bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparison of latex agglutination test with enzyme-linked immunosorbent assay for detection of antibody to varicella-zoster virus
Authors: Landry M L , Ferguson D
Published: January 2020

A new latex agglutination (LA) test (VZVscan; Becton Dickinson, Cockeysville, Md.) was compared with an enzyme-linked immunosorbent assay (ELISA) (Varicella STAT; Whittaker Bioproducts, Walkersville, Md.) for detection of varicella-zoster virus (VZV) antibody. Of 165 samples tested, 126 (76%) were positive by LA, 123 (73%) were positive by ELISA, and 35 (21%) were negative by both methods. Six samples (4%) were LA positive and ELISA negative or equivocal; three samples (2%) were ELISA positive and LA negative. However, LA failed to detect seroconversions in four adults with varicella in samples obtained 14 to 17 days after the onset of rash. These same samples had ELISA values in the high-positive range. In addition, the recommended LA screening dilution (1:2) is not supported by published data and should be changed.
VP4 serotype of the Gottfried strain of porcine rotavirus
Authors: Li B , Gorziglia M
Published: January 2020

The antigenic relationships of the VP4 serotype of porcine rotavirus Gottfried strain with other rotaviruses were determined by using antiserum to Gottfried VP4-baculovirus recombinant. This antiserum failed to react significantly with virus of serotypes P1A, P1B, P3, and OSU; however, it reacted with P2 strains. In the reciprocal assay, antiserum to VP4 of an asymptomatic strain (P2) failed to neutralize the Gottfried strain virus to a significant level. It thus appears that the Gottfried strain should be considered a subtype of P2.
Factors affecting detection of Brucella melitensis by BACTEC NR730, a nonradiometric system for hemocultures
Authors: Gamazo C , Vitas A I , López-Goñi I , Díaz R , Moriyón I
Published: January 2020

The detection of Brucella bacteremia by subculture does not always correlate with a positive signal in the BACTEC NR730 nonradiometric system (Becton Dickinson Diagnostic Instrument Systems, Sparks, Md.). The effect of the inoculum size, pH, sodium polyanetholesulfonate, carbon sources (i-erythritol, sodium pyruvate, monosodium glutamate, D-glucose, and L-alanine), and urea in the release of CO2 was evaluated by using the reference strain Brucella melitensis 16M. In standard NR6 vials with or without blood, inocula 5 to 10 times larger (at least 265 CFU per vial) than those usually found in the blood of patients with brucellosis were necessary to produce a positive growth value (GV) in 4 days or less, and similar results were obtained with vials supplemented with the substrates listed above. GVs were consistently lower in vials with sodium polyanetholesulfonate than in vials without this agent. Vials with no blood inoculated with 265 CFU per vial showed turbidity 1 day before GVs became positive, proving that the major limiting detection factor was the low level of release of CO2 and not an inadequate growth medium. In NR6 vials buffered to pH 6.2, GVs became positive faster and were higher than those in standard vials. NR6 vials at pH 6.2 with 0.3% sodium pyruvate yielded a positive GV in the first day of bacterial turbidity.
Evaluation of Clearview and Magic Lite tests, polymerase chain reaction, and cell culture for detection of Chlamydia trachomatis in urogenital specimens
Authors: Kluytmans J A , Goessens W H , Mouton J W , van Rijsoort-Vos J H , Niesters H G , Quint W G , Habbema L , Stolz E , Wagenvoort J H
Published: January 2020

The Clearview Chlamydia test (CV; Unipath Ltd., Bedford, United Kingdom), the Magic Lite Chlamydia test (ML; CIBA Corning, Medfield, Mass.), a polymerase chain reaction (PCR), and cell culture (CC) were evaluated for detection of Chlamydia trachomatis in urogenital specimens. Specimens were collected from 283 men and 724 women visiting the outpatient clinic for Sexually Transmitted Diseases at the University Hospital Rotterdam, Rotterdam, The Netherlands. ML, PCR, and CC were all performed on the same sample to prevent swab-to-swab variability. CV was performed on a separate sample. Analysis of discordant results was performed by application of the following confirmatory assays: first, PCR on the CC, second, ML was repeated, and third, PCR was repeated by using a different DNA extraction protocol. If more than one test was positive, the sample was considered true positive. If only one test was positive, which was confirmed by the confirmatory assay, the sample was also considered true positive. By using these interpretations, the following results were obtained. The sensitivity and specificity of CV for samples from men were 60.4 and 86.3%, respectively. For samples from women, these values were 62.3 and 99.7%, respectively. The low specificity for samples from men was caused by unidentified substances in the swab that was used. The use of CV on samples from men is not recommended by the manufacturer. For samples from women, the specificity of CV was high, but the low sensitivity of CV limits its use for diagnostic purposes. The sensitivities of ML were low for samples from both men and women (68.8% and 50.9% respectively), while specificities were excellent for samples from both groups (100 and 99.9%, respectively). The low sensitivity of ML limits its diagnostic value. The PCR technique was highly specific for samples from both men (99.6%) and women (99.9%). The sensitivity of PCR, however, was unexpectedly low for samples from both groups (men, 87.5%; women, 79.2%), most likely because of the sample treatment method used. The sensitivity and specificity values of CC for samples from men were 95.8 and 100%, respectively. For samples from women, these values were 100 and 99.9%, respectively. In the present study, CC was the most reliable technique for the detection of C. trachomatis.
Discrimination of epidemic and sporadic isolates of Arcobacter butzleri by polymerase chain reaction-mediated DNA fingerprinting
Authors: Vandamme P , Giesendorf B A , van Belkum A , Pierard D , Lauwers S , Kersters K , Butzler J P , Goossens H , Quint W G
Published: January 2020

