bluuebunny/crossref_metadata_2025 huggingface dataset:
Coagulase-negative staphylococci in multiple blood cultures: strain relatedness and determinants of same-strain bacteremia
Authors: Khatib R , Riederer K M , Clark J A , Khatib S , Briski L E , Wilson F M
Published: January 2020

The frequency of strain relatedness was determined among randomly selected patients with coagulase-negative staphylococcal infections as determined in multiple blood cultures by plasmid typing, determination of species, and antibiotyping. Strain relatedness was demonstrated in 21 of 47 episodes of bacteremia (44.7%) among 34 patients, with a similar percentage among patients with two or one positive blood culture in 24 h (14 of 30 [46.7%] versus 7 of 17 [41.2%], respectively). Same-strain bacteremia was more frequent in cases of infection among patients with a corresponding fever (15 of 21 [71.4%]), among patients infected with organisms from an identifiable source (7 of 9 [77.8%]) and with non-Staphylococcus epidermidis species (9 of 11 [81.8%]), and among patients with nosocomially acquired infections (18 of 36 [50%]). Comparing episodes with or without strain relatedness, no difference was noted in the time to growth (2.1 +/- 1.4 versus 1.9 +/- 0.9 days, respectively), in bacterial growth in two culture bottles (5 of 14 [35.7%] versus 8 of 24 [33.3%], respectively), and in the presence of additional negative blood cultures (9 of 21 [42.9%] versus 11 of 26 [42.3%], respectively). The antibiotypes of all related strains and 7 of 44 (15.9%) unrelated pairs were identical. These findings demonstrate that coagulase-negative staphylococci from multiple blood cultures are frequently unrelated, suggesting a high prevalence of contamination. In the absence of precise measures for demonstrating strain relatedness, the combination of a clinical assessment with antibiotype determination appears to be a suitable alternative.
Food-initiated outbreak of methicillin-resistant Staphylococcus aureus analyzed by pheno- and genotyping
Authors: Kluytmans J , van Leeuwen W , Goessens W , Hollis R , Messer S , Herwaldt L , Bruining H , Heck M , Rost J , van Leeuwen N
Published: January 2020

An outbreak of methicillin-resistant Staphylococcus aureus (MRSA) involving 27 patients and 14 health-care workers (HCW) was studied. The outbreak started in the hematology unit of the University Hospital Rotterdam, Dijkzigt, The Netherlands, and spread to the surgical unit. Twenty-one patients (77.8%) developed clinical disease, and five died. Subsequently, MRSA was detected in food and in the throat of one of the HCW who prepared food for hematology patients. Food contaminated by an HCW most likely caused the first case of MRSA septicemia. This route of transmission has not been described before. The outbreak strain was probably transmitted to the surgical unit by a colonized nurse, where it caused an explosive outbreak. Airborne probably transmitted to the surgical unit by a colonized nurse, where it caused an explosive outbreak. Airborne MRSA transmission played an important role in disseminating the organism. The outbreak was controlled within 6 months by intensifying surveillance, temporarily closing the affected wards, treating carriers, and instituting an MRSA ward outside the hospital. Phage typing, insertion sequence probing, protein A gene typing, and DNA fingerprinting by PCR revealed that all outbreak-related isolates were identical. By pulsed-field gel electrophoresis, all but one of the outbreak-related isolates were determined to be identical. Protein A gene typing identified numerous (11) repeat units in all outbreak-related isolates, which supports the suggestion that the outbreak strain may have been more virulent and more transmissible than other MRSA strains. Pheno- and genotyping studies underlined the value of DNA fingerprinting methods for investigation of MRSA epidemiology. Optimal discriminatory power was achieved by combining the results of four genotyping methods.
Comparison of two rapid commercial tests with complement fixation for serologic diagnosis of Mycoplasma pneumoniae infections
Authors: Thacker W L , Talkington D F
Published: January 2020

The complement fixation (CF) test is the current reference serologic test for the diagnosis of Mycoplasma pneumoniae infection. However, it is reported to be insensitive and nonspecific, and it is labor intensive. To determine if a faster and more sensitive diagnosis of M. pneumoniae could be obtained, we examined 50 paired serum samples from patients with suspected M. pneumoniae infection by the CF test and two commercial rapid antibody detection kits, the Remel M. pneumoniae immunoglobulin G (IgG)-IgM antibody test system (Remel, Lenexa, Kans.) and the Seradyn Color Vue M. pneumoniae IgG-IgM kit (Seradyn, Indianapolis, Ind.). The Remel test, a 5-min qualitative immunobinding assay, detected antibodies in three patient serum samples with CF titers of 32 and in all but one sample with titers of > or = 64. The Seradyn test, a 40-min qualitative agglutination test, was less sensitive than CF or Remel. The Seradyn test was positive in 68% of cases, compared with 94 and 96% of cases tested by CF or Remel, respectively. Both commercial tests are faster and less technically demanding to perform than is the CF test.
Potential sources of sporadic human infection with Yersinia enterocolitica serovar O:8 in Aomori Prefecture, Japan
Authors: Hayashidani H , Ohtomo Y , Toyokawa Y , Saito M , Kaneko K , Kosuge J , Kato M , Ogawa M , Kapperud G
Published: January 2020

