bluuebunny/crossref_metadata_2025 huggingface dataset:
Escherichia coli in bacteremia: O-acetylated K1 strains appear to be more virulent than non-O-acetylated K1 strains
Authors: Frasa H , Procee J , Torensma R , Verbruggen A , Algra A , Rozenberg-Arska M , Kraaijeveld K , Verhoef J
Published: January 2020

A total of 174 blood isolates of Escherichia coli, collected during a 5-year period at the University Hospital Utrecht, were serotyped with rabbit sera against 171 O antigens and 73 capsule (K) antigens. The four most prevalent O-antigen serotypes were O6 (n = 22), O18 (n = 19), O1 (n = 19), and O2 (n = 15). Thirty-one strains were not typeable with any of the O-antigen-typing sera. Of the 148 strains that were subjected to K-antigen serotyping, 34 strains lacked a K antigen and 41 were not typeable with the K-antigen-specific antisera used in the study. K1 was by far the most frequently found K-antigen serotype; this was followed by K2, K53, K5, K13, K7, K(A)28, and K15. Strains possessing a K1 antigen were further classified as either O-acetyl-positive (n = 12) or O-acetyl-negative (n = 21) strains. Retrospective analysis of patients infected with different E. coli isolates--nonencapsulated (n = 23), O-acetylated K1 (n = 12), and non-O-acetylated K1 (n = 21)--revealed clinical differences. More patients suffered from sepsis (94% versus 74%), and a higher rate of mortality was found in the group infected with K1 isolates (18 versus 9%) than in the group infected with nonencapsulated isolates. More patients with severe sepsis (25 versus 10%) and a higher mortality (33 versus 10%) were found in the group infected with O-acetylated K1 isolates than in the group infected with non-O-acetylated isolated. Also, the hospitalization of these patients was prolonged. Thus, O-acetylated E. coli K1 strains seem to be more virulent than non-O-acetylated K1 strains.
Molecular epidemiology of Escherichia coli O157:H7 strains by bacteriophage lambda restriction fragment length polymorphism analysis: application to a multistate foodborne outbreak and a day-care center cluster
Authors: Samadpour M , Grimm L M , Desai B , Alfi D , Ongerth J E , Tarr P I
Published: January 2020

Genomic DNAs prepared from 168 isolates of Escherichia coli O157:H7 were analyzed for restriction fragment length polymorphisms on Southern blots probed with bacteriophage lambda DNA. The isolates analyzed included strains from a recent large multistate outbreak of E. coli O157:H7 infection associated with consumption of poorly cooked beef in restaurants, a day-care center cluster, and temporally and geographically unrelated isolates. E. coli O157:H7 isolates recovered from the incriminated meat and from 61 (96.8%) of 63 patients from Washington and Nevada possessed identical lambda restriction fragment length patterns. The lambda restriction fragment length polymorphisms observed in 11 (91.7%) of 12 day-care center patients were identical, but they differed from that of the strain associated with the multistate outbreak. E. coli O157:H7 from 42 patients temporally or geographically unrelated to either cluster of infection possessed unique and different lambda restriction fragment length patterns, except for paired isolates from three separate clusters of infection. These data demonstrate that the hybridization of DNA digests of E. coli O157:H7 with radiolabelled bacteriophage lambda DNA can be a useful, stable, and discriminatory epidemiologic tool for analyzing the linkage between strains of E. coli O157:H7.
Molecular subtyping of Salmonella enteritidis phage type 8 strains from the United States
Authors: Usera M A , Popovic T , Bopp C A , Strockbine N A
Published: January 2020

Salmonella enteritidis is now the most common serotype of the genus Salmonella reported in the United States. Bacteriophage typing has been helpful for subdividing S. enteritidis strains from different sources in the United States. Most S. enteritidis outbreaks reported were egg related, and the majority of them were caused by strains of phage type 8. To determine whether restriction fragment length polymorphism of the rRNA genes (ribotyping) and of the genomic DNAs from two lysogenic phages from S. enteritidis could be used to discriminate between S. enteritidis phage type 8 strains, we conducted Southern hybridization studies on 24 isolates from different outbreaks and six non-outbreak-associated strains using DNA probes for 16S and 23S rRNA genes and S. enteritidis typing phages 1 and 2 from the Ward typing system (L. R. Ward, J. D. H. de Sa, and B. Rowe, Epidemiol. Infect. 99:291-294, 1987). Of seven restriction endonucleases screened with the probe for rRNA genes, AccI provided the best discrimination between strains; six distinct patterns were observed. AccI ribosomal DNA patterns 1 to 6 were detected among 76.7, 3.3, 6.7, 3.3, 3.3, and 6.7% of isolates tested, respectively. Strains of AccI ribosomal DNA pattern 3 could be further subdivided into two additional patterns by using SmaI. Epidemiologically related strains had identical patterns. No discrimination between strains was achieved by probes for phages 1 and 2. No sequences homologous to the phage I probe were detected among phage type 8 strains, and all strains tested with six restriction enzymes had the same hybridization pattern with the phage 2 probe. These findings demonstrate that ribotyping with AccI and SmaI provides an additional means of discriminating between some phage type 8 strains; however, ribotyping and the phage 2 hybridization results from egg-related outbreak strains support previous findings that these strains are closely related.
Respiratory syncytial virus subgroup B dominance during one winter season between 1987 and 1992 in Vancouver, Canada
Authors: Thomas E , Margach M J , Orvell C , Morrison B , Wilson E
Published: January 2020