DNA polymorphisms of Arcobacter butzleri outbreak-related strains and Arcobacter reference strains were determined by use of the polymerase chain reaction with primers aimed at repetitive sequences. The epidemiological relationship among 14 outbreak-related strains was substantiated, as they showed virtually no genomic variations. Their DNA amplification patterns were, however, clearly different from those of all Arcobacter reference strains studied; each reference strain was characterized by a unique DNA fingerprint.
Human papillomavirus DNA in urine samples compared with that in simultaneously collected urethra and cervix samples
Authors: Forslund O , Hansson B G , Rymark P , Bjerre B
Published: January 2020

A polymerase chain reaction was used to evaluate the occurrence of human papillomavirus (HPV) DNA in urine samples compared with that in urethra and cervix samples simultaneously collected with brushes. Of 138 presumably healthy military conscripts, 12 (8%) had HPV DNA-positive urethra samples and 8 (5%) had HPV DNA-positive urine samples. Both the urine and urethra cell samples of five men were positive, with identical types found in the paired specimens. Seven had HPV DNA-positive urethra samples only, and three had HPV DNA-positive urine samples only. Five of 7 urethra samples from males and 11 of 12 urethra samples from females, who were among patients consulting a clinic for adolescents, were positive for HPV DNA. Among those patients whose urethras were positive for HPV DNA, the corresponding urine samples of 3 of the 5 men and all the 11 women were also positive, with one or two HPV types being in common within the paired samples. Among female patients referred to a colposcopy clinic, 49% (241 of 489) of the cervical cell samples and 38% (187 of 489) of the urine specimens were found to be HPV DNA positive. Of the patients whose cervixes were positive for HPV DNA, 65% (158 of 241) of the simultaneously collected urine samples were also positive for HPV DNA. On the other hand, 84% (158 of 187) of the patients with HPV DNA in their urine also had HPV DNA in their cervical samples. Although not all individuals with genital HPV infections could be identified as HPV positive by analysis of urine samples, at least in epidemiological surveys in which invasive samples are difficult to obtain, such as from children, analysis of urine could be an alternative means of identifying HPV DNA.
Serological response to Chlamydia pneumoniae in adults with coronary arterial fatty streaks and fibrolipid plaques
Authors: Puolakkainen M , Kuo C C , Shor A , Wang S P , Grayston J T , Campbell L A
Published: January 2020

The antigen-specific serological response to Chlamydia pneumoniae was studied in 45 adults with coronary artery atherosclerosis and compared with that in 40 adults with acute respiratory infection. C. pneumoniae antigen and DNA were detected in lesions more frequently in patients with low immunoglobulin G titers against C. pneumoniae than in those with high immunoglobulin G titers. Reactivities with the 42-kDa (46%) and 52-kDa (31%) proteins were observed more frequently in sera from seropositive individuals with atherosclerosis than in sera from patients with acute respiratory infection. Antibodies against the C. pneumoniae-specific 42- and/or 52-kDa protein may be a marker for chronic C. pneumoniae infection.
Wound isolate of Salmonella typhimurium that became chlorate resistant after exposure to Dakin's solution: concomitant loss of hydrogen sulfide production, gas production, and nitrate reduction
Authors: Lannigan R , Hussain Z
Published: January 2020

A strain of Salmonella typhimurium isolated from a decubitus ulcer that was being treated topically with half-strength Dakin's solution became H2S negative, nitrate negative, and unable to produce gas from glucose. Experimental data suggested that these effects were associated with the development of chlorate resistance. Thirty-five other strains of Salmonella spp. that were made chlorate resistant also became negative for these three tests.
Differentiation of distribution systems, source water, and clinical coliforms by DNA analysis
Authors: Edberg S C , Patterson J E , Smith D B
Published: January 2020

During a 2-week period, Enterobacter cloacae was isolated from throughout the water distribution system in New Haven County, Connecticut. There was no forewarning of this event and no apparent reasons for it. Several epidemiologic and public health questions required rapid answers. Were these E. cloacae isolates the result of treatment failure and breakthrough or was regrowth occurring within the system? Did the E. cloacae isolates represent a health threat and were they causing infection? Pulsed-field gel electrophoresis utilizing whole-cell DNA digestion with restriction endonuclease SpeI permitted the rapid generation of specific information to answer these questions. Gel bands were stained with ethidium bromide and photographed with UV illumination. Homogeneity among isolates was confirmed by repeat digestion with XbaI. From each of the water distribution isolates, a single pattern of restriction endonuclease fragments was generated, indicating that only one clone of E. cloacae was in the distribution system. There was no homogeneity between source and distribution water E. cloacae isolates. Moreover, E. cloacae clinical isolates from patients from New Haven area hospitals showed no identity with E. cloacae isolated from the distribution system. Therefore, pulsed-field gel electrophoresis DNA analysis demonstrated that the E. cloacae from the distribution system was the result of a regrowth bloom within the system and not the result of treatment failure and that this clone was not causing a public health risk.
Differentiation of Salmonella enteritidis phage type 8 strains: evaluation of three additional phage typing systems, plasmid profiles, antibiotic susceptibility patterns, and biotyping
Authors: Stubbs A D , Hickman-Brenner F W , Cameron D N , Farmer J J
Published: January 2020