In November 1992 and June and August 1993 rectal contents from 204 small mammals living in the wild were collected and examined for the presence of Yersinia enterocolitica serovar O:8 to clarify the source of human infections caused by this microbe in the Tsugaru Region of Aomori Prefecture, Japan. Serovar O:8 was isolated from 10 (5.2%) of 193 wild rodents trapped in June 1993 (9 of 107) and August 1993 (1 of 23) but not from animals trapped in November 1992 (0 of 63). This serovar was not isolated from 11 moles. From May to September 1993, 12 human patients were found to have become ill and to be infected with Y. enterocolitica O:8. The patients lived in the same districts where the wild rodents harboring serovar O:8 were trapped. Two different patterns by restriction enzyme analysis of the virulence plasmid were observed. One pattern obtained by restriction enzyme analysis of the virulence plasmid was observed in 20 isolates from 11 human patients and 9 wild rodents, and the other was observed in 2 isolates from 1 human patient and 1 wild rodent. These findings indicate that wild rodents seem to play an important role as a source of human Y. enterocolitica O:8 infection.
Detection of Salmonella typhimurium from rectal swabs of experimentally infected beagles by short cultivation and PCR-hybridization
Authors: Stone G G , Oberst R D , Hays M P , McVey S , Galland J C , Curtiss R , Kelly S M , Chengappa M M
Published: January 2020

A rapid and sensitive cultivation and PCR-hybridization procedure for the detection and identification of Salmonella typhimurium was evaluated over a 42-day period with eight experimentally infected beagles. Rectal swabs were taken at several times postinfection, inoculated into selenite-cystine broth, and plated onto Hektoen-Enteric Enteric agar immediately after incubation for 4 and 24 h. PCRs and hybridizations were also conducted with each sample, and the results were compared with those of standard culture techniques to evaluate the efficiency of the PCR-hybridization procedure. The PCR-hybridization procedure was more sensitive than standard culture techniques at each enrichment incubation (P < 0.05). In addition, the PCR-hybridization procedure was significantly better than culture up through 3 days postinfection (P < 0.05). A nonspecific amplified product, relatively close in size to the 457-bp specifically amplified product, did not hybridize to an internal oligonucleotide probe or to a random-primed labeled probe. Subsequent sequence information revealed that the product had very little similarity to the 457-bp product but had significant similarity to an Escherichia coli aldehyde dehydrogenase gene. This study indicated that a cultivation and PCR-hybridization procedure is significantly better than culture for the identification of S. typhimurium. Additionally, the results confirm the importance of determining specificities of PCR products beyond the gel electrophoresis level by hybridization with a specific probe.
Detection of Tropheryma whippelii DNA in a patient with AIDS
Authors: Maiwald M , Meier-Willersen H J , Hartmann M , von Herbay A
Published: January 2020

A case of an AIDS patient infected with the Whipple's disease bacterium, Tropheryma whippelii, is reported. A DNA fragment with sequence specificity for the 16S rRNA gene of the bacterium was detected by PCR in a duodenal biopsy specimen from a 55-year-old male patient with AIDS and diarrhea. The biopsy specimen contained periodic acid-Schiff stain-positive macrophages which did not, however, resemble the sickleform-particle-containing cells characteristic of Whipple's disease. This observation raises two possibilities: either the patient had a coincidence of AIDS and Whipple's disease or Tropheryma whippelii acted as an opportunistic pathogen in this immunodeficient patient. The latter explanation is of interest in light of the ongoing discussion of immunologic abnormalities as predisposing factors for Whipple's disease.
Taxonomic position of deep-seated, mucosa-associated, and superficial isolates of Trichosporon cutaneum from trichosporonosis patients
Authors: Sugita T , Nishikawa A , Shinoda T , Kume H
Published: January 2020

Clinical isolates of Trichosporon cutaneum, the causative agent of trichosporonosis, were identified on the basis of DNA relatedness. Of the 10 strains from deep-seated and mucosa-associated infections, 9 were identified as T. asahii and 1 was identified as T. ovoides. The two superficial strains were identified as T. cutaneum and a variety of T. montevideense. These findings suggest that T. cutaneum is a heterogeneous species in clinical samples and that the causative agents of trichosporonosis exist in four or more species.
Value of terminal subcultures for blood cultures monitored by BACTEC 9240
Authors: Shigei J T , Shimabukuro J A , Pezzlo M T , de la Maza L M , Peterson E M
Published: January 2020

Blood cultures collected in BACTEC Plus Aerobic/F bottles and BACTEC Plus Anaerobic/F bottles were monitored for 5 days by BACTEC 9240 and subsequent terminal subcultures. Of the 13,471 bottles subcultured, 11.0% (1,477 of 13,471) were culture positive. Of these, 94.0% (1,388 of 1,477) were detected by BACTEC 9240; the additional 6.0% (89 of 1,477) were considered to be false negatives by BACTEC 9240 since they were detected by terminal subculture only. The false-negative bottles consisted of 17 BACTEC Plus Aerobic/F and 72 BACTEC Plus Anaerobic/F bottles, accounting for 2.2 (17 of 786) and 10.4% (72 of 691) of the total positive aerobic and anaerobic bottles, respectively. The positive blood culture bottles most frequently not detected by BACTEC 9240 grew Pseudomonas spp. (24), Staphylococcus spp. (21), and yeasts (24). Of the 86 blood cultures represented by the 89 false-negative bottles, 41 would not have been identified as positive since the other bottle in the blood culture set was either a false negative or a true negative. In general, terminal subcultures of false-negative BACTEC bottles had heavy growth, indicating that BACTEC Plus media were able to support the growth of microorganisms, but the BACTEC 9240 instrument was unable to detect this growth.
Genetic diversity of Pseudomonas aeruginosa strains isolated from patients with cystic fibrosis revealed by restriction fragment length polymorphism of the rRNA gene region
Authors: Martin C , Ichou M A , Massicot P , Goudeau A , Quentin R
Published: January 2020