A subgroup analysis of 613 specimens submitted to the British Columbia's Children's Hospital from 1987 to 1992 revealed that subgroups A and B of respiratory syncytial virus (RSV) were both circulating in our community, with some predominance for subgroup A during the period from October 1987 to September 1988 (the 1987-88 season) (64%), 1990-91 (60%), and 1991-92 (62%). During 1989-90 subgroup A represented the majority of isolates (80%). Subgroup B predominated during only one season, 1988-89 (94%). No microheterogeneity within subgroups was apparent as judged by the monoclonal antibody reactivity pattern. More male than female children were affected overall, but no sex-related difference between subgroup infections could be detected (P = 0.28). The majority of patients were less than 1 year of age, and no significant association between age and subgroup was detected after stratifying for year (P = 0.64). This is, to our knowledge, the first comprehensive longitudinal RSV subgroup prevalence study from the Pacific Northwest and from Canada.
Evaluation of an enzyme immunoassay for detection of Chlamydia trachomatis in urine of asymptomatic men
Authors: Sanders J W , Hook E W , Welsh L E , Shepherd M E , Quinn T C
Published: January 2020

In a study of 1,486 men attending two sexually transmitted disease clinics, of whom 891 had no symptoms of urethritis, we compared an enzyme immunoassay (EIA) (Baxter-Bartels, formerly Northumbria AntigEnz) of urine sediment to urethral culture for the detection of Chlamydia trachomatis. C. trachomatis prevalence by culture alone was 7.7% in asymptomatic men and 10.9% in symptomatic men. Discrepant results between EIA of urine and urethral culture were evaluated by direct fluorescent-antibody staining (DFA) for elementary bodies in urine sediment or in culture transport media. When chlamydial infection was defined as either a positive urethral culture or positive EIA confirmed by DFA, chlamydia prevalence increased to 8.9% in asymptomatic men and 11.6% in symptomatic men. The urine EIA sensitivity, specificity, and positive and negative predictive values for chlamydia detection in asymptomatic men were 84.8, 99.3, 91.8, and 98.5%, respectively, with nearly identical results for symptomatic men. The sensitivities of urethral culture alone compared with the combination of urethral culture and urine EIA (with DFA confirmation) were 87.3 and 94.3% for asymptomatic and symptomatic men, respectively. The present EIA of urine sediment is both highly sensitive and specific for the detection of C. trachomatis in asymptomatic men, thus providing a noninvasive screening method for chlamydia infection in asymptomatic men attending sexually transmitted disease clinics.
Comparative evaluation of nine kits for rapid diagnosis of infectious mononucleosis and Epstein-Barr virus-specific serology
Authors: Linderholm M , Boman J , Juto P , Linde A
Published: January 2020

Rapid diagnosis of Epstein-Barr virus (EBV)-associated infectious mononucleosis was compared by using nine kits and EBV-specific serology. Specific antibodies indicative of primary EBV infection were detected in 46 of 108 (43%) serum samples of infectious mononucleosis patients. The sensitivities and specificities of the rapid kits varied from 63 to 84% and 84 to 100%, respectively.
An automated enzymatic method for measurement of D-arabinitol, a metabolite of pathogenic Candida species
Authors: Switchenko A C , Miyada C G , Goodman T C , Walsh T J , Wong B , Becker M J , Ullman E F
Published: January 2020

An automated enzymatic method was developed for the measurement of D-arabinitol in human serum. The assay is based on a novel, highly specific D-arabinitol dehydrogenase from Candida tropicalis. This enzyme catalyzes the oxidation of D-arabinitol to D-ribulose and the concomitant reduction of NAD+ to NADH. The NADH produced is used in a second reaction to reduce p-iodonitrotetrazolium violet (INT) to INT-formazan, which is measured spectrophotometrically. The entire reaction sequence can be performed automatically on a COBAS MIRA-S clinical chemistry analyzer (Roche Diagnostic Systems, Inc., Montclair, N.J.). Replicate analyses of human sera supplemented with D-arabinitol over a concentration range of 0 to 40 microM demonstrated that the pentitol could be measured with an accuracy of +/- 7% and a precision (standard deviation) of +/- 0.4 microM. Serum D-arabinitol measurements correlated with those determined by gas chromatography (r = 0.94). The enzymatic method is unaffected by L-arabinitol, D-mannitol, or other polyols commonly found in human serum. Any of 17 therapeutic drugs potentially present in serum did not significantly influence assay performance. Data illustrating the application of the assay in patients for possible diagnosis of invasive candidiasis and the monitoring of therapeutic intervention are presented. The automated assay described here was developed to facilitate the investigation of D-arabinitol as a serum marker for invasive Candida infections.
Diagnosis of genital Chlamydia trachomatis infections in asymptomatic males by testing urine by PCR
Authors: Domeika M , Bassiri M , Mårdh P A
Published: January 2020

An enzyme-linked immunosorbent assay (EIA) (MikroTrak; Syva) was compared with PCR (Amplicor; Roche) for detection of Chlamydia trachomatis in first-void urine (FVU) from 184 men attending a skin and venereal disease clinic. The prevalence of C. trachomatis in the population studied was 18.5%. Discrepant results between Syva EIA and Roche PCR were retested by using major outer membrane protein primer-based PCR. After retesting, the sensitivity, the specificity, and the positive and negative predictive values for the Syva EIA were 85.3, 100, 100, and 77.5%, respectively, and those for the Roche PCR 100, 100, 100, and 100%, respectively. It was concluded that PCR provides a highly sensitive and specific noninvasive screening method for genital chlamydial infection in asymptomatic men.
PCR for capsular typing of Haemophilus influenzae
Authors: Falla T J , Crook D W , Brophy L N , Maskell D , Kroll J S , Moxon E R
Published: January 2020