Three additional phage typing systems for Salmonella enteritidis, plasmid analysis, biochemical tests, and antimicrobial susceptibility tests, were used in an attempt to subdivide 30 phage type 8 (phage typing system used by the WHO International Center for Enteric Phage Typing, London, England) isolates. These isolates represented 18 different egg-related outbreaks (21 strains) and 9 reference strains or strains that were not egg-associated. Only 7 of the 30 strains (28%) were subdivided by one or more of the methods used; this included 3 of the 21 strains from egg-related outbreaks. Twenty-seven strains contained a 55-kb plasmid that is associated with S. enteritidis. Of 65 additional phages tested, 2 from the phage typing system obtained from the Pasteur Institute, Paris, France, were useful in differentiating the three strains that lacked the 55-kb plasmid. Although the results obtained for the 21 strains from egg-related outbreaks showed that the strains had minor phenotypic differences, the overall results suggested that the strains may represent a single clone. Studies are planned to test additional phages and other typing methods to see whether strains of phage type 8 can be further differentiated.
Danger of sputum purulence screens in culture of Legionella species
Authors: Ingram J G , Plouffe J F
Published: January 2020

Previous studies have shown the benefit of sputum purulence screens in routine bacterial culture. We have correlated 944 sputum smear and Legionella pneumophila culture results and demonstrated that 47 to 84% of L. pneumophila-positive samples would have been discarded by using established sputum purulence screens. We recommend acceptance of all specimens submitted for Legionella culture.
Serological responses to different genotypes of hepatitis C virus in France
Authors: Alonso C , Qu D , Lamelin J P , de Sanjosé S , Vitvitski L , Li J , Berby F , Lambert V , Cortey M L , Trépo C
Published: January 2020

The relationship between hepatitis C virus (HCV) genotypes and antibody status was studied in 104 chronic non-A, non-B hepatitis patients and asymptomatic HCV-infected blood donors. On the basis of amplification of the nonstructural protein 3 (NS3) coding region by PCR and hybridization with specific probes, 55 and 42 patients were identified as being infected with type I and type II, respectively, according to the classification by H. Okamoto, K. Kurai, S. Okada, K. Yamamoto, H. Lizuka, T. Tanaka, S. Fukuda, F. Tsudaand, and S. Mishiro (Virology 188:331-341, 1992). All samples were tested for antibodies to 5.1.1, C-100, C-33, and C-22 proteins by a second-generation recombinant immunoblot assay. Among 97 patients with known HCV genotypes, 31 of 42 patients infected with type II and 24 of 55 infected with type I had antibodies against all four antigens (P < 0.01). In the type II-infected group, more patients had detectable antibodies to 5.11, C-33, and C-22 proteins than in the type I group (P < 0.05). No difference was found in the serological response to C-100 between the two groups.
A novel kit for rapid detection of Vibrio cholerae O1
Authors: Hasan J A , Huq A , Tamplin M L , Siebeling R J , Colwell R R
Published: January 2020

We report on the development and testing of a novel, rapid, colorimetric immunodiagnostic kit, Cholera SMART, for direct detection of the presence of Vibrio cholerae O1 in clinical specimens. Unlike conventional culture methods requiring several days to complete, the Cholera SMART kit can be used directly in the field by untrained or minimally skilled personnel to detect V. cholerae O1 in less than 15 min, without cumbersome laboratory equipment. A total of 120 clinical and environmental bacterial strains, including both O1 and non-O1 serotypes of V. cholerae isolated from samples collected from a variety of geographical regions, were tested, and positive reactions were observed only with V. cholerae O1. Also, results of a field trial in Bangladesh, employing Cholera SMART, showed 100% specificity and 96% sensitivity compared with conventional culture methods. Another field trial, in Mexico, showed that Cholera SMART was 100% in agreement with a recently described coagglutination test when 108 stool specimens were tested.
Stability of DNA fingerprint pattern produced with IS6110 in strains of Mycobacterium tuberculosis
Authors: Cave M D , Eisenach K D , Templeton G , Salfinger M , Mazurek G , Bates J H , Crawford J T
Published: January 2020

To assess the stability of IS6110 restriction fragment length polymorphism patterns, DNA fingerprints of 6 Mycobacterium bovis isolates from 1 patient and of 41 Mycobacterium tuberculosis isolates from 18 patients were compared. The fingerprint pattern for a given patient remained identical or nearly identical despite recovery of the isolates during intervals which ranged from 8 months to 4.5 years. Changes in drug resistance profile did not alter a strain's fingerprint pattern.
Characterization of PCR-ribotyping for Burkholderia (Pseudomonas) cepacia
Authors: Dasen S E , LiPuma J J , Kostman J R , Stull T L
Published: January 2020