The restriction fragment length polymorphism patterns of rDNAs from Pseudomonas aeruginosa strains isolated from the respiratory tracts of patients suffering from cystic fibrosis were obtained to evaluate the genetic polymorphism of this population of strains. Eighty-seven P. aeruginosa strains isolated from 87 patients from diverse areas of France and the ATCC 10145 strain were examined. Four restriction enzymes were used: BamHI, ClaI, EcoRI, and PstI. Forty-nine strains (56%) were in the three most frequent ribotypes (ribotypes R1 to R3). In addition, hierarchical clustering analysis of the data showed that 71 of the 88 strains (81%) clustered at a distance of less than one-third of the greatest distance observed in the total population. This indicates that clinical isolates implicated in the pathology of cystic fibrosis present a low degree of heterogeneity of rRNA operons, in contrast to the heterogeneity of strains of P. aeruginosa isolated from patients with various other pathologies. This relative homogeneity of rRNA genes was observed independently of the clinical status of the patient and the colony morphology.
Multiple strains of Streptococcus pyogenes in skin sores of aboriginal Australians
Authors: Carapetis J , Gardiner D , Currie B , Mathews J D
Published: January 2020

A molecular technique (random amplification of polymorphic DNA) was used to characterize group A streptococcal (GAS) strains among 194 isolates from 55 swabs from 12 Australian Aboriginal children and adults with multiple pyoderma lesions. Ninety-three percent of the lesions contained only one strain of GAS, but 8 of 12 individuals were infected with more than one strain. We conclude that accurate epidemiologic surveys require that more than one swab specimen be obtained from each person, whereas typing of more than one colony per swab is less informative. Characterization of GAS strains by random amplification of polymorphic DNA analysis should help to provide important insights into the epidemiology of GAS, particularly in tropical populations where many isolates are M nontypeable, and into the mechanisms of genetic variation of GAS in such populations.
Variable serum immunoglobulin responses against different Borrelia burgdorferi sensu lato species in a population at risk for and patients with Lyme disease
Authors: Bunikis J , Olsén B , Westman G , Bergstroöm S
Published: January 2020

Borrelia burgdorferi sensu lato species display considerable antigenic polymorphism. In order to evaluate the importance of this antigenic heterogeneity in the serodiagnosis of Lyme disease, the serum immunoglobulin G response in 148 healthy individuals from an area in northern Sweden where Lyme disease is endemic and in 40 American patients with Lyme disease was assessed. In a seroprevalence study, the control group included 173 individuals from a region of northern Sweden where Lyme disease is not endemic. The two enzyme immunoassays used were based on outer membrane-associated proteins of either B. burgdorferi sensu stricto or Borrelia garinii. The Swedish populations were also screened for antiflagellum seroreactivity. The individuals from the area of endemicity were significantly more seropositive for the subcellular protein fraction of the local B. garinii isolate NBS16 than the control group (11.5 versus 2.9%; P = 0.005) but were not significantly more positive for the other antigens used. In contrast, American patients with Lyme disease were significantly more reactive against the North American B. burgdorferi sensu stricto strain B31 than against B. garinii NBS16 (57.5 versus 15.0%; P = 0.0001). Immunoblot analysis suggests that the borrelial outer surface protein C is involved in triggering the production of species-specific antibody during localized Lyme disease. We conclude that a species-specific immune response develops during infection with Lyme disease Borrelia spp. Thus, the reliability of a serological investigation of Lyme disease increases when one measures antibody titers against the outer membrane proteins of Lyme disease Borrelia spp. occurring in a particular geographic region.
Ethanol fixation of sputum sediments for DNA-based detection of Mycobacterium tuberculosis
Authors: Williams D L , Gillis T P , Dupree W G
Published: January 2020

The effect of ethanol fixation on PCR detection and viability of Mycobacterium tuberculosis in human sputum sediments was evaluated. M. tuberculosis seeded into sputum sediments was efficiently killed when treated for 1 h with 50, 70, or 95% ethanol. PCR amplification of a 123-bp fragment of the M. tuberculosis-specific IS6110 was not affected in ethanol-treated samples even when fixation was extended to 24 h. Ethanol fixation of sputum sediments did not affect the PCR detection of M. tuberculosis in clinical samples. PCR results from ethanol-treated clinical samples containing M. tuberculosis (smear positive and smear negative) or other respiratory pathogens correlated directly with the results by conventional detection methods for M. tuberculosis. Our results show that ethanol fixation of human sputum sediments containing M. tuberculosis significantly reduces the potential exposure of workers to viable M. tuberculosis without affecting DNA analysis by PCR. Also, ethanol fixation of sputum sediments provides a simple and inexpensive way to store and transport clinical specimens identified for DNA-based diagnostics without refrigeration.
Analysis of enzymatic activities for differentiation of two species of nutritionally variant streptococci, Streptococcus defectivus and Streptococcus adjacens
Authors: Beighton D , Homer K A , Bouvet A , Storey A R
Published: January 2020