A PCR method for the unequivocal assignment of Haemophilus influenzae capsular type (types a to f) was developed. PCR primers were designed from capsule type-specific DNA sequences cloned from the capsular gene cluster of each of the six capsular types. PCR product was amplified only from the capsular type for which the primers were designed. Product was confirmed by using either an internal oligonucleotide or restriction endonuclease digestion. A total of 172 H. influenzae strains of known capsular type (determined genetically) comprising all capsular types and noncapsulate strains were tested by PCR capsular typing. In all cases the PCR capsular type corresponded to the capsular genotype determined by restriction fragment length polymorphism analysis of the cap region. When used in conjunction with PCR primers derived from the capsular gene bexA, capsulate, noncapsulate, and capsule-deficient type b mutant strains could be differentiated. PCR capsular typing overcomes the problems of cross-reaction and autoagglutination associated with the serotyping of H. influenzae strains. The rapid and unequivocal capsular typing method that is described will be particularly important for typing invasive H. influenzae strains isolated from recipients of H. influenzae type b vaccine.
Sensitivities of radioimmunoprecipitation assay and PCR for detection of human T-lymphotropic type II infection
Authors: Gallo D , Penning L M , Diggs J L , Hanson C V
Published: January 2020

Five hundred forty-eight uncoagulated blood specimens from intravenous drug users infected with human T-lymphotropic virus type II (HTLV-II) were used to evaluate the sensitivities of the radioimmunoprecipitation assay (RIPA) and PCR for detecting HTLV-II-infected people. The sensitivities of both RIPA and PCR were found to be dependent on the HTLV-II antibody titer, as determined by the immunofluorescence assay. Neither of these recommended confirmatory methods was as sensitive for detecting weakly reactive HTLV-II specimens as the immunofluorescence assay, Western blotting (immunoblotting), or a modified licensed enzyme immunoassay. Use of RIPA and PCR to determine the reliabilities of other tests may sometimes give erroneous results.
Distribution of Borrelia burgdorferi in host mice in Pennsylvania
Authors: Lord R D , Lord V R , Humphreys J G , McLean R G
Published: January 2020

Host mice (Peromyscus leucopus and Peromyscus maniculatus) were sampled throughout the state of Pennsylvania to determine the geographical and ecological distribution of the Lyme disease spirochete Borrelia burgdorferi. All 67 counties of the state were sampled. A total of 1,619 mice were captured from a total of 157 sites during the period 1990 to 1993 for an overall capture rate of 29.69%. A total of 112 (6.92%) isolations of B. burgdorferi were made. The distribution of isolations revealed the reason for the correlated distribution of human cases of Lyme disease in the state. Significantly more mice were captured and significantly more isolations were made from hemlock (Tsuga canadensis) habitat than from deciduous species forest. Nevertheless, high isolation rates from counties of the southeastern corner of the state illustrate well that hemlock habitat is not essential. Evidence suggests that in some areas, transmission between mice is occurring in some way other than through ticks as vectors. Host mice proved useful for determining the geographical and ecological distribution of B. burgdorferi.
Multicenter evaluation of a fully automated screening test, VIDAS HIV 1 + 2, for antibodies to human immunodeficiency virus types 1 and 2
Authors: Azevedo-Pereira J M , Lourenço M H , Barin F , Cisterna R , Denis F , Moncharmont P , Grillo R , Santos-Ferreira M O
Published: January 2020

A multicenter study was done to evaluate the sensitivity, specificity, and efficiency of a new screening test for the simultaneous detection of human immunodeficiency virus type 1 and 2 (HIV-1 and HIV-2) antibodies. The VIDAS HIV 1 + 2 (bioMérieux, Marcy l'Etoile, France) is a fully automated enzyme-linked fluorescent immunoassay that uses synthetic peptides from immunodominant regions of gp41 of HIV-1 and gp36 of HIV-2 as antigens. A total of 2,984 samples were evaluated with this system in six different laboratories, and the results were compared to those obtained with other enzyme-linked immunosorbent assays. The VIDAS HIV 1 + 2 assay showed a very good performance in terms of sensitivity (100%) and specificity (99.6%), requiring minimal manipulation and short incubation time (32 min) to give results similar to or better than those of the other enzyme-linked immunosorbent assays used for screening.
Testing antimicrobial susceptibilities of adherent bacteria by a method that incorporates guidelines of the National Committee for Clinical Laboratory Standards
Authors: Domingue G , Ellis B , Dasgupta M , Costerton J W
Published: January 2020

Membrane filters (0.22-microns pore size) were colonized with 10(4) CFU of logarithmic-phase bacteria per filter under laminar flow conditions in a Modified Robbins Device. Colonized filters were then placed upon agar dilution plates for MIC determinations. Subsequently, filters were transferred to control plates and incubated to obtain MBCs.
Optimum recovery of Mycobacterium avium complex from blood specimens of human immunodeficiency virus-positive patients by using small volumes of isolator concentrate inoculated into BACTEC 12B bottles
Authors: Doern G V , Westerling J A
Published: January 2020