Ribotyping, a method of genotyping bacterial isolates for epidemiologic study, uses rRNA as a probe to detect chromosomal restriction fragment length polymorphisms. Although ribotyping is accurate, its utility is limited by the labor and time necessary for Southern blot analysis. PCR-ribotyping uses PCR to amplify the 16S-23S intergenic spacer region of the bacterial rRNA operon. Length heterogeneity in the spacer region has previously been found to be useful as an alternative to standard ribotyping in a study of Burkholderia (Pseudomonas) cepacia. To further analyze the accuracy of PCR-ribotyping, three groups of previously characterized isolates of B. cepacia were investigated. PCR-ribotyping grouped 90 isolates recovered from seven well-defined epidemics into the correct outbreak group with a mean concordance of 93%. Both standard ribotyping and PCR-ribotyping separated 15 unrelated isolates into 14 types. In an analysis of 83 B. cepacia isolates from chronically colonized cystic fibrosis patients, the concordance of PCR-ribotyping with standard ribotyping ranged from 83 to 100%, with a mean of 98%. One isolate from a chronically colonized patient had a different type by standard ribotyping but was identical to the other isolates from this patient by PCR-ribotyping. Thus, PCR-ribotyping is a rapid and accurate method for typing B. cepacia and is less labor intensive than standard ribotyping.
Identification of human herpesvirus 6 variants A and B by amplimer hybridization with variant-specific oligonucleotides and amplification with variant-specific primers
Authors: Aubin J T , Poirel L , Robert C , Huraux J M , Agut H
Published: January 2020

Two distinct PCR-based procedures were evaluated for the detection and identification of human herpesvirus 6 (HHV-6) variants A and B in uncultured human samples. Variant-specific oligonucleotide hybridization (VSOH) is based on the amplification of two distinct regions of the HHV-6 genome, followed by hybridization of amplimers with variant-specific oligonucleotide probes. Variant-specific primer PCR (VSPP) is based on the amplification of each variant by using variant-specific primers. The study of 10 well-characterized HHV-6 strains allowed us to demonstrate the high sensitivity and specificity of both methods. With variant mixtures, however, some limitations of VSOH were evidenced and VSPP was required to obtain unambiguous results. The combination of VSOH and VSPP was applied to the direct study of 300 peripheral blood mononuclear cell samples from French subjects. HHV-6 was detected in 15 samples: 11 corresponded to variant B, 3 corresponded to variant A, and 1 corresponded to a mixture of both variants.
Determination of parallelism and nonparallelism in bioassay dilution curves
Authors: Plikaytis B D , Holder P F , Pais L B , Maslanka S E , Gheesling L L , Carlone G M
Published: January 2020

There is a lack of consensus among investigators who use a variety of immunoassay techniques (e.g., enzyme-linked immunosorbent assay [ELISA] and radioimmunoassay) regarding the protocols for describing and forming standard reference or calibration curves and interpolating serum antibody concentrations. This confounds the issue of detecting the presence or absence of parallelism between standard reference serum and serially diluted serum sample curves. These curves must be parallel to support the assumption that the antibody-binding characteristics are similar enough to allow the determination of antibody levels in the diluted serum sample. There is no universal and widely adopted strategy for assessing parallelism in bioassays, and without an assurance of parallelism, investigators are not able to calculate reliable estimates for antibody concentrations in serum samples. Furthermore, single-point (dilution) serum assays do not provide information related to parallelism and nonparallelism, and this, too, may lead to considerable error when calculating antibody concentrations. When assay methodology, technique, and precision improve to the extent that standard reference serum and serially diluted serum sample curves are fit with little error, standard analysis of variance techniques are overly sensitive to negligible departures from parallelism. We present a series of guidelines that compose a protocol for assessing parallelism between bioassay dilution curves that are applicable to data derived from ELISAs. These criteria should be applicable, with minor modifications, to most immunoassay experimental situations and, most importantly, are not dependent on the mathematical model used to form the standard reference curve. These guidelines have evolved in our laboratories over the past 4 years during the performance of thousands of ELISAs for antibodies to the capsular polysaccharides of Neisseria meningitidis groups A and C and Haemophilus influenzae type b.
Development of PCR assays to detect ampicillin resistance genes in cerebrospinal fluid samples containing Haemophilus influenzae
Authors: Tenover F C , Huang M B , Rasheed J K , Persing D H
Published: January 2020

We developed PCR primers specific for the blaTEM and blaROB ampicillin resistance genes. The specificity of the primers was confirmed by testing a series of Escherichia coli isolates containing a variety of ampicillin resistance genes and a series of ampicillin-resistant and ampicillin-susceptible Haemophilus influenzae isolates. There was a perfect correlation between ampicillin MICs, the presence of beta-lactamase (as determined by the nitrocefin test), and the results with the blaTEM and blaROB primers. Isolates of H. influenzae and Streptococcus pneumoniae obtained from 25 frozen cerebrospinal fluid (CSF) specimens were also tested. Four of 14 H. influenzae isolates were positive with the blaTEM primers; none were positive with the blaROB primers. Ampicillin MICs were determined for the H. influenzae isolates, and penicillin MICs were determined for the S. pneumoniae isolates. Only the four PCR-positive H. influenzae isolates had elevated MICs of ampicillin and were beta-lactamase positive. None of the H. influenzae isolates contained the blaROB gene, and none of the S. pneumoniae isolates produced positive reactions with either primer set. We then used universal primers directed to conserved regions of rRNA and a Haemophilus detection probe to identify which of the 25 frozen samples of CSF contained H. influenzae. Fourteen of the 25 CSF specimens were positive for H. influenzae, which correlated with the number of organisms obtained by culture of the CSF samples. Four of the CSF samples were positive with the blaTEM primer set, and these correlated with the four H. influenzae isolates that were positive when tested directly by PCR. The blaTEM assay required the use of native Taq polymerase because Amplitaq preparations were contaminated with vector DNA that contained the blaTEM-1 gene.
Transmission of Rickettsia tsutsugamushi strains among humans, wild rodents, and trombiculid mites in an area of Japan in which tsutsugamushi disease is newly endemic
Authors: Yamashita T , Kasuya S , Noda N , Nagano I , Kang J S
Published: January 2020