Strains of nutritionally variant streptococci, Streptococcus defectivus (n = 10) and Streptococcus adjacens (n = 20), were studied for the production of glycosidic and proteolytic enzyme activities. S. defectivus strains produced neuraminidase and alpha-fucosidase, while S. adjacens strains produced only neuraminidase. The S. adjacens strains produced a very wide range of proteolytic activities with the ability to hydrolyze the majority of aminopeptidase substrates tested, while S. defectivus strains hydrolyzed only a minority of aminopeptidase substrates. These data provide additional phenotypic characteristics which may be useful in distinguishing between these two species and suggest that they may have different nutritional requirements.
Species identification of members of the Streptococcus milleri group isolated from the vagina by ID 32 Strep system and differential phenotypic characteristics
Authors: Ahmet Z , Warren M , Houang E T
Published: January 2020

The importance of bacterial vaginosis as a risk factor in obstetric and gynecological infections has recently been recognized. The bacterial vaginosis group of organisms includes members of the Streptococcus milleri group, the identification of which has caused much confusion. We prospectively surveyed the rates of carriage of S. milleri group organisms in 397 high vaginal swabs received in our laboratory. For the identification of 99 clinical isolates and 23 control strains, we compared the results obtained by the rapid ID 32 Strep system (Analytab Products) and by a scheme utilizing six differential phenotypic characteristics (presence of beta-N-acetylglucosaminidase, alpha-glucosidase, beta-D-fucosidase, beta-galactosidase, beta-N-acetylgalactosaminidase, and beta-glucosidase) as described by Whiley et al. (R. A. Whiley, H. Fraser, J. M. Hardie, and D. Beighton, J. Clin. Microbiol. 28:1497-1501, 1990). We identified Streptococcus anginosus in 18% and Streptococcus constellatus in 0.05% of the specimens examined. Of the isolates of S. anginosus that reacted with grouping antisera, 20 of 25 belonged to Lancefield group F. The incubation conditions for bacterial cultures and for reaction mixtures affected the results of phenotypic characterization in the production of alpha-glucosidase, beta-galactosidase, and beta-glucosidase. However, by using bacterial cultures grown under hypercapnic conditions and incubating the reaction mixtures aerobically, consistent phenotypic characteristics were obtained, allowing identification similar to that obtained by the ID 32 Strep system. We therefore recommend the phenotypic scheme as an inexpensive, reliable, and convenient method for the initial identification of species of the S. milleri group.
Experience with a novel selective medium for isolation of Actinomyces spp. from medical and dental specimens
Authors: Lewis R , McKenzie D , Bagg J , Dickie A
Published: January 2020

A selective medium containing mupirocin and metronidazole was evaluated for the isolation of Actinomyces spp. from clinical material. The study was a one-year prospective comparison of the method with existing methods, which use nonselective media, at a general hospital and a dental hospital. Significantly more Actinomyces spp. were isolated on the selective medium than on nonselective media from both dental specimens and intrauterine contraceptive devices. However, differentiating between Actinomyces spp. and related nonsporulating gram-positive rods remains a slow and sometimes uncertain process which can introduce lengthy delays in reporting.
Comparison of rapid immunofluorescence procedure with TestPack RSV and Directigen FLU-A for diagnosis of respiratory syncytial virus and influenza A virus
Authors: Todd S J , Minnich L , Waner J L
Published: January 2020

A rapid immunofluorescence format requiring 20 min for completion was as effective as conventional indirect and direct immunofluorescence procedures for detecting respiratory syncytial virus and influenza A virus antigens in clinical specimens. Rapid immunofluorescence was more sensitive than TestPack RSV and comparable to Directigen FLU-A immunosorbent assays, which require 20 min for completion.
Human herpesvirus 6 and human herpesvirus 7 in chronic fatigue syndrome
Authors: Di Luca D , Zorzenon M , Mirandola P , Colle R , Botta G A , Cassai E
Published: January 2020

We analyzed lymphocytes of patients with chronic fatigue syndrome (CFS) for the presence of human herpesvirus 6 (HHV-6) and HHV-7 DNA. HHV-7 was present in over 80% of CFS patients and healthy controls, while the prevalence of HHV-6 variant A increased significantly in CFS cases (22 versus 4%; P = 0.05).
Analysis of strains of Campylobacter fetus by pulsed-field gel electrophoresis
Authors: Fujita M , Fujimoto S , Morooka T , Amako K
Published: January 2020

Campylobacter fetus chromosomal DNA from 21 strains was analyzed by pulsed-field gel electrophoresis. The fingerprint patterns generated with SmaI and SalI were distinctive. Using the profiles obtained by pulsed-field gel electrophoresis, we established the phylogenetic dendrogram of C. fetus to identify the genetic relationship of the strains.
Variation of human papillomavirus type 6 (HPV-6) and HPV-11 genomes sampled throughout the world
Authors: Heinzel P A , Chan S Y , Ho L , O'Connor M , Balaram P , Campo M S , Fujinaga K , Kiviat N , Kuypers J , Pfister H
Published: January 2020