Recovery of Mycobacterium avium complex organisms from 412 lysis-centrifugation (Isolator) concentrates of blood specimens obtained from human immunodeficiency virus-positive individuals was attempted with the following media (and Isolator concentrate inoculum volumes): BACTEC 12B broth (0.2 and 1.0 ml), Lowenstein-Jensen slants (0.1 ml), and Middlebrook 7H10/11 agar (0.1 ml). A total of 42 M. avium complex isolates were recovered. The highest rates of recovery and shortest detection times were noted with BACTEC 12B bottles inoculated with 0.2 ml of Isolator concentrate. Middlebrook agar was superior to Lowenstein-Jensen slants. Fluorochrome acid-fast smears performed directly upon Isolator concentrates were of no utility.
Plasmodium vivax ookinetes in human peripheral blood
Authors: Hummert B A
Published: January 2020

Microscopic examination of a peripheral blood smear revealed ookinetes of Plasmodium vivax. This unusual finding was probably due to an excessive delay between blood collection and smear preparation. Ookinete formation normally occurs in the mosquito gut. When seen in blood smears, it can cause confusion and misidentification of the parasite.
Performance of third-generation confirmatory tests for detection of antibody to hepatitis C virus
Authors: Chaudhary R K , Jacobsen H
Published: January 2020

We investigated three immunoblot assays (RIBA 3.0 from Chiron, Matrix from Abbott Laboratories, and LiaTek III from Organon Teknika) for the detection of antibody to hepatitis C virus. RIBA 3.0 and Matrix require reactivity to two antigens and LiaTek III requires reactivity to one for a sample to be positive. We tested 80 samples that were positive in repeat enzyme immunoassays by supplemental tests. The results showed that 55, 46, and 28% were reactive by LiaTek III, RIBA 3.0, and Matrix, respectively; 54, 33, and 13% of the samples were indeterminate by Matrix, RIBA 3.0, and LiaTek III, respectively. There were 32, 21, and 16% nonreactive samples by LiaTek III, RIBA 3.0, and Matrix, respectively. Of the samples positive by RIBA 3.0, only 50 and 76% were reactive by Matrix and LiaTek III, respectively. A large number of samples that were indeterminate by RIBA 3.0 were positive by LiaTek III (52%). The core antigen was the most reactive antigen in all three tests (48 to 57%). The NS4 antigen in Matrix (20%) and LiaTek III (16%) was poorly reactive, although it performed better in RIBA 3.0 (45%). The NS5 and E2/NS1 antigens made minor contributions to reactivity. The combinations of the core, NS3, and NS4 antigens produced 77% of the RIBA 3.0 and 100% of the Matrix reactive samples. The results showed a poor correlation among the three tests.
Clonal outbreaks of extended-spectrum beta-lactamase-producing strains of Klebsiella pneumoniae demonstrated by antibiotic susceptibility testing, beta-lactamase typing, and multilocus enzyme electrophoresis
Authors: Nouvellon M , Pons J L , Sirot D , Combe M L , Lemeland J F
Published: January 2020

Nineteen extended-spectrum beta-lactamase (ESBla)-producing Klebsiella pneumoniae isolates from Rouen Hospital were investigated for their implication in nosocomial outbreaks: in addition to antibiotic susceptibility testing, the ESBlas were characterized by isoelectric focusing, and the genetic relationships between the strains were analyzed by multilocus enzyme electrophoresis using a combined polyacrylamide electrophoresis-electrophoretic transfer technique. Four isoelectric focusing beta-lactamase patterns and 11 enzyme electrophoretic types (ETs) among the strains tested were described. Three strains isolated in the same neurological unit over a 7-day period exhibited an SHV 3 beta-lactamase (pI 7.0) and were assigned to a common ET. Three of five strains isolated from patients in a rehabilitation center over a 6-week period harbored an SHV 4 beta-lactamase (pI 7.8) and exhibited the same ET. These results differentiate nosocomial transmission from sporadic cases and provide evidence that multilocus enzyme electrophoresis is a potential tool for studying genetic relationships between strains harboring a common ESBla.
Routine identification of Mycobacterium tuberculosis complex isolates by automated hybridization
Authors: Mabilat C , Desvarenne S , Panteix G , Machabert N , Bernillon M H , Guardiola G , Cros P
Published: January 2020

Methodologies for biochemical identification of mycobacteria isolated from clinical samples are still cumbersome, taking skilled technicians 3 to 6 weeks. We describe here a 2-h identification system for mycobacterial isolates belonging to the Mycobacterium tuberculosis complex using a DNA probe. After 30 min of hands-off sample preparation, the 1.5-h hybridization test is totally automated in the newly developed VIDAS system (bioMérieux, Marcyl'Etoile, France), which performs solid-phase specific hybridization of 16S rRNA at 37 degrees C. The strain collection of actinomycetes tested was composed of 662 isolates from 27 species: 461 members of the M. tuberculosis complex (443 M. tuberculosis, 10 M. bovis, and 8 M. bovis BCG isolates) and 201 isolates of other species, including 55 M. avium-intracellulare isolates). They were identified by traditional methods: growth rate, colonial morphology, pigmentation, and biochemical profiles. The automated probe assay displayed an excellent correlation with the reference results. The four members of the Nocardia and Rhodococcus genera tested did not cross-hybridize. This flexible random-access and automated technology was shown to suit the routine context of the laboratory by rapidly delivering the results.
Surgical wound infection caused by Rahnella aquatilis
Authors: Maraki S , Samonis G , Marnelakis E , Tselentis Y
Published: January 2020

Rahnella aquatilis is a water-residing gram-negative rod, a member of the family Enterobacteriaceae, isolated rarely from clinical specimens of immunocompromised patients. A case of a surgical wound infection caused by R. aquatilis in a patient who underwent a prosthetic surgical intervention is reported. The presence of inducible beta-lactamase was suggested by the disk induction test and the conventional agar dilution assay. Literature on R. aquatilis infections in humans is reviewed.
Comparison of six typing methods for Actinobacillus actinomycetemcomitans
Authors: van Steenbergen T J , Bosch-Tijhof C J , van Winkelhoff A J , Gmür R , de Graaff J
Published: January 2020

