bluuebunny/crossref_metadata_2025 huggingface dataset:
Automated interpretation of disk diffusion antibiotic susceptibility tests with the radial profile analysis algorithm
Authors: Hejblum G , Jarlier V , Grosset J , Aurengo A
Published: January 2020

An original algorithm referred to as the radial profile analysis algorithm was implemented on a Macintosh Quadra 700 computer to provide an automatic determination of the inhibition zone diameters of antibiotic susceptibility tests performed with the disk diffusion method. After digitization of the petri plate image, each antibiotic disk is recognized and labeled. Pixels of the local zone around each disk are then used for generating a profile pattern that is subjected to decision rules. The resulting estimate of the inhibition zone diameter is then automatically compared with conventional breakpoints for classifying the tested strain in one of the clinical categories of antibiotic susceptibility. The program is also able to request a human reading for some rare plates difficult to interpret. The algorithm accuracy was tested by comparing the results with a combination of independent human measurements performed on the tested plates. The test sample was composed of 98 strains, and 2,552 tests of 40 distinct antibiotics were subjected to the analysis. The difference between the automatic and human diameter estimates was less than 4 mm in 90% of the tests. The agreement between the automatic and human clinical categorizations amounted to 95.5%, and severe (major and very major) disagreements were found in 5.6% of the tests performed with staphylococci but only 0.3% of the tests with gram-negative rods. We conclude that the radial profile analysis algorithm is a solid backbone for an automatic system dedicated to the clinical interpretation of disk diffusion antibiotic susceptibility tests.
Epidemiologic application of a standardized ribotype scheme for Vibrio cholerae O1
Authors: Popovic T , Bopp C , Olsvik O , Wachsmuth K
Published: January 2020

A standardized scheme of 27 different BglI ribotypes and subtypes of Vibrio cholerae O1 strains is proposed on the basis of data from 214 human and environmental strains isolated in 35 countries and 14 U.S. states over the past 60 years. The ribotype patterns obtained are reproducible and stable over time. Seven different but very similar ribotypes (1a to 1g) were observed among 16 strains of the classical biotype. Twenty ribotypes and subtypes were identified among 198 V. cholerae O1 strains of the El Tor biotype. Six different patterns were found among the strains causing the current seventh pandemic. Strains of ribotype 8 originated only in central African countries, while those of ribotype 3 originated mainly in Asia and the Pacific Islands. The most widely distributed strains were those of ribotype 6, which was subdivided into three very similar but still distinguishable subtypes. The present Latin American epidemic is caused by strains of ribotype 5. Strains of this ribotype were isolated from several other geographic locations but can be differentiated from the Latin American strains by other molecular methods. Strains associated with two documented environmental reservoirs exhibited three distinct ribotype patterns; those isolated from patients who ate food from the U.S. Gulf waters were all of ribotype 2, while the strains related to the northeast Australian rivers were of ribotypes 9 and 10. Nontoxigenic V. cholerae O1 strains originating in Latin America and the U.S. Gulf Coast did not form a specific cluster of ribotypes. Ribotyping in combination with other well-defined methods can assist in epidemiologic investigations, helping to trace the movement of strains and to identify their geographic origins.
Analysis of respiratory syncytial virus strain variation in successive epidemics in one city
Authors: Cane P A , Matthews D A , Pringle C R
Published: January 2020

The variability of respiratory syncytial virus isolates from five successive epidemics in an urban population was determined. A total of 187 isolates of respiratory syncytial virus from the southern part of Birmingham, United Kingdom, were classified into subgroups A and B and were then further assigned to genetic lineages. Allocation of isolates into lineages was achieved by reverse transcription of infected cell RNA and then PCR amplification of selected regions of the genome; PCR products were examined by restriction mapping or nucleotide sequencing of parts of the nucleoprotein gene, the small hydrophobic protein gene, and the attachment protein gene. Previous work has shown that estimations of genetic diversity by analysis of genes coding for proteins likely (attachment protein) and unlikely (nucleoprotein and small hydrophobic protein) to be under immune pressure gave concordant results. Six genetic lineages of subgroup A isolates have been defined by this procedure; these isolates differ by up to 20% in the amino acid sequences of their attachment proteins; likewise, subgroup B isolates can be divided into two categories by restriction mapping of parts of their nucleoprotein and attachment protein genes. The same genetic lineages appeared to be present worldwide during the same period. The analysis of isolates from successive epidemics showed that different lineages predominated in each epidemic and that not all lineages were present in every epidemic. Some lineages appeared to increase in numbers over several years and then decline, possibly indicating a buildup of resistance in the community to a particular genotype.
Direct PCR enables detection of Mycoplasma pneumoniae in patients with respiratory tract infections
Authors: Tjhie J H , van Kuppeveld F J , Roosendaal R , Melchers W J , Gordijn R , MacLaren D M , Walboomers J M , Meijer C J , van den Brule A J
Published: January 2020