Thirty-two newly isolated strains of Rickettsia tsutsugamushi, 14 from patients with tsutsugamushi disease, 12 from wild rodents, and 6 from trombiculid mites parasitizing rodents in Gifu Prefecture, Japan, were examined for reactivities to 12 monoclonal antibodies by an indirect fluorescent-antibody test to classify their antigenicities. All of the isolates could be classified into one of six groups (KN-1, KN-2, KN-3, GJ-1, R158, and R161) according to their reactivities to the monoclonal antibodies. The KN-1 and GJ-1 strains that are prevalent among patients from Gifu Prefecture had the same reactivities as the Kawasaki and Kuroki strains, respectively, which have been isolated and are prevalent in the Miyazaki and Kagoshima prefectures in southwest Japan. The isolates from patients were different in serotype from those from rodents and mites (Leptotrombidium pallidum). The KN-2 and KN-3 strains were most prevalent among patients and among rodents and mites, respectively. No close similarity between KN-2 and other strains tested was observed. KN-3 is only a minor contributor to diseases in patients in Gifu Prefecture; however, it was proven that the same strain was prevalent in Niigata Prefecture in northern Japan. Thus, Gifu Prefecture is an area where southern, northern, and local strains are found. We hypothesize that humans are prone to infection with KN-2, GJ-1 (very similar to Kuroki), and KN-1 (very similar to Kawasaki), probably by infestation with Leptotrombidium scutellare. While both L. scutellare and L. pallidum parasitize wild rodents and may carry any rickettsial strain, the most virulent strain, KN-3, is predominant among wild rodents. Antigenic analysis using monoclonal antibodies to R. tsutsugamushi should be useful for epidemiological studies of infection with this organism.
The superoxide dismutase gene, a target for detection and identification of mycobacteria by PCR
Authors: Zolg J W , Philippi-Schulz S
Published: January 2020

The superoxide dismutase gene has been identified as a target in screening for the presence of mycobacteria on the genus level and differentiating relevant mycobacterial species from one another by PCR. Consensus primers deduced from known superoxide dismutase gene sequences allowed the amplification of DNAs from a variety of bacteria, fungi, and protozoa. Selected amplicons from Actinomyces viscosus, Corynebacterium diphtheriae, Corynebacterium pseudodiphtheriticum, Mycobacterium avium, M. fortuitum, M. gordonae, M. intracellulare, M. kansasii, M. scrofulaceum, M. simiae, M. tuberculosis, M. xenopi, and Nocardia asteroides were subsequently cloned and sequenced. The alignment of those sequences facilitated the selection of primers targeting conserved regions present in myobacterial species but absent in nonmycobacterial species and thus allowed the genus-specific amplification of all 28 different mycobacterial species tested. A pool of genus-specific allowed the genus-specific amplification of all 28 different mycobacterial species tested. A pool of genus-specific probes recognized 23 of the 28 mycobacterial species and did not cross-react with any of the 96 nonmycobacterial species tested. In addition, probes recognizing species-specific variable regions within the superoxide dismutase genes of M. avium, M. fortuitum, M. gordonae, M. intracellulare, M. kansasii, M. scrofulaceum, M. simiae, the M. tuberculosis complex, and M. xenopi were identified. All probes recognized only the species from which they were derived and did not cross-react with any other mycobacterial species or with any of the nonmycobacterial species tested. We conclude that the superoxide dismutase gene is a suitable target for amplifying mycobacteria by PCR on the genus level, confirming correct amplification by genus-specific probes, and differentiating relevant species from one another by a set of species-specific probes.
Value of PCR for evaluating occurrence of parasitemia in immunocompromised patients with cerebral and extracerebral toxoplasmosis
Authors: Khalifa K el-S  , Roth A , Roth B , Arasteh K N , Janitschke K
Published: January 2020

PCR was used to evaluate the occurrence of Toxoplasma gondii parasitemia by detection of the B1 gene in blood samples in two groups of immunosuppressed patients (148 subjects) suspected of having cerebral or extracerebral infection, respectively. Group I consisted of 52 patients with AIDS with suspected cerebral toxoplasmosis. The diagnosis was clinically proven in 15 cases. Parasitemia was detected by PCR in only two of these patients (13.3%), both showing evidence of disseminated infection. Group II consisted of 96 immunocompromised patients, either with AIDS or receiving iatrogenic immunosuppressive therapy. Of these patients, 65 (34 with AIDS and 31 others) showed abnormalities only in chest radiography and were first screened for the presence of Toxoplasma DNA in bronchoalveolar lavage fluid. Blood was then analyzed when the parasite was detected in the bronchoalveolar lavage fluid. The remaining 31 subjects (22 with AIDS and 9 others) were suspected of having extracerebral, pulmonary, or disseminated toxoplasmosis, and blood was studied directly in these cases. Among the nine patients with clinically diagnosed extracerebral infection in group II, the parasite was detected by PCR in the blood of five patients (55.5%), all having pulmonary toxoplasmosis. If all patients with clinical manifestations of extracerebral toxoplasmosis (from both groups) who had not received antitoxoplasma therapy when the samples were collected are considered, PCR detected parasitemia in seven of the nine cases (77.8%). The present study indicates that examination of blood by PCR may be valuable in cases of extracerebral toxoplasmosis because of the disseminated nature of the disease. Since most cases of cerebral toxoplasmosis result from the local reactivation of latent brain cysts, detection of parasitemia by PCR is useful only in cases associated with severe cerebral infection or dissemination of this disease.
Osteomyelitis caused by Neosartorya pseudofischeri
Authors: Padhye A A , Godfrey J H , Chandler F W , Peterson S W
Published: January 2020