Actinobacillus actinomycetemcomitans is an important pathogen in the etiology of severe periodontitis. For epidemiological studies on the prevalence of certain pathogenic clones and transmission of this bacterium, adequate typing methods are necessary. The purpose of this study was to compare six different typing methods for A. actinomycetemcomitans. Five reference strains and 27 fresh clinical isolates from periodontitis patients were used. Serotyping showed 12 serotype a strains, 13 type b strains, 6 type c strains, and 1 nontypeable strain. Biotyping on the basis of the fermentation of mannose, mannitol, and xylose resulted in six biotypes. Antibiogram typing was evaluated by measuring the inhibition zones of seven antibiotics in agar diffusion tests. With this method eight main types which could be further differentiated into 15 subtypes were found. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis patterns of outer membrane proteins were similar among all isolates tested. Restriction endonuclease analysis (REA) of whole chromosomal DNA resulted in five main types. These five main types were further differentiated into 24 subtypes on the basis of DNA fragment differences in the high-molecular-weight region. Hybridization of DNA fragments with ribosomal DNA (ribotyping) resulted in 22 to 24 different types, depending on the restriction endonuclease used. Ribotype patterns were easy to interpret and provided an univocal distinction between different strains compared with REA results. When applied to epidemiologically related isolates, all methods were able to discriminate two clonal types among five isolates from five children from one family. We conclude that serotyping, biotyping, and outer membrane patterns were reproducible but had a low discriminatory potential. REA and ribotyping were reproducible and gave the highest number of distinct types. When the DNA typing methodis were compared, all strains tested could be distinguished. These findings confirm the heterogeneity found within the species A. actinomycetemcomitans.
Use of an ATP bioluminescent assay to evaluate viability of Pneumocystis carinii from rats
Authors: Chen F , Cushion M T
Published: January 2020

A bioluminescent assay which employs the luciferin-luciferase ATP-dependent reaction was used to evaluate the viability of populations of Pneumocystis carinii derived from infected rat lungs. Contamination with host cells was reduced by a purification method which involved a combination of low- and high-speed centrifugations resulting in a 1,000-fold reduction of the rat cells while enriching for the trophic form of P. carinii. A linear correlation for the number of P. carinii nuclei versus the amount of ATP was observed. The ATP content of the organism populations could be maintained at inoculum levels for one week, although the number of organisms did not increase. Addition of respiratory chain inhibitors dramatically decreased the ATP content of the P. carinii after 24 h of incubation, with the exception of the antibiotic oligomycin B. Low concentrations of trimethoprim-sulfamethoxazole and pentamidine isethionate reduced the organism ATP content by over 50% after 24 h of exposure, while no effect was observed with 100-fold greater concentrations of ampicillin. The bioluminescent assay was found to be a more sensitive indicator of viability than a dual fluorescent staining technique. This assay does not require replication of P. carinii and should be a useful method for in vitro drug screening and viability assessment of P. carinii populations.
Validation of respiratory syncytial virus enzyme immunoassay and shell vial assay results
Authors: Pedneault L , Robillard L , Turgeon J P
Published: January 2020

The Pathfinder respiratory syncytial virus (RSV) enzyme immunoassay (EIA) (Kallestad), the shell vial (SV) technique, and conventional cell culture (CC) were compared for detection of RSV in nasopharyngeal aspirates. We found sensitivities, specificities, and positive and negative predictive values of 58.4, 100, 100, and 68.2%; 80.7, 97.2, 97.0, and 81.9%; and 77.6, 97.2, 96.9, and 79.5% for the CC, EIA, and SV methods, respectively. The SV and EIA techniques were both more sensitive than CC (P < 0.001). Finally, 29 respiratory viruses other than RSV were identified by CC.
Misidentification of clinical yeast isolates by using the updated Vitek Yeast Biochemical Card
Authors: Dooley D P , Beckius M L , Jeffrey B S
Published: January 2020

The Vitek Yeast Biochemical Card (YBC) is widely used as a rapid identification (RI) (within 48 h) system for clinical yeast isolates. We compared the RI results obtained by the YBC technique with matched results obtained with the API 20C system. The RI of germ tube-negative yeasts isolated from 222 clinical specimens was performed with the YBC system, and the results were compared with those of standard identifications obtained by using the API 20C system and morphology, with additional biochemical reactions performed as required. Commonly isolated yeasts (Candida albicans [n = 29], Candida tropicalis [n = 40], Torulopsis [Candida] glabrata [n = 28], Candida parapsilosis [n = 12], and Cryptococcus neoformans [n = 14]) were generally well identified (115 of 123 [93%] identified correctly, with only C. albicans, C. tropicalis, and C. neoformans mis- or unidentified more than once). The RI of less commonly isolated yeasts included in the YBC database, however, was less successful (54 of 99 [55%] correct). The YBC card failed to identify 42% (10 of 24) of Candida krusei isolates, 80% (4 of 5) of Candida lambica isolates, 88% (7 of 8) of Trichosporon beigelii isolates, and 83% (10 of 12) of Cryptococcus isolates (non-C. neoformans species). For most identification failures (79%; 42 of 53) there was no identification by the end of 48 h; the other identification failures (21%; 11 of 53) gave definite but incorrect identifications. Of eight rare clinical yeast isolates not included in the Vitek database, six were correctly, not identified, while two (25%) were falsely assigned a definite RI (one Hansenula fabianii isolate was identified as Rhodotorula glutinis, and one Hansenula isolate [non-Hansenula anomala] was identified as Hansenula anomala). While the Vitek YBC rapidly and adequately identifies common yeast isolates, it fails in the RI of more unusual organisms.
Early diagnosis of Lassa fever by reverse transcription-PCR
Authors: Demby A H , Chamberlain J , Brown D W , Clegg C S
Published: January 2020

