobacter, Helicobacter, or Arcobacter taxon, and a reference strain of Staphylococcus aureus were used to assess the reproducibility of nine enzyme detection tests used in the identification of campylobacters. For five of the tests (alkaline phosphatase, DNase, and H2S production, indoxyl acetate hydrolysis, and nitrate reduction), more than one procedure was employed to determine the most suitable method. Alkaline phosphatase test results were better defined and more reproducible if read after 1 h of incubation. Detection of DNase was fully reproducible with each method (except with Helicobacter pylori), but reactions were generally weaker than those of other DNase-producing organisms. Both procedures for determining H2S production were irreproducible for the same strains. The reproducibility of indoxyl acetate hydrolysis was improved by using disks impregnated with 25 microliters of substrate. Reduction of nitrate was best determined by Cook's plate method. Results for the other tests examined (catalase, oxidase, and urease production and hippurate hydrolysis) were both pertinent and fully reproducible for all strains.
Effects of anticoagulants and storage of blood samples on efficacy of the polymerase chain reaction assay for hepatitis C virus
Authors: Wang J T , Wang T H , Sheu J C , Lin S M , Lin J T , Chen D S
Published: January 2020

Blood samples from 11 patients with posttransfusion hepatitis C virus infection were collected. Each sample was divided into three fractions to obtain sera, sodium-citrated plasma, and heparinized plasma and then tested for HCV RNA by a nested polymerase chain reaction (PCR). Of them, eight sodium-citrated plasma samples, seven serum samples, and no heparinized plasma samples were PCR positive. Eight PCR-positive sodium-citrated plasma samples were exposed to different physical conditions and semiquantified for HCV RNA after serial dilutions. Samples stored at -70 degrees C showed the best preservation of HCV RNA, and storage at the other conditions resulted in only minimal loss of the PCR signal. Therefore, serum or sodium-citrated plasma specimens are satisfactory for detecting HCV RNA by PCR, but heparinized blood specimens are not.
Highly sensitive enhanced chemiluminescence immunodetection method for herpes simplex virus type 2 Western immunoblot
Authors: Dalessio J , Ashley R
Published: January 2020

Three Western blot (immunoblot) methods for detecting antibodies to herpes simplex virus type 2 were compared: (i) nitrocellulose blots with 4-chloro-1-naphthol immunostaining (4CN-WB); (ii) polyvinylidene difluoride (PVDF) blots with 3,3',5,5'-tetramethylbenzidene immunostaining (TMB-WB); and (iii) PVDF blots with enhanced chemiluminescence (ECL-WB). TMB-WB was 10-fold more sensitive than 4CN-WB, while ECL-WB was as much as 500-fold more sensitive.
Nonmycetomic Actinomadura madurae infection in a patient with AIDS
Authors: McNeil M M , Brown J M , Scalise G , Piersimoni C
Published: January 2020

Actinomadura madurae is an aerobic actinomycete which is best known worldwide as the cause of actinomycotic mycetomas. It has not previously been reported to have caused invasive pulmonary or disseminated infection in humans. We describe an AIDS patient with opportunistic A. madurae-induced pneumonia and bacteremia. The isolate from the patient's blood was subjected to dilutional antimicrobial susceptibility tests with 12 antimicrobial agents and was found to have a wide spectrum of susceptibility. This unusual microorganism may be a cause of infections in severely immunosuppressed patients.
Comparison of immunotyping of Chlamydia trachomatis by indirect fluorescent-antibody staining and radioimmunoassay
Authors: van der Pol B J , Jones R B
Published: January 2020

Chlamydia trachomatis isolates from 58 patients were immunotyped by radioimmunoassay (RIA) and fluorescent-antibody staining (FA) by using monoclonal antibodies. Unambiguous typing was observed with 55 isolates by FA and 53 isolates by RIA, although 3 isolates were typed by RIA but not FA. FA required less antigen and was more sensitive in detecting mixed infections.
Rapid identification of legionellae by a colony blot assay based on a genus-specific monoclonal antibody
Authors: Steinmetz I , Rheinheimer C , Bitter-Suermann D
Published: January 2020

We recently developed a monoclonal antibody immunoglobulin G2a (2125) recognizing a genus-specific epitope on the 60-kDa heat shock protein of all Legionella species. In the current study, this antibody was used in a colony blot enzyme-linked immunosorbent assay for the rapid identification of Legionella cultures on agar plates. The whole protocol was completed in less than 2 h. All 59 Legionella species and serogroups that were tested gave a positive signal. No unspecific reactions with nonlegionellae were observed. This test is a rapid procedure for the identification of legionellae growing on agar medium to the genus level.
Interpretive criteria for disk diffusion tests using 5-microgram cefdinir disks with rapidly growing clinical isolates
Authors: Jones R N , Erwin M E , Gooding B B
Published: January 2020