The sensitivities of three methods of detection of Mycoplasma pneumoniae by a 16S rDNA PCR were compared by using a serial dilution of M. pneumoniae. These methods consisted of a PCR performed directly on cells after a proteinase K pretreatment (direct PCR), a PCR after purification of nucleic acids (DNA-PCR), and a PCR with rRNA sequences as the target after reverse transcription. The direct PCR and the reverse transcription PCR had a sensitivity of 1.5 CFU (approximately 250 genomes). By purification, a 10-fold loss of target DNA occurred, as shown by a 10-fold decrease in sensitivity (15 CFU) of the DNA-PCR. The presence of an excess of human background DNA did not influence the sensitivity of either PCR. The direct PCR was evaluated on samples from patients with respiratory complaints. Direct PCR amplification was possible in 94.9% of the samples, which were tested by amplification of a 326-bp fragment of the beta-globin gene, which was performed to test for the suitability of amplification. Nucleic acid purification was performed on the beta-globin-negative samples, after which only 2% remained negative. A positive correlation between the direct M. pneumoniae PCR and serology, as tested by the microparticle agglutination assay (MAG assay), was found in 88.1% of the cases. A positive MAG assay result was found for samples from 10 (17%) of the patients; samples from 6 (10.2%) of these patients were also positive by PCR. Samples from three patients were found to be positive by the M. pneumoniae PCR and negative by the MAG assay. Persistence of M. pneumoniae, as detected by PCR was observed in three patients. These results indicate that the direct PCR with 16S rDNA could prove to be useful in the detection of M. pneumoniae in respiratory tract samples, although more studies are needed to evaluate the correlation between clinical symptoms and positive test results.
Evaluation of affirm VP Microbial Identification Test for Gardnerella vaginalis and Trichomonas vaginalis
Authors: Briselden A M , Hillier S L
Published: January 2020

A commercial system (Affirm VP Microbial Identification Test; MicroProbe Corp.) for detection of vaginal pathogens was evaluated with 176 consecutive women attending a sexually transmitted disease clinic for genital complaints. Vaginal swab specimens were used for culture of Gardnerella vaginalis and Trichomonas vaginalis, preparation of a vaginal smear for Gram stain interpretation, and wet mount evaluation. An additional swab was used to evaluate the 30-min nonisotopic oligonucleotide probe test. The automated probe system detected G. vaginalis in 69 (95%) of 73 women having > 5 x 10(5) CFU of G. vaginalis per ml by culture, and 20 (43%) of 47 specimens with < or = 5 x 10(5) CFU of G. vaginalis per ml. There were three false positives and four false negatives for the Affirm VP test compared with > 5 x 10(5) CFU of G. vaginalis per ml. The probe system detected G. vaginalis in 57 (90%) of 63 vaginal specimens from women having clue cells on wet mount examination, and in only 3 (3%) of 113 women without clue cells, suggesting that the Affirm probe for G. vaginalis could be used as a surrogate for wet mount examination for clue cells. The T. vaginalis probe was positive for 12 of 12 specimens positive by wet mount and 12 of 15 specimens positive by culture. There were no false positives and three false negatives for the Affirm VP test compared with culture and/or wet mount for T. vaginalis. The Affirm VP Microbial Identification System is a rapid, objective, and automated test for the detection of T. vaginalis and clinically significant levels of G. vaginalis that is comparable to wet mount examination for clue cells and is superior to wet mount examination for the detection of trichomonads.
Detection of penicillin and extended-spectrum cephalosporin resistance among Streptococcus pneumoniae clinical isolates by use of the E test
Authors: Jorgensen J H , Ferraro M J , McElmeel M L , Spargo J , Swenson J M , Tenover F C
Published: January 2020

Increasing penicillin resistance and the initial recognition of resistance to extended-spectrum cephalosporins among Streptococcus pneumoniae isolates have placed greater emphasis on accurate methods for susceptibility testing of clinical isolates. This study has evaluated the use of the E test (AB Biodisk NA, Piscataway, N.J.) for the detection of penicillin and cefotaxime resistance among 147 pneumococcal clinical isolates in three geographically separate laboratories. These included 42 penicillin-resistant (MIC, > or = 2 micrograms/ml) and 14 cefotaxime-resistant (defined here as an MIC of > or = 2 micrograms/ml) isolates. E test strips were applied to the surface of Mueller-Hinton sheep blood agar plates and incubated at 35 degrees C in 5% CO2 for 20 to 24 h. E test MICs were compared with MICs determined with lysed horse blood-supplemented Mueller-Hinton broth in a microdilution format as recommended by the National Committee for Clinical Laboratory Standards. Penicillin MICs agreed within one log2 dilution for 136 of 147 (92.5%) isolates, and cefotaxime MICs agreed within one log2 dilution for 142 of 147 (96.6%) isolates. No very major or major interpretive errors occurred with either penicillin or cefotaxime E test MIC results. There were 9.5 and 5.4% minor interpretive category errors with penicillin and cefotaxime E test MICs, respectively. These data indicate that the E test represents a convenient and reliable method for the detection of penicillin or cephalosporin resistance in pneumococci.
Pneumocystis carinii pneumonia in thoroughbred foals: identification of a genetically distinct organism by DNA amplification
Authors: Peters S E , Wakefield A E , Whitwell K E , Hopkin J M
Published: January 2020

Genetically distinct forms of Pneumocystis carinii infect several mammalian hosts. We report the amplification of P. carinii DNA from samples of two infected thoroughbred foal lungs by using primers designed from the sequence of a P. carinii mitochondrial rRNA gene; these primers also prime the amplification of P. carinii DNA from other hosts. The nucleotide sequence of part of the mitochondrial rRNA gene amplified from P. carinii infecting one of the foals was determined and found to be distinct from that of published rat-, rabbit-, ferret-, and human-derived P. carinii sequences.
Comparison of siderophore production and utilization in pathogenic and environmental isolates of Yersinia enterocolitica
Authors: Chambers C E , Sokol P A
Published: January 2020