We developed a method based on a coupled reverse transcription-PCR (RT-PCR) for the detection of Lassa virus using primers specific for regions of the S RNA segment which are well conserved between isolates from Sierra Leone, Liberia, and Nigeria. The specificity of the assay was confirmed by Southern blotting with a chemiluminescent probe. The assay was able to detect 1 to 10 copies of a plasmid or an RNA transcript containing the target sequence. There was complete concordance between RT-PCR and virus culture for the detection of Lassa virus in a set of 29 positive and 32 negative serum samples obtained on admission to the hospital from patients suspected of having Lassa fever in Sierra Leone. Specificity was confirmed by the failure of amplification of specific products from serum samples collected from 129 healthy blood donors in Sierra Leone or from tissue culture supernatants from cells infected with related arenaviruses (Mopeia, lymphocytic choriomeningitis, Tacaribe, and Pichinde viruses). Sequential serum samples from 29 hospitalized patients confirmed to have Lassa fever were tested by RT-PCR and for Lassa virus-specific antibodies by indirect immunofluorescence (IF). RT-PCR detected virus RNA in 79% of the patients at the time of admission, comparing favorably with IF, which detected antibodies in only 21% of the patients. Lassa virus RNA was detected by RT-PCR in all 29 patients by the third day of admission, whereas antibody was detectable by IF in only 52% of the patients. These results point to an important role for RT-PCR in the management of suspected cases of Lassa fever.
Pneumococci in nasopharyngeal samples from Filipino children with acute respiratory infections
Authors: Lankinen K S , Leinonen M , Tupasi T E , Haikala R , Ruutu P
Published: January 2020

The presence of Streptococcus pneumoniae in the upper respiratory tract was studied in 318 Filipino children less than 5 years old with an acute lower respiratory tract infection. Nasopharyngeal samples were obtained from 292 children. With both quantitative bacterial culture and detection of capsular polysaccharide antigens by coagglutination, counterimmunoelectrophoresis, and latex agglutination, pneumococci were found in 160 (70%) of the 227 samples eligible for analysis. Culture was positive in 115 samples and antigen was positive in 140 samples. The culture isolation rate was significantly lower if the patient had received antimicrobial agents in the 48 h prior to the sampling. The seven most common types or groups of pneumococci were 6, 14, 19, 23, 15, 7, and 11, which together accounted for 64% of all pneumococcal findings.
Evaluation of new transport medium for detection of herpes simplex virus by culture and direct enzyme-linked immunosorbent assay
Authors: Ogburn J R , Hoffpauir J T , Cole E , Hood K , Michael D , Nguyen T , Raden S , Raju B , Reisinger V , Oefinger P E
Published: January 2020

The transport medium Multi-Microbe Media (M4) was evaluated prospectively by culture and direct enzyme-linked immunosorbent assay (ELISA) for detection of herpes simplex virus from 473 specimens. In addition, 377 specimens in Bartels Viral Transport Medium were evaluated. By using culture as a "gold standard," the ELISA sensitivity was approximately 85%, while the specificities exceeded 96% for both media.
Serotypic differentiation of rotaviruses in field samples from diarrheic pigs by using nucleic acid probes specific for porcine VP4 and human and porcine VP7 genes
Authors: Rosen B I , Parwani A V , Lopez S , Flores J , Saif L J
Published: January 2020

Of 216 fecal and intestinal samples collected from nursing or weaned diarrheic pigs in the United States and Canada, 57 were identified as group A rotavirus positive by RNA electrophoresis and silver staining. Fifty-seven and 52 rotavirus-positive samples were analyzed by hybridization with Gottfried and OSU PCR-derived gene 9 and 4 probes, respectively. Only 17 samples were identified with either homologous VP4 (P)- or VP7 (G)-coding genes or both. One rotavirus identified as G4 and P7 was similar to the previously characterized interserotype rotavirus, SB-1A. Additional hybridization analyses were performed with PCR-derived probes prepared from gene 9 cDNA of the human rotaviruses Wa (G1), DS-1 (G2), and P (G3) and of the porcine rotavirus YM (G11). Eleven of 52 samples collected and analyzed from swine in Ohio, California, and Nebraska were identified as G11. No samples with G1-, G2-, or G3-type specificities were detected among the 25 of 57 rotavirus-positive samples analyzed with human rotavirus-derived probes. Further investigations with a PCR-derived gene 4 probe prepared from porcine rotavirus YM revealed hybridization specificities similar to those of the OSU gene 4 probe.
Correlation between antibiotic resistance, phage-like particle presence, and virulence in Rhodococcus equi human isolates
Authors: Nordmann P , Keller M , Espinasse F , Ronco E
Published: January 2020