Preliminary interpretive zone diameter criteria were calculated for the 5-micrograms cefdinir disk diffusion test by using two potential MIC breakpoints (less than or equal to 0.5 and less than or equal to 1 micrograms/ml). The absolute agreement between tests ranged from 85.9 to 92.4%, and the false-susceptibility errors were principally contributed by the Enterobacter spp. (2.2% error). One proposed criterion was greater than or equal to 20-mm zone diameter (less than or equal to 1 micrograms/ml) for susceptibility and less than or equal to 16-mm zone diameter (greater than 2 micrograms/ml) for resistance to cefdinir. Clinical laboratory users of the disk diffusion method should be cautioned about the possibility of very major interpretive errors among enterobacter isolates.
Isolation of Mycobacterium thermoresistibile following augmentation mammaplasty
Authors: Wolfe J M , Moore D F
Published: January 2020

This is the first case report of a Mycobacterium thermoresistibile infection following augmentation mammaplasty and is the fourth human case report of M. thermoresistibile infection. Antimicrobial susceptibility results determined by a modified proportion method using a 3-day incubation were the same as those determined by the standard 3-week assay.
Detection of human cytomegalovirus in clinical specimens by centrifugation culture with a nonhuman cell line
Authors: Gleaves C A , Hursh D A , Meyers J D
Published: January 2020

The sensitivities of MRC-5 and mink lung (ML) cells in centrifugation culture were compared simultaneously for the detection of cytomegalovirus (CMV) IE antigen (immediate-early antigen) from clinical specimens. Of 413 samples assayed, 51 (12%) were positive for CMV by both centrifugation and standard cell culture. At 20 h postinoculation (p.i.), 46 of 51 (90.2%) CMV-positive specimens were detected in ML cells. At 40 h p.i., 50 of 51 (98.0%) CMV-positive specimens were detected in ML cells, compared with 48 of 51 (94.0%) in MRC-5 cells. There was no significant difference in the detection of CMV in either cell line by centrifugation culture. However, in 19 of 23 positive samples that had countable foci at 20 h p.i., there was a 25% increase in the number of positive foci observed for ML cells compared with MRC-5 cells. Less toxicity was also noted for ML cells than for MRC cells, particularly in viral blood specimens. These data suggest that ML cells are comparable to MRC-5 cells for the rapid detection of CMV by centrifugation culture.
Determination of growth value thresholds for BACTEC PLUS aerobic blood culture vials
Authors: McGowan J E , Metchock B G
Published: January 2020

Growth value thresholds used to identify positive blood culture vials can be defined by users for each BACTEC NR-660 bacteremia detection instrument. Growth values were compared with the recovery of organisms from vials flagged as positive during the testing of 3.056 high-volume vials containing aerobic (BACTEC PLUS 26) medium over a 2-month period. Results showed that optimal threshold values for our use of these vials varied from those recommended by the manufacturer; if the thresholds defined from these data had been used during the study period, total vials flagged as positive from which no organisms were recovered (false alarms) would have been reduced from 181 (5.9/100 vials tested) to 71 (2.3/100 vials tested), with a minimal decrease in the identification of vials containing usual or occasional pathogens (hits). Adjustments of growth value thresholds by the individual user can make the use of BACTEC instruments more efficient by decreasing further processing of vials from which no organisms are recovered.
Reactivity patterns and infection status of serum samples with indeterminate Western immunoblot tests for antibody to human immunodeficiency virus type 1
Authors: Ramirez E , Uribe P , Escanilla D , Sanchez G , Espejo R T
Published: January 2020

Serum samples with indeterminate Western blot (WB) tests from 61 individuals whose sera were positive by enzyme-linked immunosorbent assay (ELISA) were studied in order to characterize their putative reactions with the human immunodeficiency virus (HIV) proteins and to resolve the HIV infection status of these individuals. The reaction observed by WB could not be confirmed either by radioimmunoprecipitation assay and subsequent electrophoresis (RIPA) or by use of LiaTek (Organon Teknika, Turnbout, The Netherlands) in 28% of the samples. Of the 86 samples that were indeterminate by WB, 66 reacted with p24 by WB; this reaction was confirmed by RIPA in only 21 (32%) and by LiaTek in 49 (74%) of the 66 samples. On the other hand, none of the indeterminate samples that reacted with HIV envelope proteins by WB did so by LiaTek, while 50% precipitated at least some of these proteins in the RIPA. The sensitivities of the three methods for detecting the antibody reaction with the different HIV proteins, which were studied with serial dilutions of positive serum samples, were similar. Thus, a lower sensitivity of RIPA or LiaTek does not seem to be the cause for the lack of reaction of the WB-indeterminate samples by these two methods. Sequential samples from individuals whose serum samples reacted by the three methods gave reproducible results, but all showed low antibody titers. Peripheral blood mononuclear cells obtained from three of the four individuals with sequential samples that reacted with HIV env proteins by WB and RIPA were negative for HIV provirus DNA after amplification by the polymerase chain reaction.
Detection of rubella virus-specific immunoglobulin G (IgG), IgM, and IgA antibodies by immunoblot assays
Authors: Zhang T , Mauracher C A , Mitchell L A , Tingle A J
Published: January 2020