Yersinia enterocolitica strains of serotypes lethal to mice have been reported previously to produce an endogenous siderophore. In this study, an ethyl acetate-extractable siderophore was characterized and given the name yersiniophore. Yersiniophore was produced by 16 of 16 human isolates of serotypes O:4, O:4,32, O:8, O:21, and one nonhuman isolate of serotype O:21. It was not produced by isolates of serotype O:3, O:5, or O:9. One strain of Yersinia pseudotuberculosis produced yersiniophore, but strains of Yersinia kristensenii, Yersinia frederiksenii, and Yersinia intermedia did not produce or utilize yersiniophore. Food and water isolates of Y. enterocolitica produced a water-soluble siderophore but not yersiniophore. Sixty-two strains of Y. enterocolitica including 42 isolates from human infections, 2 animal isolates, and 18 water and food isolates were examined for utilization of yersiniophore, the water-soluble siderophore, and ferrioxamine. Yersiniophore promoted growth rate, iron binding, and uptake in 17 of 62 strains, all of which produced yersiniophore. Ten of 17 food and water isolates and one human isolate were capable of utilizing the water-soluble siderophore. Utilization studies suggest that at least one additional water-soluble siderophore may be produced. Ferrioxamine promoted the growth of 60 of 62 strains examined; however, only the 17 strains which produced yersiniophore actively accumulated [59Fe]ferrioxamine. Yersiniophore production and utilization may be important in clinical infections since all human strains belonging to serotype O:8 produced yersinophore. The water-soluble siderophore was not detected in human isolates.
Evaluation of a new immunodiagnostic assay for Helicobacter pylori antibody detection: correlation with histopathological and microbiological results
Authors: Pronovost A D , Rose S L , Pawlak J W , Robin H , Schneider R
Published: January 2020

Infection with Helicobacter pylori has been associated with the pathogenesis of chronic active gastritis and gastric and duodenal ulcer disease. Detection of immunoglobulin G antibodies to H. pylori offers a simple alternative to direct detection of the organism in biopsied tissue by culture or histopathological methods. A rapid flow-through membrane-based enzyme immunoassay for the detection of human immunoglobulin G antibodies to H. pylori has been developed and evaluated. Clinical evaluations were performed with 256 patient serum samples obtained from four clinical sites. Biopsy samples were obtained by endoscopic procedures at the same time as the serum samples, and were histopathologically and microbiologically categorized for the presence or absence of H. pylori. Sensitivity and specificity for this rapid enzyme immunoassay were 92 and 88%, respectively, compared directly with endoscopy results. After discordant results were resolved by a quantitative microwell enzyme-linked immunosorbent assay, the resulting sensitivity and specificity were 94 and > 99%, respectively. These results indicate that this rapid enzyme immunoassay is a useful technique to determine H. pylori infection status and is a viable alternative to invasive endoscopic procedures.
Use of various genetic markers in differentiation of Mycobacterium bovis strains from animals and humans and for studying epidemiology of bovine tuberculosis
Authors: van Soolingen D , de Haas P E , Haagsma J , Eger T , Hermans P W , Ritacco V , Alito A , van Embden J D
Published: January 2020

One hundred fifty-three Mycobacterium bovis strains from cattle, various animal species from zoos and wild parks, and humans were analyzed for three different genetic markers for use in the epidemiology of bovine tuberculosis. M. bovis strains isolated from cattle were found to carry a single IS6110 element, whereas the majority of strains from other animals such as antelopes, monkeys, and seals harbored multiple IS6110 elements, suggesting that the reservoirs in cattle and wild animals are separated. Because the single IS6110 element in cattle strains is located at the same chromosomal position, strain differentiation by insertion sequence fingerprinting was hampered. Therefore, we investigated the usefulness of the direct repeat and polymorphic GC-rich repeat elements for strain differentiation. Both markers allowed sufficient strain discrimination for epidemiological purposes. Evidence is presented that in Argentina, most human M. bovis infections are due to transmission from cattle, whereas M. bovis infections among humans in the Netherlands are mainly contracted from animals other than cattle. Various outbreaks of M. bovis among animals and humans are described, including a small one which likely involved transmission from human to human.
High doses of purified influenza A virus hemagglutinin significantly augment serum and nasal secretion antibody responses in healthy young adults
Authors: Keitel W A , Couch R B , Cate T R , Hess K R , Baxter B , Quarles J M , Atmar R L , Six H R
Published: January 2020

The reactogenicity and immunogenicity of purified influenza virus hemagglutinin (HA) vaccines administered intramuscularly were evaluated in two placebo-controlled clinical trials. A total of 139 healthy young adults were randomized to receive increasing doses of monovalent influenza A/Taiwan/1/86 (H1N1) virus HA (range, 0 to 405 micrograms per dose [study 1]). An additional 139 subjects were given increasing doses of a trivalent HA vaccine containing equal amounts of A/H1N1 virus, A/Shanghai/16/89 (H3N2) virus, and influenza B/Yamagata/16/88 virus HA (range, 0 to 135 micrograms of HA per strain, 0 to 405 micrograms per dose) or a standard dose of commercial influenza vaccine (study 2). Increasing doses of HA were associated with increasing frequencies of symptoms at the vaccination site early after vaccination, but all doses were well tolerated. Occurrence of systemic symptoms was unrelated to dose. Increasing the dose of HA resulted in increasingly higher postimmunization levels of serum hemagglutination inhibiting and neutralizing antibody levels versus influenza A/H1N1 virus in study 1 (P < 0.05); these enhanced responses persisted for up to 6 months. Nasal secretory immunoglobulin A and G antibody responses were assessed 2 weeks after immunization with monovalent H1N1 virus HA; the frequencies of significant responses also increased in a dose-related fashion. Similar increases in serum antibody levels were noted for both A/H1N1 and A/H3N2 viruses in study 2. These data provide a basis for proceeding with the evaluation of high doses of purified HA in the elderly.
Brucella melitensis cell envelope protein and lipopolysaccharide epitopes involved in humoral immune responses of naturally and experimentally infected sheep
Authors: Zygmunt M S , Cloeckaert A , Dubray G
Published: January 2020