Rhodococcus equi is a gram-positive coccobacillus that appears to be emerging as a pulmonary pathogen in AIDS patients. In four human clinical isolates, two antibiotic resistance phenotypes were found to coexist: one beta-lactam resistant and the other beta-lactam susceptible. In vitro, beta-lactam-resistant mutants were obtained at a frequency of 1 x 10(-5) to 5 x 10(-5) from beta-lactam-susceptible strains on cephalothin-containing plates. Neither beta-lactamase nor plasmid DNA was detected in beta-lactam-resistant or -susceptible strains. The penicillin-binding protein patterns for the two types of strains were identical. Electron microscopy revealed that the beta-lactam-resistant strains possessed cell-surface-associated appendages and produced phage-like particles. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of total cell protein showed at least three additional bands of 42, 39, and 30 kDa found only in the beta-lactam-resistant strains. Testing for virulence in Swiss mice revealed that (i) phage-like-particle-producing strains had lower 50% lethal doses when injected intravenously in euthymic and nude mice than the non-phage-like-particle-producing strains did and (ii) intravenous inoculation of a sublethal dose (5 x 10(6) CFU) in nude mice led to chronic infection by the phage-like-particle-producing bacteria only. Finally, in vitro growth curves indicated that the phage-like-particle-producing strains possessed an ecological selection advantage. These results suggest that, among R. equi human isolates, the antibiotic resistance phenotype is associated with virulence and may be phage mediated.
Monoclonal antibody to native P39 protein from Borrelia burgdorferi
Authors: Sullivan T J , Hechemy K E , Harris H L , Rudofsky U H , Samsonoff W A , Peterson A J , Evans B D , Balaban S L
Published: January 2020

We have produced, by using a sonicate of Borrelia burgdorferi, a monoclonal antibody (MAb), NYSP39H, that is specific for the P39 protein band. This MAb reacted with 13 isolates of B. burgdorferi but not with eight different spirochetes (four borrelias, two leptospiras, and two treponemas). Surface labeling of B. burgdorferi with biotin and subsequent treatment with Nonidet P-40 showed that P39 was not biotinylated but was extracted with Nonidet P-40, indicating that it is present within the outer membrane, but not on the surface of the spirochete. Immunoelectron microscopy revealed the immunogold probe primarily at the cytoplasmic membrane region of the spirochete. The MAb detected B. burgdorferi in the indirect fluorescent-antibody test only when the spirochetes from a culture or in a tick homogenate were fixed with polylysine and not with acetone. NYSP39H appears to be an appropriate probe for use in the specific detection of B. burgdorferi.
Variation of DNA sequence in immediate-early gene of human herpesvirus 6 and variant identification by PCR
Authors: Yamamoto T , Mukai T , Kondo K , Yamanishi K
Published: January 2020

The complete nucleotide sequence of one of the immediate-early genes of human herpesvirus 6 variant B was determined and compared with that of variant A reported by Martin et al. (M.D. Martin, J. Nicholas, B. J. Thomson, C. Newman, and R. W. Honess, J. Virol. 65:5381-5390, 1991). While it was reported that two open reading frames exist in this region of variant A, only one open reading frame was found in variant B and the putative coding region of variant B was 2,679 nucleotides long. Furthermore, two additive regions of 108 and 228 bp were found in variant B. Primers covering one of these regions deleted in variant A were synthesized and used for PCR amplification. Twelve isolates from patients were clearly classified into variants A and B by PCR amplification with these primers. All isolates from patients with exanthem subitum were variant B.
Identification of a highly cross-reactive outer surface protein B epitope among diverse geographic isolates of Borrelia spp. causing Lyme disease
Authors: Shoberg R J , Jonsson M , Sadziene A , Bergström S , Thomas D D
Published: January 2020

The outer surface lipoprotein B (OspB) of Borrelia burgdorferi is a major component of the borrelial protein profile and has been shown to be highly immunogenic in experimentally immunized and infected mammals. However, the ospB loci of different strains show considerable heterology at the nucleic acid sequence level, and the progeny of a clonal strain of B. burgdorferi exhibited OspB polymorphisms with respect to apparent molecular weights and reactivities with monoclonal antibodies. These data suggest that OspB is not a good candidate for vaccination or diagnostic purposes. The present study describes a monoclonal antibody, designated 84C, directed against a very highly conserved domain of the OspB lipoprotein. Western immunoblot analysis with 84C demonstrated reactivity in 84.2% of human, tick, and other vertebrate isolate strains examined from widely diverse geographic regions, including strains of B. burgdorferi sensu stricto and two closely related species, B. garinii and B. afzelii. The 84C-binding region was delimited to a highly conserved 11-amino-acid region in the carboxyl terminus of OspB as demonstrated by (i) DNA sequence analysis of wild-type and 84C-resistant mutant ospB alleles and (ii) deletion mutagenesis of a recombinant ospB gene in Escherichia coli. Finally, the 84C epitope was demonstrated to be exposed on the borrelial surface in situ as (i) the monoclonal antibody 84C was able to agglutinate borrelias in culture and (ii) 84C-resistant escape variants were isolated. These data suggest that the potential value of OspB as a vaccine candidate or diagnostic tool be examined more closely, in the context of the 84C-reactive domain.
Hemagglutinating properties of enteroaggregative Escherichia coli
Authors: Qadri F , Haque A , Faruque S M , Bettelheim K A , Robins-Browne R , Albert M J
Published: January 2020