We examined the genomic diversity of human papillomavirus type 6 (HPV-6) and HPV-11 isolates from different parts of the world by comparing the nucleotide sequences of part of the long control region of three reference clones and 62 HPV-6 and 40 HPV-11 isolates from Africa, Europe, Asia, and North and South America. The genomic sequence of the HPV-6b reference type had to be amended by inclusion of a 94-bp segment, which is also present with minor differences in HPV-6a. Aside from two small inserts typical of all variants related to HPV-6a and three inserts found in HPV-11 variants, no major alterations to the size of the long control regions of these viruses were observed. This corrects the previous impression that these two HPV types are highly polymorphic. Altogether, 19 HPV-6 and 10 HPV-11 variant genomes could be distinguished, and most of the differences were due to point substitutions. The variants of either type were continuously connected in phylogenetic trees rather than clustered separately into subtype groups. Thirteen mutations, namely, the two HPV-6a inserts and 11 substitutions in HPV-6 or HPV-11 variants, reduced the dissimilarity between the types, but they bridged only a small fraction of the genetic distance between the two types. Genomes more obviously intermediate between HPV-6 and HPV-11 were not found and probably do not exist any more. A single HPV-11 variant was found in Africa, but otherwise, no significant correlations of HPV-6 or HPV-11 variants with geography or ethnicity of the patient cohort were found. Functional analysis of diverse enhancer variants showed activities that differed two-to threefold, and it must be considered that transcriptional differences may alter the biology or pathology of these viruses. Similar variants were found in lesions from anatomically different sites and in both benign and malignant lesions.
Detection and quantification of virus DNA in plasma of patients with Epstein-Barr virus-associated diseases
Authors: Yamamoto M , Kimura H , Hironaka T , Hirai K , Hasegawa S , Kuzushima K , Shibata M , Morishima T
Published: January 2020

Epstein-Barr virus (EBV) causes various diseases, such as infectious mononucleosis (IM), fatal IM, EBV-associated hemophagocytic syndrome (EBVAHS), and chronic active EBV infection (CAEBV). In the present study, cell-free EBV DNA was detected in the plasma of patients with EBV-associated diseases by PCR assay. The patients included 20 patients with IM, 2 patients with fatal IM, 4 patients with EBVAHS, 4 patients with CAEBV, and 38 healthy children (20 EBV seropositive and 18 EBV seronegative). In patients with IM, plasma samples were positive for EBV DNA in all patients (100%) in the acute phase and in 44% of the patients in the convalescent phase, but plasma samples from the 38 healthy control children were negative (0%) for EBV DNA. Quantitative PCR assay revealed that plasma from patients with IM contained the highest amount of virus DNA within 7 days following the onset of disease (mean, 6 x 10(4) copies per ml). The EBV DNA concentration decreased thereafter as the patients recovered. Plasma from patients with fatal IM contained more than 100 times more copies of EBV DNA (3 x 10(7) copies per ml) than plasma from patients with IM. Plasma from patients with the acute phase of EBVAHS contained 10 times more copies of EBV DNA (5 x 10(5) copies per ml) than plasma from IM, and then patients with the number of copies decreased similarly in both groups of patients in the convalescent phase (2 x 10(4) copies per ml). The amount of virus DNA in patients with CAEBV (6 x 10(4) copies per ml) was similar to that noted in patients with IM; however, it became higher (1 x 10(6) copies per ml) when the patients' clinical status deteriorated. These data suggest that the presence of cell-free EBV DNA in plasma is a common phenomenon in patients with EBV-associated diseases. The concentration of EBV DNA in plasma seems to be higher in patients with the more severe clinical categories of EBV diseases.
Molecular analysis of genes encoding outer surface protein C (OspC) of Borrelia burgdorferi sensu lato: relationship to ospA genotype and evidence of lateral gene exchange of ospC
Authors: Jauris-Heipke S , Liegl G , Preac-Mursic V , Rössler D , Schwab E , Soutschek E , Will G , Wilske B
Published: January 2020

It has been shown by analysis with monoclonal and polyclonal antibodies that outer surface protein C (OspC) of Borrelia burgdorferi sensu lato is highly heterogeneous. To determine if the heterogeneity has a genetic basis, the genes of 18 different B. burgdorferi sensu lato strains have been amplified by PCR, cloned, and sequenced. The ospC genes could be amplified from all strains tested, even from two strains which did not express OspC in detectable amounts. Among the 18 strains, 16 significantly different types of ospC sequences have been found. The sequence identities of the deduced amino acid sequences of different ospC genotypes range between 62 and 80% (determined without the leader peptide). The sequences range between 62 and 80% (determined without the leader peptide). The sequences correspond to one of the 13 OspC types distinguishable by analysis with monoclonal antibodies (B. Wilske, S. Jauris-Heipke, R. Lobentanzer, I. Pradel, V. Preac-Mursic, D. Roessler, E. Soutschek, and R. C. Johnson, J. Clin. Microbiol. 33:103-109, 1995) or represent additional types. Two completely new types were found, and OspC type 8 (which was found in Borrelia afzelii and Borrelia garinii) could be divided into two groups with different sequences but the same antibody pattern. Thus, strains belonging to different species or OspA serotypes were always significantly different in their ospC sequences. This was also confirmed by ospA sequence analysis. Interestingly, some strains of the same OspA serotype or genotype were very heterogeneous with respect to OspC, while others had nearly identical OspC proteins. Such groups of strains were found among B. burgdorferi sensu stricto, B. afzelii, and B. garinii strains. Cluster analysis of 5'-terminal and 3'-terminal stretches of ospC suggested recent intragenic recombination events in the ospC gene at least one B. afzelii strain. In addition, other recombination events between ancestors of strains belonging to the same or different species were evidenced by this type of analysis.
Comparison of BACTEC 9240 and BacT/Alert blood culture systems in an adult hospital
Authors: Smith J A , Bryce E A , Ngui-Yen J H , Roberts F J
Published: January 2020