The first case of osteomyelitis caused by Neosartorya pseudofischeri is reported. The patient, a 77-year-old male with a history of silicosis and tuberculosis, on X-ray examination revealed lytic lesions of L2 and L3 vertebrae suspicious for metastatic lesions. Histologic examination of biopsy specimens from vertebral bodies showed short, distorted, extra- and intracellular, hyaline hyphal fragments. The culture from the biopsy tissue produced numerous, evanescent asci containing eight ellipsoidal ascospores with two distinctive equatorial bands ca. 1 micron wide. When examined by a scanning electron microscope, ascospores exhibited a convex surface ornamented with raised flaps of tissue, in shape resembling triangular projections or long ridge lines. The conidial state (anamorph) was identified as Aspergillus thermomutatus on the basis of conidial columns which were smaller and less tightly packed as well as of a lighter shade of green than those observed in Aspergillus fumigatus. On the basis of the morphologic features of the ascospores, the teleomorph was identified as N. pseudofischeri.
Propionibacterium acnes as a cause of aggressive aortic valve endocarditis and importance of tissue grinding: case report and review
Authors: Günthard H , Hany A , Turina M , Wüst J
Published: January 2020

A case of prosthetic valve endocarditis with Propionibacterium acnes is described. The diagnosis was documented by histology and isolation of P. acnes from both blood and anulus tissue. Grinding of the tissue, which was first omitted to avoid contamination, was indispensable for cultivating the agent. The literature for P. acnes endocarditis is reviewed.
Abortion associated with Campylobacter upsaliensis
Authors: Gurgan T , Diker K S
Published: January 2020

Campylobacter upsaliensis was isolated from the blood and fetoplacental material of an 18-week-pregnant woman who had contact with a household cat. We believe this is the first report of abortion associated with C. upsaliensis infection.
Nested PCR for specific detection and rapid identification of human picornaviruses
Authors: Kämmerer U , Kunkel B , Korn K
Published: January 2020

A nested PCR for the detection and rapid identification of human picornaviruses is described. Enteroviruses and rhinoviruses were amplified with the same set of four primers from the 5'-noncoding region. The nested primers allowed the detection of far less than 1 PFU in diluted virus stocks without Southern blot hybridization. In patients with neurological disorders (mainly aseptic meningitis), 43% of 37 specimens (11 of 21 cerebrospinal fluid specimens, 2 of 10 serum specimens, and 3 of 6 stool specimens) were positive by PCR. A total of 21% (10 of 47 specimens) of heart biopsy specimens from patients with dilative cardiomyopathy were PCR positive, whereas 3% (2 of 70 specimens) of control biopsy specimens from patients with coronary artery disease were PCR positive. PCR-amplified fragments from 27 of 29 clinical isolates and 14 of 28 patient samples were successfully serotyped by restriction enzyme digestion. Two specimens were further investigated by direct sequencing of PCR products, leading to the identification of a poliovirus type 3 isolate with a sequence that was highly divergent from previously published sequences.
Evaluation of differential inoculum disk diffusion method and Vitek GPS-SA card for detection of oxacillin-resistant staphylococci
Authors: Knapp C C , Ludwig M D , Washington J A
Published: January 2020

This study was conducted in order to compare the accuracy of detection of oxacillin-resistant staphylococci, defined by microdilution MICs, population analyses, and mec gene hybridization, with the Vitek GPS-SA Susceptibility Card with that of the standard inoculum (10(7) CFU) and high-inoculum (10(9) CFU) disk diffusion tests. By the standard inoculum disk diffusion test, 10 of 67 (15%) isolates of oxacillin-resistant Staphylococcus aureus and 3 of 47 (6%) isolates of Staphylococcus epidermidis were falsely susceptible after 24 h of incubation at 35 degrees C. By the high-inoculum disk diffusion test (10(9) CFU), 4 of the 10 isolates of S. aureus remained falsely susceptible, whereas none of the isolates of S. epidermidis was falsely susceptible. Of the 10 isolates of S. aureus falsely susceptible by the standard disk test, only one remained falsely susceptible after an additional 24 h of incubation at 22 degrees C. All four isolates of S. aureus that were falsely susceptible by the high-inoculum disk diffusion test after 24 h of incubation at 35 degrees C became resistant after an additional 24 h of incubation at 22 degrees C. Thus, extended incubation of both the standard and high-inoculum disk diffusion tests increased their accuracy in detecting oxacillin resistance. All isolates of oxacillin-resistant staphylococci were accurately detected with the Vitek software upgrades (6.1 and 7.1) of the GPS-SA card.
Predictive accuracy of disk diffusion test for Proteus vulgaris and Providencia species against five newer orally administered cephalosporins, cefdinir, cefetamet, cefprozil, cefuroxime, and loracarbef
Authors: Biedenbach D J , Jones R N
Published: January 2020