Many intestinal bacterial pathogens possess hemagglutinating properties, which are indicative of their adhesive properties to the intestinal mucosal surface. To understand the bacteria-mucosa interaction, 41 strains of enteroaggregative Escherichia coli (EAggEC), a recently described category of diarrheagenic E. coli, isolated mostly from children with diarrhea in Bangladesh, India, Thailand, Central America, and South America were screened for mannose-sensitive hemagglutination and mannose-resistant hemagglutination of erythrocytes from humans, rats, mice, sheep, cattle, and rabbits. Some strains demonstrated mannose-sensitive hemagglutination of erythrocytes. Most isolates showed mannose-resistant hemagglutination of erythrocytes from all species except rabbits. The hemagglutination patterns could be classified into 18 groups. Studies with three selected isolates suggested that hemagglutinins are cell bound and are protein in nature. On the basis of the pattern of inhibition of hemagglutination by various chemicals, 39 isolates were classified into 19 groups. Hemagglutinations of many isolates were inhibited by sialic acid-containing compounds, suggesting that these compounds may be the receptors for these organisms on erythrocytes and possibly on the intestinal mucosa. These data indicate that strains of EAggEC are a heterogeneous group of organisms with different types of hemagglutinins or adhesins for the intestinal mucosal surface. Also, the adhesion characteristics of EAggEC strains may be too complex to be assessed by simple hemagglutination tests.
Improved PCR sensitivity for direct genotyping of Chlamydia trachomatis serovars by using a nested PCR
Authors: Lan J , Ossewaarde J M , Walboomers J M , Meijer C J , van den Brule A J
Published: January 2020

Successful amplification of omp1 DNA by PCR is crucial in the genotyping of Chlamydia trachomatis when directly performed with clinical samples (J. Lan, J. M. M. Walboomers, R. Roosendaal, G. J. van Doornum, D. M. McLaren, C. J. L. M. Meijer, and A. J. C. van den Brule, J. Clin. Microbiol. 31:1060-1065, 1993). Several primers flanking the four variable domains of the omp1 gene were selected and tested for sensitivity in several nested PCRs with serial dilutions of serovar G. The optimal sensitivity obtained was 0.1 to 0.01 inclusion-forming units, similar to that obtained in the C. trachomatis plasmid PCR. With this approach, any C. trachomatis PCR-positive sample can be typed.
Cross-reactivity of genetic probe for detection of Mycobacterium tuberculosis with newly described species Mycobacterium celatum
Authors: Butler W R , O'Connor S P , Yakrus M A , Gross W M
Published: January 2020

An acridinium ester-labeled DNA probe (AccuProbe; Gen-Probe Inc., San Diego, Calif.) for the identification of the Mycobacterium tuberculosis complex gave discrepant results with the newly described species M. celatum. Examination of 20 strains of M. celatum showed that 8 were positive with the probe; the remaining 12 were negative.
Rapid assay for detection of toxic shock syndrome toxin 1 from human sera
Authors: Miwa K , Fukuyama M , Kunitomo T , Igarashi H
Published: January 2020

A noncompetitive enzyme-linked immunosorbent assay that enables the quantitation of toxic shock syndrome toxin 1 (TSST-1) to as little as 30 pg/ml and the detection of TSST-1 to 10 pg/ml in phosphate-buffered saline including 33% human serum or plasma was developed. It takes only 3 h to complete this assay after plate preparation. In this study, 64 human serum samples obtained from 30 patients with toxic shock syndrome or toxic shock syndrome-like symptoms were subjected to testing for the detection of TSST-1. With a cutoff level for TSST-1 of less than 100 pg/ml, 28 samples obtained from 12 patients were positive for TSST-1. The mean and maximum concentrations for these TSST-1-positive samples were 440 and 5,450 pg/ml, respectively. Of these 12 patients, 8 were Staphylococcus aureus culture positive, 3 were negative upon bacterial culturing, and 1 had no cultures done.
Microflora of abdominal sepsis by locus of infection
Authors: Walker A P , Krepel C J , Gohr C M , Edmiston C E
Published: January 2020

Specimens from 152 abdominal infections were examined to determine which groups of endogenous bacteria participate in infection emanating from different sites in the gastrointestinal tract. A notable finding was the predominance of anaerobic microflora from infections of ischemic versus perforated small bowel. Empiric antibiotic treatment for ischemic bowel should include focused coverage for anaerobes.
Evaluation of a p21e-spiked western blot (immunoblot) in confirming human T-cell lymphotropic virus type I or II infection in volunteer blood donors. The Retrovirus Epidemiology Donor Study Group
Authors: Kleinman S H , Kaplan J E , Khabbaz R F , Calabro M A , Thomson R , Busch M
Published: January 2020

Current algorithms for the serologic confirmation of human T-cell lymphotropic virus type I and II (HTLV-I/II) antibody reactivity are complicated. We evaluated the performance of an HTLV-I Western blot (immunoblot) spiked with recombinant p21e protein (p21e WB) as an alternative to current confirmatory methods. These methods include the HTLV-I viral lysate Western blot and either a radioimmunoprecipitation assay or a p21e enzyme-linked immunosorbent assay. Five hundred fifty nine blood donations obtained from five U.S. blood centers and classified as HTLV-I/II seropositive (n = 149) or seroindeterminate (n = 410) by routine testing methods were further evaluated by PCR for proviral DNA and by the p21e WB. On the basis of serologic and PCR testing, 155 donations were classified as HTLV-I/II infected. The sensitivity of the p21e WB was 97.4%, slightly exceeding that of routine confirmatory testing. The specificity of the p21e WB was 97.5%, as determined by testing of 404 seroindeterminate samples that were negative in the PCR. The positive predictive value of the p21e WB was 94%. In contrast, the specificity and positive predictive value of routine confirmatory testing were both 100%. Follow-up sampling of presumptive p21e WB false-positive donors substantiated the absence of HTLV-I/II infection. Although the p21e WB used in this study has high sensitivity and may be useful as a confirmatory assay in epidemiologic research studies, it may not be ideal as a confirmatory test for the notification of blood donors.