In a study comparing the BACTEC 9240 (Becton Dickinson, Canada Inc., Mississauga, Ontario, Canada) and the BacT/Alert (Organon Teknika, Scarborough, Ontario, Canada) blood culture systems, 6,437 sets of four bottles of 10,412 cultures submitted were deemed acceptable for evaluation. There were 586 clinically significant isolates from 544 positive blood cultures. Of the 544 positive blood cultures, 329 were positive in both systems, 157 were positive in the BACTEC 9240 system alone, and 58 were positive in the BacT/Alert system alone (P < 0.001). These cultures represented 421 bacteremic episodes, of which 290 were detected by both systems, 84 were detected by the BACTEC 9240 system alone, and 47 were detected by the BacT/Alert system alone (P < 0.001). The difference between the two systems in terms of microbial recovery was attributable largely to differences in the ability to grow staphylococci. Of 82 monomicrobial isolates of Staphylococcus aureus, 46 were isolated by both systems, 30 were isolated by the BACTEC 9240 system alone, and 6 were isolated by the BacT/Alert system alone (P < 0.0001). A similar pattern was noted in the case of coagulase-negative staphylococci, of which 48 were isolated in both systems, 46 were isolated in the BACTEC 9240 system alone, and 5 were isolated in the BacT/Alert system alone (P < 0.0001). The false-positive rate in the BACTEC 9240 system was 0.5%, and that in the BacT/Alert system was 1.2%. Both systems provided satisfactory service mechanically, and in terms of computer software and costs, both systems were similar. The differences in the recovery of organisms, notably staphylococci, may be due in part to the fact that in the present study the BacT/Alert medium did not contain antimicrobial agent-removing devices, whereas the BACTEC 9240 medium did. Until this difference is resolved, we consider the BACTEC 9240 system to more adequately meet our hospital's requirements.
The DNA-binding protein pUL57 of human cytomegalovirus is a major target antigen for the immunoglobulin M antibody response during acute infection
Authors: Vornhagen R , Hinderer W , Sonneborn H H , Bein G , Matter L , The T H , Jahn G , Plachter B
Published: January 2020

A small polypeptide from pUL57 of human cytomegalovirus was identified as a major target for the immunoglobulin M antibody response. This antigen seems to be superior to antigenic fragments from pp150 and p52 in the identification of sera from acutely infected patients. It may therefore represent an essential antigen for recombinant immunoglobulin M antibody tests for human cytomegalovirus.
Comparison of pigment production and motility tests with PCR for reliable identification of intrinsically vancomycin-resistant enterococci
Authors: Cartwright C P , Stock F , Fahle G A , Gill V J
Published: January 2020

Forty-eight clinical isolates identified as either Enterococcus faecium or Enterococcus faecalis with the MicroScan system (Dade International, MicroScan Inc., West Sacramento, Calif.) were further characterized by two supplementary biochemical tests (pigment production and motility). Twenty isolates (42%), all initially identified as E. faecium, were motile. Of these 20, 8 isolates (17%) produced yellow pigment and were identified as Enterococcus casseliflavus and the remaining 12 (25%) were nonpigmented and were identified as Enterococcus gallinarum. Identical identification results were obtained when PCR amplification of regions of the vanC gene was used as a technique for differentiating these organisms. The results of this study indicate that motility and pigment production tests together with commercial test systems are sufficient for reliable identifications of E. faecium, E. casseliflavus, and E. gallinarum.
Algorithm for use of nucleic acid probes for identifying Mycobacterium tuberculosis from BACTEC 12B bottles
Authors: Metchock B , Diem L
Published: January 2020

Nucleic acid probes (Gen-Probe, San Diego, Calif.) can be used to identify mycobacteria in BACTEC 12B broth cultures prior to detection of growth on solid media. We developed an algorithm that can be used to make an initial choice of a probe (either Mycobacterium tuberculosis complex [MTB] or M. avium complex [MAC]) for use in testing respiratory specimens. The algorithm was based on both the fluorochrome smear result of the concentrated specimen and the time from inoculation until the BACTEC 12B broth culture is flagged (growth index 10) as presumptively positive. The MTB probe is used first for all 4+ smear specimens, 3+ smear specimens positive in 5 days, 2+ and 1+ smear specimens positive in 7 days, and smear-negative specimens positive in 11 days. The MAC probe is used for all other specimens. The algorithm is used when other information about the culture (e.g., previous positive cultures and colonial morphology of growth on solid media) is unknown. Use of the algorithm to probe 102 respiratory BACTEC 12B broth cultures (35 with MTB; 1 with MTB, MAC, and M. gordonae; 47 with MAC; and 19 with other mycobacterial species) from 1 September through 30 November 1992 resulted in the initial use of the MTB probe for 35 (97%) of the cultures positive for MTB and the use of the MAC probe for 35 (73%) of the cultures positive for MAC. Use of the algorithm aided in the efficient use of laboratory resources without delaying the time to identification of MTB isolates.
Retrospective case-control study of diffusely adhering Escherichia coli and clinical features in children with diarrhea
Authors: Poitrineau P , Forestier C , Meyer M , Jallat C , Rich C , Malpuech G , De Champs C
Published: January 2020

A retrospective case-control study with a small population group revealed that, among clinical signs, vomiting but not diarrhea was significantly associated with the presence of diffusely adhering Escherichia coli (DAEC) in children suffering from gastroenteritidis (P < 0.05). Of the children carrying DAEC strains, those who were F1845 DNA probe positive had a significantly longer hospital stay than those who were F1845 DNA probe negative. We believe that the heterogeneity of DAEC strains is responsible for the discrepant results concerning their involvement in disease and that only some of these strains are really pathogenic for children.
Errors arising from incorrect orientation of E test strips
Authors: Hamilton-Miller J M , Shah S , Yam T S
Published: January 2020