Cell envelope fraction (CEF) of Brucella melitensis B115 was used to investigate antibody responses of B. melitensis naturally and strain H38 experimentally infected sheep by immunoblotting, indirect enzyme-linked immunosorbent assay (ELISA) (I-ELISA), and competitive ELISA (C-ELISA) with monoclonal antibodies (MAbs). MAbs used were directed to outer membrane proteins with molecular masses of 10, 16.5, 19, 25 to 27, 31 to 34, 36 to 38, and 89 kDa; to the heat shock protein DnaK, to O-polysaccharide (O-PS) common (C) and M epitopes; and to rough lipopolysaccharide (R-LPS) epitopes. In immunoblotting, all infected sheep sera tested recognized a large number of protein bands, including the above-cited proteins and other proteins for which MAbs have not been defined. The antibody response pattern was different from one animal to another, even within the experimentally infected sheep which were infected under the same experimental conditions. A number of protein bands were recognized by the sheep sera prior to experimental infection and by other uninfected sheep sera. The antibody reactivity to these antigens and others might explain the nonspecific antibody reactivity of sera in I-ELISA with CEF. C-ELISA confirmed also the individual variability of the antibody responses of infected sheep to protein antigens. Antibody responses to O-PS C and M epitopes were detected in all experimentally infected sheep and in half of the naturally infected sheep, but these responses were also heterogeneous in relation to their intensities. Antibody responses to R-LPS epitopes detected by use of C-ELISA with the anti-R-LPS MAbs were low or negative in most of the infected animals. Despite antibody response heterogeneity for CEF antigens, immunoblot and C-ELISA results indicated that, among the CEF antigens, the O-PS epitopes (C and M epitopes) and epitopes of the major 25- to 27- and 31- to 34-kDa outer membrane proteins seem to be the most promising for detecting B. melitensis infection in sheep.
Bacteremia caused by a novel Bordetella species, "B. hinzii"
Authors: Cookson B T , Vandamme P , Carlson L C , Larson A M , Sheffield J V , Kersters K , Spach D H
Published: January 2020

Bordetella spp. cause respiratory tract diseases in warm-blooded animals. Only Bordetella bronchiseptica has been reported to cause bacteremia in humans, and this rare infection usually occurs with pneumonia in immunocompromised patients. We describe "Bordetella hinzii" bacteremia in an AIDS patient without a respiratory illness. Combining biochemical phenotyping with fatty acid analysis permitted preliminary identification of this previously undescribed pathogen; identity was confirmed by DNA-DNA hybridization. This report extends the spectrum of human infections caused by the bordetellae.
Diagnosis of enteroviral meningitis by using PCR with a colorimetric microwell detection assay
Authors: Rotbart H A , Sawyer M H , Fast S , Lewinski C , Murphy N , Keyser E F , Spadoro J , Kao S Y , Loeffelholz M
Published: January 2020

A 5-h, user-friendly PCR assay for the diagnosis of enteroviral meningitis was developed. Reverse transcription and amplification were performed in a one-step reaction using rTth polymerase. Carryover contamination was prevented with dUTP and uracil N-glycosylate. Detection was performed colorimetrically on a microwell titer plate. Sensitivity, specificity, positive predictive value, and negative predictive value were 94.7, 97.4, 94.7, and 97.4%, respectively.
Rapid screening for early detection of mother-to-child transmission of human immunodeficiency virus type 1
Authors: Cassol S , Butcher A , Kinard S , Spadoro J , Sy T , Lapointe N , Read S , Gomez P , Fauvel M , Major C
Published: January 2020

The testing of dried blood spots (DBSs) for the presence of human immunodeficiency type 1 (HIV-1) proviral DNA by PCR was first described in 1991. The technology has proven to be particularly valuable for resolving the infection status in HIV-1-indeterminate infants born to HIV-1-seropositive mothers. To broaden the applicability of DBS PCR, we adapted it to a standardized, commercially available microwell plate amplification and detection kit, Amplicor HIV-1, produced by Roche Diagnostic Systems. The microwell assay is rapid and easy to perform and uses equipment that is readily available in routine diagnostic laboratories. The high level of performance of the assay was demonstrated in 1,168 duplicate tests performed on 584 DBSs from 178 uninfected and 100 HIV-1-infected individuals, including 56 children with perinatally acquired HIV-1. Of 12 infants who were followed prospectively from birth, 3 (25%) were infected in utero (PCR positive at birth) and 9 (75%) were infected intrapartum (PCR negative, culture negative at birth). Overall, HIV-1 DNA was identified in 3 of 11 (27.3%) DBSs collected from infected infants during the first 4 days of life, 8 of 9 (88.9%) DBSs collected between 10 and 15 days postpartum, and 166 of 167 (99.4%) DBSs collected after 15 days of age. All 320 DBSs from uninfected children were PCR DNA negative. These findings indicate that use of the Roche microwell DBS PCR assay provides a powerful new approach for large-scale perinatal screening programs and population-based studies of vertical transmission.
Evaluation of two rapid antigen assays, BioStar Strep A OIA and Pacific Biotech CARDS O.S., and culture for detection of group A streptococci in throat swabs
Authors: Dale J C , Vetter E A , Contezac J M , Iverson L K , Wollan P C , Cockerill F R
Published: January 2020