Three members of the tribe Proteeae (Proteus vulgaris, Providencia rettgeri, and Providencia stuartii) were tested against five newer orally administered cephalosporins (cefdinir, cefprozil, cefuroxime, cefetamet, and loracarbef) by the disk diffusion and reference broth microdilution methods. One hundred strains of these organisms were tested to confirm the excessive interpretive error rates that previously had been noted for the disk diffusion test. The results indicate that the suggested disk diffusion breakpoints for cefetamet and cefuroxime were without serious interpretive errors. In contrast, loracarbef, cefdinir, and cefprozil results exceeded acceptable interpretive error rates, with very major (false-susceptible) errors of 4, 5, and 9% respectively. Loracarbef currently has a warning in the National Committee for Clinical Laboratory Standards table footnotes addressing this problem. We recommend including cefdinir and cefprozil in the list of compounds requiring this warning. In addition, MICs of cefazolin, cefaclor, and cephalothin were determined to establish whether a class MIC concept to predict susceptibility for these agents was possible. When the indole-positive Proteus strains are tested, cefazolin MICs can be used to predict MICs of all tested orally administered cephems (8 to 13% total errors, with only a 0 to 1% very major error.
Rapid identification of mycolic acid patterns of mycobacteria by high-performance liquid chromatography using pattern recognition software and a Mycobacterium library
Authors: Glickman S E , Kilburn J O , Butler W R , Ramos L S
Published: January 2020

Current methods for identifying mycobacteria by high-performance liquid chromatography (HPLC) require a visual assessment of the generated chromatographic data, which often involves time-consuming hand calculations and the use of flow charts. Our laboratory has developed a personal computer-based file containing patterns of mycolic acids detected in 45 species of Mycobacterium, including both slowly and rapidly growing species, as well as Tsukamurella paurometabolum and members of the genera Corynebacterium, Nocardia, Rhodococcus, and Gordona. The library was designed to be used in conjunction with a commercially available pattern recognition software package, Pirouette (Infometrix, Seattle, Wash.). Pirouette uses the K-nearest neighbor algorithm, a similarity-based classification method, to categorize unknown samples on the basis of their multivariate proximities to samples of a preassigned category. Multivariate proximity is calculated from peak height data, while peak heights are named by retention time matching. The system was tested for accuracy by using 24 species of Mycobacterium. Of the 1,333 strains evaluated, > or = 97% were correctly identified. Identification of M. tuberculosis (n = 649) was 99.85% accurate, and identification of the M. avium complex (n = 211) was > or = 98% accurate; > or = 95% of strains of both double-cluster and single-cluster M. gordonae (n = 47) were correctly identified. This system provides a rapid, highly reliable assessment of HPLC-generated chromatographic data for the identification of mycobacteria.
Bias of culture techniques for diagnosing mixed Mycobacterium genavense and Mycobacterium avium infection in AIDS
Authors: Kirschner P , Vogel U , Hein R , Böttger E C
Published: January 2020

Disseminated Mycobacterium avium infection is a common complication in late-stage AIDS. We describe a patient seropositive for human immunodeficiency virus with a disseminated infection caused by mycobacteria. Acid-fast rods were visible by microscopy in stool and bone marrow and in specimens from skin, liver, spleen, lung, and lymph nodes. Using molecular biology techniques and standard culture techniques, we found evidence of a mixed mycobacterial infection with M. genavense and M. avium. We suggest that the prevalence of M. genavense infection in patients with AIDS is underestimated because of the bias toward M. avium when using standard techniques for the detection and identification of mycobacteria.
Enzyme-linked immunosorbent assay-enzyme-linked coagulation assay for detection of antibodies to Clostridium botulinum neurotoxins A, B, and E and solution-phase complexes
Authors: Doellgast G J , Triscott M X , Beard G A , Bottoms J D
Published: January 2020

The solution-phase complex assay for toxins A, B, and E from Clostridium botulinum (Elcatech, Inc., Winston-Salem, N.C.) was modified to measure antibody. The addition of unlabeled polyclonal antibodies to a mixture consisting of toxin with chicken antibody and RVV-XA-labeled horse antibody reduces the sensitivity of detection of neurotoxin. This reduction in sensitivity can be used as a measure of the specific antibody titer.
Differentiation of toxigenic from nontoxigenic isolates of Pasteurella multocida by PCR
Authors: Nagai S , Someno S , Yagihashi T
Published: January 2020

A PCR assay was developed for the differentiation of toxigenic Pasteurella multocida subsp. multocida strains, the major etiologic agent for progressive atrophic rhinitis in pigs, from nontoxigenic strains. The PCR targeted a toxA gene encoding a 143-kDa dermonecrotic toxin that is considered to be the central etiologic factor in progressive atrophic rhinitis. toxA fragments were amplified from toxigenic P. multocida isolates but not from nontoxigenic isolates or other bacteria isolated from pigs. The sensitivity of the reaction was as low as 10 pg of chromosomal DNA from a toxigenic strain. The results obtained by PCR of the DNAs of 187 field isolates of P. multocida were consistent with those obtained by the guinea pig skin test and Western blot (immunoblot) analysis. Restriction fragment analysis of the PCR-amplified fragments from 67 field isolates and comparison of the DNA sequences of fragments from capsular serotype A and D strains suggest that the PCR-amplified region, which is considered to encode the major immunologic determinants of the toxin, would be the same among P. multocida strains. The PCR that we describe should be useful for the diagnosis and the etiologic survey of progressive atrophic rhinitis.
Development and evaluation of PCR test for detection of Taylorella equigenitalis
Authors: Bleumink-Pluym N M , Werdler M E , Houwers D J , Parlevliet J M , Colenbrander B , van der Zeijst B A
Published: January 2020