Immunoblot (IB) assays were developed for detection of rubella virus (RV)-specific immunoglobulin G (IgG), IgM, and IgA antibodies in human serum following natural infection or immunization. IB assays performed under nonreducing conditions were compared with those performed under reducing conditions and with immunoprecipitation assays. Significant loss of antigenicity (greater than 90%) of RV E1 and E2 proteins was observed when IB assays were performed in the presence of 2-mercaptoethanol as compared with assays under nonreducing conditions. In contrast, the antigenicity of RV capsid protein was not influenced by reducing agents. Sensitivity of IB for RV-specific IgG antibodies was determined to be 0.01 IU/ml under nonreducing conditions. In the determination of RV-specific IgM and IgA antibodies by IB, pretreatment of serum with protein G to remove competing high-affinity RV-specific IgG or rheumatoid factor significantly improved assay sensitivity. IB assays were observed to be superior to immunoprecipitation assays in their ability to better define the specificities of RV-specific antibodies and to detect antibodies of all immunoglobulin classes. However, the conformational sensitivity of RV protein antigenicity should be an important consideration in the interpretation of RV-specific antibodies by IB assays.
Infection of peripheral blood mononuclear cells and cell lines by cell-free human T-cell lymphoma/leukemia virus type I
Authors: Fan N , Gavalchin J , Paul B , Wells K H , Lane M J , Poiesz B J
Published: January 2020

Previous studies of in vitro infection by human T-cell lymphoma/leukemia virus type I (HTLV-I) have required cocultivation of target cells with HTLV-I cell lines or vesicular stomatitis virus pseudotypes containing HTLV-I envelope proteins. We report here the development of a cell-free infection assay for HTLV-I. Target cells were incubated with purified, DNase-treated HTLV-I virions for 4 h at 37 degrees C. Target cell DNA was then analyzed for the presence of newly synthesized HTLV-I proviral DNA by the highly sensitive polymerase chain reaction. Using this assay system, we have been able to consistently detect in vitro infection of a variety of cellular targets by different HTLV-I isolates. Optimal infection required the presence of 10 micrograms of DEAE-dextran per ml. The assay was dose dependent with respect to virus input. In general, the amount of proviral DNA detected correlated with the level of HTLV-I receptors present on the surface of the target cells, as measured by fluorochrome-labelled HTLV-I binding. Finally, the specificity of the assay was confirmed by demonstrating that the cell line, L1q, a somatic cell hybrid containing human chromosome 17q, to which the gene for the HTLV-I receptor has been mapped, was susceptible to infection by HTLV-I, while the parental mouse cell line from which it was derived, LMTK-, which lacks human chromosome 17q, was not.
Detection of hepatitis C virus sequences in sera with controversial serology by nested polymerase chain reaction
Authors: Lazizi Y , Elfassi E , Pillot J
Published: January 2020

The specificity of first-generation enzyme-linked immunosorbent assays (ELIAs) for antibody detection in individuals with hepatitis C virus (HCV) infection has been questioned in some pathological situations. We observed a surprisingly high prevalence of anti-HCV antibodies in alcoholic patients, and thus, false-positive reactions in anti-HCV tests were strongly suspected. The introduction of new epitopes, particularly a core protein, C22 (second-generation tests), seems to increase the sensitivity of anti-HCV detection. In order to study the specificity of the second-generation tests, 60 serum samples from alcoholic patients found to be positive by the first-generation anti-HCV ELISA (Ortho) were reexamined by a second-generation anti-HCV enzyme immunoassay (Abbott) and a recombinant immunoblot assay (RIBA II; Chiron). Fifteen serum samples gave contradictory results when they were tested by the two assays. We performed nested polymerase chain reactions (PCRs) to confirm that the discrepancies that we observed could be due to the presence of low levels of anti-HCV antibodies, which were detected by a more sensitive test, or to unspecific positive reactions. Nested PCR revealed the presence of HCV RNA sequences in all anti-HCV-positive sera or sera that were weakly positive by ELISA. Anti-HCV positive by RIBA II was always correlated with the presence of viral RNA in serum, but HCV RNA was detected in RIBA II-negative sera. These results indicate that the specificity of the second-generation tests is an important improvement but that an HCV infection can still persist without detectable antibodies. PCR remains the reference assay to clear up controversial serology results and to detect HCV infection in patients with no anti-HCV-detectable immune response.