Two rapid methods, BioStar Strep A OIA (OIA; BioStar, Inc., Boulder, Colo.), an optical immunoassay, and CARDS O.S. (O.S.; Pacific Biotech, Inc., San Diego, Calif.), a color immunochromographic assay, and two culture methods, one with 5% sheep blood agar (SBA) and one with Todd-Hewitt broth (TH; Remel, Lenexa, Kans.), were evaluated for use in the detection of Streptococcus pyogenes from pharnygeal swabs. Seven hundred forty-six double swabs (Culturette II) were processed, with OIA and SBA culture performed on one swab and O.S. and SBA culture performed on the other swab. The pledget from the Culturette II was incubated overnight in TH and was subcultured onto SBA for an additional 48 h in ambient air. All beta-hemolytic streptococci from culture were tested by a direct fluorescent-antibody test (Difco Laboratories, Detroit, Mich.). Specimens with discordant fluorescent-antibody test and rapid test results were also tested by using the Streptex latex agglutination reagent (Murex Diagnostics Limited, Dartford, England). The results obtained by all testing methods were compared with a combined test result ("gold standard"), which was defined as any positive culture detected by the SBA or TH culture methods and confirmed by Streptex latex agglutination or, in the case of negative results by both culture methods, a concomitant positive result by OIA and O.S. antigen testing. Sensitivity and specificity results for each of the methods were as follows, respectively: OIA, 81.0 and 97.5%; O.S., 74.4 and 99.0%; SBA culture, 92.3 and 98.3%; and TH culture, 86.4 and 100%. Both OIA and O.S. are suitable screening methods for detecting S. pyogenes directly from throat swabs but are of insufficient sensitivity to eliminate the need for backup cultures for specimens with negative OIA and O.S. results.
Polyclonal and monoclonal antibody and PCR-amplified small-subunit rRNA identification of a microsporidian, Encephalitozoon hellem, isolated from an AIDS patient with disseminated infection
Authors: Visvesvara G S , Leitch G J , da Silva A J , Croppo G P , Moura H , Wallace S , Slemenda S B , Schwartz D A , Moss D , Bryan R T
Published: January 2020

Microsporidia are primitive, spore-forming, mitochondria-lacking, eukaryotic protozoa that are obligate intracellular parasites. They are known to parasitize almost every group of animals including humans. Recently, microsporidia have increasingly been found to infect patients with AIDS. Five genera (Encephalitozoon, Enterocytozoon, Nosema, Septata, and Pleistophora) of microsporidia are known to infect humans. Enterocytozoon organisms cause gastrointestinal disease in a majority of AIDS patients with microsporidiosis. However, a smaller, but an expanding, number of patients with AIDS are being diagnosed with ocular and disseminated infection with Encephalitozoon hellem. Although microsporidial spores can be identified in clinical samples by a staining technique such as one with Weber's chromotrope stain, identification to the species level is dependent on cumbersome and time-consuming electron microscopy. We have recently isolated and established in continuous culture several strains of E. hellem from urine, bronchoalveolar lavage, and sputum samples from AIDS patients with disseminated microsporidiosis. We developed polyclonal and monoclonal antibodies and PCR primers to a strain of E. hellem that can be used successfully to identify E. hellem from other species of microsporidia either in clinical specimens or in cultures established from clinical specimens. Since patients infected with Encephalitozoon spp. are known to respond favorably to albendazole, identification of the parasite to the species level would be invaluable in the treatment of disseminated microsporidiosis.
Efficient subtyping of pathogenic Yersinia enterocolitica strains by pulsed-field gel electrophoresis
Authors: Najdenski H , Iteman I , Carniel E
Published: January 2020

Yersinia enterocolitica is an enteropathogen that has recently and rapidly expanded over the world. There is a close correlation between the biotypes, serotypes, and phage types of the strains, making it virtually impossible to distinguish isolates of the same serotype with the classical phenotypic markers. In the present study, pulsed-field gel electrophoresis (PFGE) was used to compare the NotI genomic profile (i.e., pulsotype) of 20 strains each of serotypes O:3, O:9, and O:5. Eleven, 12, and 18 different pulsotypes were obtained, respectively, indicating that this technique is very efficient for subtyping pathogenic isolates of Y. enterocolitica. Within strains of serotype O:5, PFGE differentiated two subgroups that corresponded to two biotypes (biotypes 1A and 3). Comparison of the pulsotypes of three strains of biotype 3 and serotype O:3 (referred to as 3/O:3) with those of strains 4/O:3 and 3/O:5 suggested that the pulsotype is closer to the biotype than to the serotype. The pulsotypes of five pairs of strains isolated from the same patient or siblings were also analyzed. In four pairs, the two strains displayed identical pulsotypes, indicating that PFGE might be a powerful epidemiological tool. In the fifth pair, one restriction fragment differed, suggesting that genomic polymorphism may occur in vivo in Y. enterocolitica. Finally, the in vitro genomic stabilities of one strain each of Y. enterocolitica O:3, O:9, and O:5 were investigated. The pulsotypes of 10 isolated colonies were identical within each strain, indicating that in vitro, the genome of Y. enterocolitica is much more stable than that of Y. pestis.
Genetic diversity in human-derived Pneumocystis carinii isolates from four geographical locations shown by analysis of mitochondrial rRNA gene sequences
Authors: Wakefield A E , Fritscher C C , Malin A S , Gwanzura L , Hughes W T , Miller R F
Published: January 2020