A PCR for the detection of Taylorella equigenitalis, the causative agent of contagious equine metritis, was developed and evaluated. A genus-specific primer-probe set was derived from the 16S ribosomal DNA sequences. The PCR was specific and amplified a 585-bp product from all 64 available T. equigenitalis isolates. This PCR product hybridized with a specific probe in a dot spot assay. A variety of microorganisms from the genital tracts of horses or with a close phylogenetic relationship to T. equigenitalis did not yield a visible PCR product and were all negative in the dot spot hybridization assay. The results of the PCR assay were compared with those of culture by using 191 genital swabs from horses of several breeds. We demonstrate that the sensitivity of the PCR assay is superior to that of culture. The assay is most sensitive when DNA from culture plates incubated for at least 2 days is used. Of the tested samples, 1.5% were positive in the culture assay, whereas 35% were positive in the culture PCR assay. PCR-positive samples were obtained from all breeds tested. This means that many T. equigenitalis-carrying horses go unidentified by the current culturing technique. This affects current views about the spread and control of T. equigenitalis.
Direct detection of Mycobacterium tuberculosis complex in respiratory specimens by a target-amplified test system
Authors: Pfyffer G E , Kissling P , Wirth R , Weber R
Published: January 2020

A total of 938 respiratory specimens (633 sputa, 249 bronchial and tracheal aspirates, and 56 bronchoalveolar lavages) from 589 patients were tested for direct detection of Mycobacterium tuberculosis complex by the Gen-Probe amplified Mycobacterium tuberculosis direct test (MTD), and the results were compared with those of the conventional methods of fluorescence microscopy and cultivation (solid and radiometric media). One series of specimens (n = 515) was decontaminated with N-acetyl-L-cysteine (NALC)-NaOH: the other one (n = 423) was decontaminated with sodium dodecyl (lauryl) sulfate (SDS)-NaOH. Of the specimens decontaminated with NALC, 39 were MTD and culture positive, 455 were MTD and culture negative, 18 were MTD positive and culture negative, and 3 were MTD negative and culture positive, indicating a sensitivity of 92.9% and a specificity of 96.2% for the MTD. Of the specimens decontaminated with SDS, 35 were MTD and culture positive, 372 were MTD and culture negative, 15 were MTD positive and culture negative, and 1 was MTD negative and culture positive, indicating a sensitivity of 97.2% and a specificity of 96.1% for the MTD. After resolution of discrepant results by review of the patients' clinical data, the sensitivity of the MTD was 93.9%, the specificity was 97.6%, the positive predictive value was 80.7%, and the negative predictive value was 99.3% for the NALC series; the corresponding values were 97.4, 96.9, 76.0, and 99.7%, respectively, for the SDS series. In conclusion, the MTD is a highly sensitive and specific technique for detecting M. tuberculosis complex within hours in both smear-positive and smear-negative respiratory specimens.
Evaluation of RapID onE system for identification of 379 strains in the family Enterobacteriaceae and oxidase-negative, gram-negative nonfermenters
Authors: Kitch T T , Jacobs M R , Appelbaum P C
Published: January 2020

The ability of the RapID onE system (Innovative Diagnostic Systems, Inc., Norcross, Ga.) to identify 364 strains in the family Enterobacteriaceae and 15 oxidase-negative, gram-negative, nonfermentative rods was evaluated. Kits were inoculated with no. 2 McFarland standard suspensions, and reactions were interpreted after 4 h of incubation at 35 degrees C. Overall, the method correctly identified (to the species level or to the genus level for salmonellas and non-Shigella sonnei Shigella species) 363 strains (95.8%) without additional tests. For four strains (1.0%), additional tests were required to delineate the correct identification from a range of two or more possibilities; these included one Serratia liquefaciens (Serratia marcescens or Serratia liquefaciens), one Serratia rubidaea (Serratia rubidaea or Serratia odorifera), one Salmonella typhi (Leminorella richardii or Salmonella sp.) and one Yersinia enterocolitica (Yersinia frederiksenii, Yersinia intermedia, or Yersinia enterocolitica). Twelve strains (3.2%) were misidentified or yielded codes with no identification; these comprised one Citrobacter amalonaticus (no identification), three Enterobacter hormaechei (not in the RapID onE database; two Enterobacter amnigenus, one Enterobacter sp.), one Serratia liquefaciens (Enterobacter cloacae), one Serratia rubidaea (no identification), four Serratia fonticola (not in RapID onE database; two Enterobacter aerogenes, one Serratia marcescens, one not identified), one Proteus mirabilis (Proteus penneri), and one Proteus vulgaris (Providencia rustigianii). If the seven strains not included in the database had been excluded, correct identification rates would have risen to 97.6% without additional tests and 98.7% with additional tests, with misidentification rates dropping to 1.3%. The RapID onE system is easy to set up and the results are easy to read, and the system provides an accurate, nonautomated commercially available method for the same-day identification of members of the family Enterobacteriaceae and oxidase-negative, gram-negative nonfermenters.