Major errors arise if E test strips are placed upside down. Asymmetric zones, or no zone at all, may result. MICs indicated by upside-down tests were almost always considerably higher than true values. This situation differs markedly from that for conventional testing, where orientation of disks is not important.
Expression of Sendai virus nucleocapsid protein in a baculovirus expression system and application to diagnostic assays for Sendai virus infection
Authors: Wan C H , Riley M I , Hook R R , Franklin C L , Besch-Williford C L , Riley L K
Published: January 2020

The most common diagnostic technique for the detection of Sendai virus infection in rodents is serological evaluation by enzyme-linked immunosorbent assay (ELISA) with semipurified preparations of whole virions as antigens. This assay often suffers from a lack of specificity. The goal of the present project was to develop more specific antigens for use in diagnostic testing by producing recombinant antigens in insect cells. To identify viral proteins immunoreactive in multiple laboratory rodent species, Western blots (immunoblots) of viral polypeptides were probed with immune sera from mice, rats, and hamsters. The nucleocapsid protein (NP) reacted with immune sera from all species tested. Therefore, the NP gene was selected for cloning and expression in a baculovirus. To construct the recombinant, complementary DNA was synthesized by reverse transcription PCR from Sendai virus RNA with primers from the 5' and 3' termini of the NP-coding region. Amplified DNA was cloned into a baculovirus transfer vector (pBlueBacHis A) and was cotransfected with wild-type baculovirus into insect cells. Baculovirus recombinants containing the NP gene were identified by PCR. Evaluation of the recombinant proteins expressed in insect cells by Western blot analysis revealed specific reactivity with immune sera. In comparison with conventional ELISAs that use whole virions as the antigen, ELISAs that use recombinant NP were more specific.
Investigation of sheep-associated malignant catarrhal fever virus infection in ruminants by PCR and competitive inhibition enzyme-linked immunosorbent assay
Authors: Li H , Shen D T , O'Toole D , Knowles D P , Gorham J R , Crawford T B
Published: January 2020

Development of control measures for the gammaherpesviral disease of cattle known as sheep-associated malignant catarrhal fever (SA-MCF) has been hampered by a lack of accurate diagnostic tests either for the causative virus or for antibody against that virus. A recently developed competitive-inhibition enzyme-linked immunosorbent assay (CI-ELISA) for the detection of antibody to malignant catarrhal fever (MCF) virus (MCFV) in ruminants based on a monoclonal antibody to a widely conserved epitope of MCFV (H. Li, D. T. Shen, D. P. Knowles, J. R. Gorham, and T. B. Crawford, J. Clin. Microbiol. 32:1674-1679, 1994) and a PCR assay based on previously reported primers (S. I. F. Baxter, I. Pow, A. Bridgen, and H. W. Reid, Arch. Virol. 132:145-159, 1993) were used to detect anti-MCFV antibody and SA-MCFV DNA in sheep and other ruminants. The PCR amplified a specific 238-bp SA-MCFV genomic DNA fragment from peripheral blood lymphocytes of adult sheep and other ruminants with clinical MCF. Of 144 samples from randomly selected healthy adult sheep, 143 (99%) were positive by PCR and 136 (94%) were positive by CI-ELISA. The agreement between the two assays exceeded 95%. Of nine samples collected from cattle and deer with clinical MCF of apparent sheep origin, seven were CI-ELISA positive and all 9 were PCR positive. Among 59 serum samples from presuckling lambs, none contained antibody detectable by CI-ELISA. After suckling, maternal anti-MCFV antibody was detectable for about 10 +/- 3 weeks. Although all colostrum and milk samples from infected ewes were strongly PCR positive, the appearance of detectable SA-MCFV DNA in lambs was correlated generally with antibody patterns, which suggests that the natural infection event in sheep may not occur during the perinatal period but occurs sometime later in life.
Use of a 24-kilodalton Trypanosoma cruzi recombinant protein to monitor cure of human Chagas' disease
Authors: Krautz G M , Galvão L M , Cançado J R , Guevara-Espinoza A , Ouaissi A , Krettli A U
Published: January 2020

A 24-kDa recombinant protein from Trypanosoma cruzi (rTc24) was evaluated by enzyme-linked immunosorbent assay (ELISA) and Western blot (immunoblot) tests to identify treated chagasic patients considered parasitologically cured on the basis of persistently negative tests of hemocultures and lytic antibodies. Some of these patients were termed dissociated because their sera, although negative by the complement-mediated lysis test, were positive by conventional serology. The negative lysis test indicates the absence of active infection after specific treatment, but this assay requires live and infectious parasites and cannot be used easily in a laboratory routine. Here we tested rTc24 by ELISA and Western blotting as an alternative for the complement-mediated lysis test. For the group of patients with active infection despite the treatment (uncured patients), all the sera tested recognized rTc24 in both tests. For the dissociated patients, approximately 80% of the sera did not react with rTc24 in the ELISA or in Western blots, in agreement with the negative complement-mediated lysis tests. Thus, the 24-kDa T. cruzi recombinant antigen, when used for initial trials to evaluate cure of chagasic patients submitted to specific treatment, will allow the identification of most, but not all, cases.
Quantitative PCR for human herpesviruses 6 and 7
Authors: Secchiero P , Zella D , Crowley R W , Gallo R C , Lusso P
Published: January 2020