The opportunistic fungal pathogen Pneumocystis carinii is a frequent cause of pneumonia in immunocompromised hosts. In this study, we have compared the DNA sequences of a portion of the mitochondrial large-subunit rRNA gene of P. carinii (an informative locus showing up to 27% differences among isolates of P. carinii from human-, rat-, mouse-, ferret-, rabbit-, and horse-infected lungs) obtained from human-derived isolates from widely disparate geographical areas, including Britain, the United States, Brazil, and Zimbabwe. A single-base polymorphism which varied among samples was identified. Apart from this nucleotide, the DNA sequences of all samples were identical. The sequences of the British samples were shown to be stable over a period of 4 years. These data suggest that there is relatively low genetic diversity among isolates of human-derived P. carinii from different global regions.
Use of Gen-Probe AccuProbes to identify Mycobacterium avium complex, Mycobacterium tuberculosis complex, Mycobacterium kansasii, and Mycobacterium gordonae directly from BACTEC TB broth cultures
Authors: Reisner B S , Gatson A M , Woods G L
Published: January 2020

To evaluate the utility of Gen-Probe AccuProbes for the identification of mycobacteria directly from BACTEC TB 12B vials containing acid-fast bacilli, culture results for 11,375 clinical specimens other than blood received from 1 January 1992 to 30 September 1993 were reviewed retrospectively. During this period, a total of 359 of 11,375 BACTEC vials were positive for acid-fast bacilli and were evaluated for mycobacteria with one or more probes: 224 were probed for Mycobacterium tuberculosis complex, 253 were probed for Mycobacterium avium complex, 64 were probed for Mycobacterium kansasii, and 77 were probed for Mycobacterium gordonae. After initial testing with the probes, 75 vials were positive for M. tuberculosis complex, 99 were positive for M. avium complex, 11 were positive for M. kansasii, and 55 were positive for M. gordonae. Repeat testing of vials that were initially probe negative or testing of colonies from subcultures of these vials identified an additional 11 M. tuberculosis, 27 M. avium complex, 1 M. kansasii, and 9 M. gordonae that were not detected on initial screening. On the basis of these data, the percentage of organisms identified directly from the BACTEC TB 12B vials upon initial screening with each of the four AccuProbes was 87.2% for M. tuberculosis complex, 78.6% for M. avium complex, 91.7% for M. kansasii, and 85.9% for M. gordonae.
Clinical significance of serum hepatitis C virus (HCV) RNA as marker of HCV infection
Authors: Marin M G , Bresciani S , Puoti M , Rodella A , Gussago A , Ravaggi A , Pizzocolo G , Albertini A , Cariani E
Published: January 2020

We have evaluated the clinical significance of hepatitis C virus (HCV) RNA determination by analyzing a group of 221 hospitalized patients with abnormal liver function tests. Serum HCV RNA was detected by "nested" PCR amplification followed by nonisotopic hybridization. Of the 200 (90.5%) patients with anti-HCV-positive enzyme-linked immunosorbent assay results, 152 (76%) were RIBA reactive, 47 (23.5%) had indeterminate results, and 1 (0.5%) was nonreactive. Of the 180 (90%) patients positive for anti-HCV and HCV RNA, 138 (76.7%) were RIBA reactive and 42 (23.3%) were RIBA indeterminate. The pattern of RIBA reactivity did not correlate with the presence of HCV RNA. Elevated alanine aminotransferase levels were associated neither with the presence of viremia nor with the RIBA pattern. Histological findings consistent with non-A non-B hepatitis correlated with the presence of HCV RNA but not with the RIBA pattern. HCV RNA was detected in 11 of 21 (52.4%) anti-HCV-negative patients. These 11 patients were either immunosuppressed or in the prodromic phase of acute hepatitis C. Circulating HCV RNA can therefore be described as being predictive of virus-induced liver damage in anti-HCV-positive patients and may be useful in the diagnosis of HCV infection in anti-HCV-negative immunosuppressed patients or in those with early acute infection.
Heterogeneity of rRNA gene restriction patterns of multiresistant serotype 6B Streptococcus pneumoniae strains
Authors: Harakeh H , Bosley G S , Keihlbauch J A , Fields B S
Published: January 2020

Three multiresistant serotype 6B Streptococcus pneumoniae strains were isolated from the middle ear fluids of children undergoing tympanostomy in Atlanta. Because multiresistant 6B pneumococci have been reported to spread from a single clone, the three isolates were compared with 13 other multiresistant 6B pneumococci by hybridization of endonuclease-restricted DNA fragments with a digoxigenin-labeled cDNA probe complementary to 16 and 23S rRNAs (ribotyping). The ear isolates were heterogeneous, whereas six of the other pneumococcal isolates were alike, indicating a need for additional studies to determine the possibility of clonal spread.
Correlation between detection of herpes simplex virus in oral secretions by PCR and susceptibility to experimental UV radiation-induced herpes labialis
Authors: Kriesel J D , Pisani P L , McKeough M B , Baringer J R , Spruance S L
Published: January 2020

We examined the oral secretions of 25 patients for herpes simplex virus (HSV) at the time of and following experimental UV radiation (UVR). HSV was detected in one or more oral secretion specimens in 5 of 12 (42%) cases by cell culture and in 8 of 12 (67%) cases by PCR. On the day of UVR, HSV was detected in 1 of 12 (8%) patients who developed a lip lesion and 2 of 16 (13%) patients who did not (the difference is not significant). We conclude that PCR is more sensitive than culture in the detection of HSV and that HSV is not shed with increased frequency from the oral cavity before the development of UVR-induced herpes labialis.
Asymptomatic carriage of Neisseria meningitidis in a randomly sampled population
Authors: Caugant D A , Høiby E A , Magnus P , Scheel O , Hoel T , Bjune G , Wedege E , Eng J , Frøholm L O
Published: January 2020

To estimate the extent of meningococcal carriage in the Norwegian population and to investigate the relationship of several characteristics of the population to the carrier state, 1,500 individuals living in rural and small-town areas near Oslo were selected at random from the Norwegian National Population Registry. These persons were asked to complete a questionnaire and to volunteer for a bacteriological tonsillopharyngeal swab sampling. Sixty-three percent of the selected persons participated in the survey. Ninety-one (9.6%) of the volunteers harbored Neisseria meningitidis. The isolates were serogrouped, serotyped, tested for antibiotic resistance, and analyzed by multilocus enzyme electrophoresis. Eight (8.8%) of the 91 isolates represented clones of the two clone complexes that have been responsible for most of the systemic meningococal disease in Norway in the 1980s. Age between 15 and 24, male sex, and active and passive smoking were found to be independently associated with meningococcal carriage in logistic regression analyses. Working outside the home and having an occupation in transportation or industry also increased the risk for meningococcal carriage in individuals older than 17, when corrections for gender and smoking were made. Assuming that our sample is representative of the Norwegian population, we estimated that about 40,000 individuals in Norway are asymptomatic carriers of isolates with epidemic potential. Thus, carriage eradication among close contacts of persons with systemic disease is unlikely to have a significant impact on the overall epidemiological situation.
Clotting activity in Staphylococcus schleiferi subspecies from human patients
Authors: Vandenesch F , Lebeau C , Bes M , Lina G , Lina B , Greenland T , Benito Y , Brun Y , Fleurette J , Etienne J
Published: January 2020

Staphylococcus schleiferi subsp. schleiferi is a coagulase-negative staphylococcus, usually present as a contaminant in human specimens. A near relative, S. schleiferi subsp. coagulans, possesses coagulase activity but has not been reported from humans. We here describe three isolates of pseudocoagulase-positive S. schleiferi subsp. schleiferi and one isolate of S. schleiferi subsp. coagulans from human patients. The pseudocoagulase from the S. schleiferi subsp. schleiferi isolates differs from S. aureus staphylocoagulase by being sensitive to a combination of protease inhibitors (aprotinin, N-ethylmaleimide, and heparin). These isolates could all easily be confused with S. aureus in a typical clinical laboratory, since they all possess a heat-stable DNase and promote clotting formation. Moreover, S. schleiferi subsp. coagulans produces protein A, and S. schleiferi subsp. schleiferi expresses a clumping factor (fibrinogen affinity factor). Southern blot hybridization with an S. aureus coa-specific probe revealed no sequence related to the coa gene in any of the S. schleiferi isolates, and their riboprobe profiles and biochemical characteristics were typical of S. schleiferi subspecies, not of S. aureus. This study demonstrates that both subspecies of S. schleiferi can promote clotting of rabbit plasma in the standard tube test for coagulase.
Loofah sponges as reservoirs and vehicles in the transmission of potentially pathogenic bacterial species to human skin
Authors: Bottone E J , Perez A A , Oeser J L
Published: January 2020

Loofah sponges are natural products used as exfoliative beauty aids. As a consequence of tracing a case of Pseudomonas aeruginosa folliculitis to a contaminated loofah sponge, we assessed the role of loofah sponges in supporting the growth of a wide variety of bacterial species. Our data show growth enhancement of sterile loofah fragments for numerous gram-negative (Pseudomonas, Xanthomonas, and Klebsiella) and gram-positive (Enterococcus and group B Streptococcus) species of human and environmental origin. Furthermore, hydrated new, unused loofah sponges undergo a shift in bacterial flora from sparse colonies of Bacillus spp. and Staphylococcus epidermidis to a predominantly gram-negative flora. The growth-promoting potential of loofah sponges (and other exfoliatives) can be further augmented by desquamated epithelial cells entrapped in the loofah fibrous matrix. Therefore, as loofah sponges (and other exfoliatives) can serve as a reservoir and a vehicle for the transmission of potentially pathogenic species to the human skin, we recommend their decontamination with hypochlorite (10%) bleach at regular intervals.
Genetic variation in swine influenza virus A isolate associated with proliferative and necrotizing pneumonia in pigs
Authors: Rekik M R , Arora D J , Dea S
Published: January 2020

A new antigenic variant of H1N1 swine influenza virus A (Sw/QC/5393/91 [QC/91]) has been found to be associated with porcine proliferative and necrotizing pneumonia. Analysis of its genomic RNA by T1 oligonucleotide mapping revealed that considerable genomic divergence exists between QC/91 and the swine influenza viruses currently circulating in North American swine herds. Analysis of the nucleotide sequence of the HA1 region of the hemagglutinin RNA of QC/91, in comparison with those of most common H1N1 human and swine influenza A viruses, showed the presence of multiple point mutations. Two amino acid substitutions appeared to be located in antigenic sites Sb and Ca. This correlates with antigenic variations demonstrated between A/NJ/8/76, A/Sw/WI/49/76, and Québec isolate A/Sw/QC/5393/91 of swine influenza virus A. Another mutation was responsible for the loss of a glycosylation site, which may have also affected the antigenicity. The other mutations seem to have been accumulated progressively over time. This significant constancy in the fixation of mutations with time suggests that genetic diversity of these viruses may best be interpreted as the result of drifts in the population of circulating swine influenza viruses in Québec.
Optical immunoassay for streptococcal pharyngitis: evaluation of accuracy with routine and mucoid strains associated with acute rheumatic fever outbreak in the intermountain area of the United States
Authors: Daly J A , Korgenski E K , Munson A C , Llausas-Magana E
Published: January 2020