A quantitative PCR assay for the detection of human herpesvirus 6 (HHV-6) (variants A and B) and HHV-7 DNAs in clinical samples was developed. The assay uses a nonhomologous internal standard (IS) for each virus that is coamplified with the wild-type target sequence in the same vial and with the same pair of primers. This method allows for a correction of the variability of efficiency of the PCR technique. A standard curve is constructed for each experiment by coamplification of known quantities of the cloned HHV-6 or HHV-7 target templates with the respective IS. Absolute quantitation of the test samples is then achieved by determining the viral target/IS ratio of the hybridization signals of the amplification products and plotting this value against the standard curve. Using this assay, we quantitated the amount of HHV-6 or HHV-7 DNA in infected cell cultures and demonstrated an inhibitory effect of phosphonoformic acid on the replication of HHV-6 and HHV-7 in vitro. As the first clinical application of this procedure, we performed preliminary measurements of the loads of HHV-6 and HHV-7 in lymph nodes from patients with Hodgkin's disease and AIDS. Application of this quantitative PCR method should be helpful for elucidating the pathogenic roles of HHV-6 and HHV-7.
Molecular epidemiology of Escherichia coli O157:H7 by restriction fragment length polymorphism using Shiga-like toxin genes
Authors: Samadpour M
Published: January 2020

The purpose of this study was to assess a simplified method for interstrain differentiation of Escherichia coli O157:H7 and other Shiga-like toxin-producing E. coli (SLTEC) strains. A method based on the use of nucleic acid probes from Shiga-like toxin (SLT) I and II structural genes was used to generate restriction fragment length polymorphism (RFLP) patterns of SLTEC strains, (SLT-RFLP patterns) resulting from digestion of isolated genomic DNA with four different restriction enzymes (BamHI, EcoRI, HindIII, and PvuII) used separately. A total of 165 SLTEC strains from clinical, food, and environmental sources, including O157:H7 isolates from four food-borne outbreaks in Canada and the United States, were analyzed in the study. SLT-RFLP demonstrated that E. coli O157:H7 strains from each food-borne outbreak had the same unique SLT-RFLP pattern. Fifty-two SLT-RFLP types were found among 96 E. coli O157:H7 isolates from sporadic cases of hemorrhagic colitis and hemolytic uremic syndrome in Washington state. The use of the SLT probes proved to be a very powerful method for interstrain differentiation of SLTEC strains. Although the use of each of the enzymes alone did not give enough differentiative power to be used in epidemiological studies, the combination of patterns generated by two restriction enzymes (EcoRI and PvuII, used separately) provided the desired sensitivity for such studies. The results clearly demonstrate the usefulness of the method for studying the molecular epidemiology of E. coli O157:H7. The method is also suitable for establishing an epidemiological database, in terms of both sensitivity and ease of compilation and interpretation of results.
Tear fluid of hepatitis C virus carriers could be infectious
Authors: Feucht H H , Zöllner B , Schröter M , Altrogge H , Laufs R
Published: January 2020

For up to 20 to 40% of patients chronically infected with hepatitis C virus (HCV), the mode of transmission is still unknown. We demonstrate that tear fluid contains HCV RNA-carrying material with the properties of infectious virus and conclude that smear infection with tear fluid may play a role in HCV transmission.
Symptomatic Shigella sonnei urinary tract infection
Authors: Papasian C J , Enna-Kifer S , Garrison B
Published: January 2020

The clinical course for a patient with symptomatic urinary tract infection due to Shigella sonnei is described. The role of Shigella spp. as urinary pathogens is reviewed.
Detection of microsporidia (Enterocytozoon bieneusi) in intestinal biopsy specimens from human immunodeficiency virus-infected patients by PCR
Authors: Franzen C , Müller A , Hegener P , Salzberger B , Hartmann P , Fätkenheuer G , Diehl V , Schrappe M
Published: January 2020

Intestinal microsporidiosis has been implicated as a major cause of chronic diarrhea in human immunodeficiency virus (HIV)-infected patients. So far diagnosis depends on direct visualization of the parasites by light and transmission electron microscopy. We evaluated the diagnostic value of microsporidian DNA amplification by PCR on duodenal biopsy specimens obtained from patients with and without intestinal microsporidiosis caused by Enterocytozoon bieneusi. Thirteen HIV-infected patients (all CDC stage C3) were studied. Eight patients had intestinal microsporidiosis caused by E. bieneusi (n = 6), Septata intestinalis (n = 1), and Encephalitozoon cuniculi (n = 1); microsporidioses were diagnosed by light microscopy of stool samples and confirmed by light and electron microscopy of intestinal biopsy specimens. Five patients had no microsporidia in their stool samples or in their intestinal biopsy specimens, as examined by light and electron microscopy. Additionally, DNA prepared from Toxoplasma gondii derived from mouse ascites was used as a further control. A 353-bp DNA fragment of the small-subunit rRNA gene could be amplified from all six biopsy specimens infected with E. bieneusi, and the nature of the PCR products was confirmed by Southern blot hybridization. No amplification of DNA fragments was seen by using DNA extracted from biopsy specimens with S. intestinalis or E. cuniculi infection or without microsporidian infection and with template DNA extracted from T. gondii. The results suggest that PCR testing of intestinal biopsy specimens may be a useful approach to diagnosing microsporidiosis in HIV-infected patients.