The Strep A OIA (BioStar, Inc., Boulder, Colo.) rapid detection system is an intriguing technology that utilizes an immunoassay relying on changes in reflected light to directly detect group A streptococcal antigen from specimens. In this evaluation, 424 routine pediatric throat specimens and 20 simulated oropharyngeal specimens with added mucoid (M type 3, 18) strains were cultured and tested by the Strep A OIA. The respective sensitivities and specificities were as follows: Strep A OIA versus enhanced broth culturing, 84.2 and 95.7%; and streptococcus-SXT agar (BBL Microbiology Systems, Cockeysville, Md.) culturing versus enhanced broth culturing, 82.9 and 98.6%. The Strep A OIA is an 8-min, technologist-friendly, accurate technique with an 89.4% agreement with traditional culturing.
Treponemal infections in hares in The Netherlands
Authors: Lumeij J T , de Koning J , Bosma R B , van der Sluis J J , Schellekens J F
Published: January 2020

Treponemal infections in wild European brown hares (Lepus europaeus) from The Netherlands were diagnosed by means of serological tests for human syphilis and histological demonstration of spirochetes by the Bosma-Steiner silver impregnation method in histological sections of skin lesions. The treponeme should probably be classified as "Treponema paraluisleporis."
Diagnosis of paracoccidioidomycosis by dot immunobinding assay for antibody detection using the purified and specific antigen gp43
Authors: Taborda C P , Camargo Z P
Published: January 2020

The dot immunobinding assay, a rapid, visually read test, was adapted for serodiagnosis and follow-up of paracoccidioidomycosis (PCM). Purified gp43 antigen was tested before and after sodium metaperiodate treatment. To evaluate the assay, it was tested with sera from PCM, histoplasmosis, Jorge Lobo's disease, aspergillosis, candidiasis, and cryptococcosis patients and healthy subjects. Native gp43 gave positive results with all sera from PCM patients and weakly positive results with sera from Jorge Lobo's disease patients (31.3%). No false-positive results were obtained when periodate-treated gp43 was used as the antigen. These results indicate that the dot immunobinding test is sensitive, specific, economical, and fast for serodiagnosis and follow-up studies of PCM.
Detection of equine arteritis virus following amplification of structural and nonstructural viral genes by reverse transcription-PCR
Authors: St-Laurent G , Morin G , Archambault D
Published: January 2020

A reverse transcription (RT)-PCR assay was developed for the detection of equine arteritis virus (EAV) in cell culture supernatant and in horse semen. Four different sets of oligonucleotide primers complementary to sequences located in the 3' end of the polymerase gene (open reading frame [ORF] 1b) and to sequences representing the entire ORFs 3, 4, and 7, which encode for nonstructural (ORFs 3 and 4) or viral nucleocapsid (ORF 7) proteins, were compared for their abilities to amplify the targeted EAV sequences by the RT-PCR procedure. The sensitivities of the RT-PCR for amplification of EAV sequences located in the 3' end of ORF 1b and ORF 4 were 2 median tissue culture infective doses (TCID50s) of viral particles in the EAV-infected cell culture supernatant for both ORFs and 20 and 200 TCID50s of viral particles, respectively, in virus-containing horse semen. The sensitivities were much lower when primers complementary to ORFs 3 and 7 were used in the RT-PCR, with a minimum detection limit of only 2 x 10(4) TCID50s of viral particles in virally infected cell culture supernatant, as determined by analyzing the resulting RT-PCR products on ethidium bromide-stained agarose gels. The specificities of the RT-PCR assays for all primer sets tested were confirmed when the amplified cDNA products of the expected size reacted positively with the corresponding virus-specific digoxigenin-labeled cDNA probes in the chemiluminescence assays. Although the sensitivity of the RT-PCR for amplification of ORF 3 and 7 sequences was lower, all sets or primers were capable of amplifying several cell culture-adapted EAV field isolates when the virus was present in high enough quanities in the test sample. When horse semen samples were analyzed for the presence of EAV by the RT-PCR with primers specific to the ORF 1b 3' end and ORF 4 sequences and by virus isolation in cell cultures, there was 100% concordance among the assays. The RT-PCR assay targeting the 3' end of ORF 1b and/or ORF 4 EAV RNA may be an alternative to conventional methods for the diagnosis of EAV infection in horses.
Alpha-mannosidase: a rapid test for identification of Arcanobacterium haemolyticum
Authors: Carlson P , Kontiainen S
Published: January 2020

A 4-h alpha-mannosidase test for identification of Arcanobacterium haemolyticum strains (n = 139) and differentiation of A. haemolyticum from Actinomyces pyogenes strains (n = 30) and other gram-positive rods was evaluated. Practically all A. haemolyticum strains (138 of 139) and the Listeria monocytogenes type strain were alpha-mannosidase positive, while all A. pyogenes strains and Corynebacterium (n = .25) strains as well as the Rhodococcus equi and Erysipelothrix rhusiopathiae type strains were negative. The rapid alpha-mannosidase test, in conjunction with a Gram stain and catalase and reverse CAMP tests, was useful in identification of A. haemolyticum and in differentiation of A. haemolyticum from A. pyogenes and Corynebacterium spp.