bluuebunny/crossref_metadata_2025 huggingface dataset:
Immunoelectron microscopy for rapid diagnosis of varicella-zoster virus in a complicated case of human T-cell lymphotropic virus type 1 infection
Authors: Folkers E , Vreeswijk J , Wagenaar F , Kapsenberg J G , Hulsebosch H J , Oranje A P
Published: January 2020

Rapid techniques for the detection of herpes simplex virus (HSV) and varicella-zoster virus (VZV) are needed for optimal therapeutic management. VZV infection poses a serious threat, especially to seriously ill patients, for instance, immunocompromised patients. We report a case of human T-cell lymphotropic virus type 1-positive leukemia complicated by atypical multidermatomal herpes zoster. Viral culture and standard serological tests failed to prove VZV infection. Herpesvirus infection was confirmed by cytodiagnosis (Tzanck test). The final diagnosis of VZV was made by immunoelectron microscopy (IEM), which can differentiate between HSV and VZV. Immunoglobulin M antibodies in serum directed against VZV were detected by IEM but not by immunofluorescence. Because IEM was able to identify virus and analyze sera in only 2 h, it is considered a valuable additional tool for the rapid diagnosis of HSV and VZV infections.
Fatal pulmonary sporotrichosis caused by Sporothrix schenckii var. luriei in India
Authors: Padhye A A , Kaufman L , Durry E , Banerjee C K , Jindal S K , Talwar P , Chakrabarti A
Published: January 2020

The first case of fatal pulmonary sporotrichosis caused by Sporothrix schenckii var. luriei in a patient from the northwestern region of India is described. In the absence of cultures, the diagnosis was suspected by notation, in lung tissue, of large, thick-walled, hyaline fungal cells that divided internally by septation or a budding process. The thick-walled, internally septated cells often became muriform. The presence of an "eyeglass" configuration of incompletely separated cells characteristic of S. schenckii var. luriei in large numbers aided the diagnosis. The identity of the etiologic agent was confirmed by application of a fluorescent-antibody reagent specific for S. schenckii.
Use of polymerase chain reaction for diagnosis of picornavirus infection in subjects with and without respiratory symptoms
Authors: Johnston S L , Sanderson G , Pattemore P K , Smith S , Bardin P G , Bruce C B , Lambden P R , Tyrrell D A , Holgate S T
Published: January 2020

Rhinoviruses and enteroviruses are the major members of the picornavirus genus that cause human disease. We compared the polymerase chain reaction and viral culture for the identification of picornaviruses in nasal aspirates from children during episodes of respiratory symptoms and when asymptomatic and from asymptomatic adults. One hundred eight children, aged 9 to 11 years, completed a year-long study. Within 24 to 48 h of a report of respiratory symptoms, a nasal aspirate was taken in the home. Nasal aspirates were also taken from 65 of the children and from 33 normal adults when they had been free of respiratory symptoms for at least 2 weeks. Picornaviruses were isolated by culture for three passages in Ohio HeLa cells in rolling tubes at 33 degrees C and pH 7.0. For the polymerase chain reaction, duplicate 50-microliters samples were amplified with conserved primers from the 5' noncoding region. Picornaviruses generated approximately 380-bp bands in agarose gel electrophoresis; the specificity of these bands was confirmed by filter hybridization with a conserved internal probe. Picornaviruses were isolated by culture in 47 (46 rhinoviruses) of 292 symptomatic episodes (16%), whereas the polymerase chain reaction identified picornavirus genomic material in 146 episodes (50%), including all but one of the culture-positive episodes. As for asymptomatic samples, eight (12%) children and two (4%) adults were positive by the polymerase chain reaction, whereas only one child's specimen was positive by culture. This polymerase chain reaction assay represents a clear advance in the identification of picornavirus infection, with a detection rate threefold greater than the virus culture method.
Measles virus-specific cellular immunity in patients with vaccine failure
Authors: Wu V H , McFarland H , Mayo K , Hanger L , Griffin D E , Dhib-Jalbut S
Published: January 2020

The cytotoxic T-lymphocyte (CTL) response to measles virus (MV) was studied in blood samples from 13 acute- and early convalescent-phase patients with measles infection despite previous vaccination with the live-MV vaccine. MV CTL responses were also measured in six healthy peer controls who had live-MV vaccination during childhood and in five healthy adults who had a remote history of natural measles. All patients recovered from illness without complication. Acute MV infection was diagnosed on the basis of the Centers for Disease Control criteria and by measuring MV-specific immunoglobulin G (IgG) and IgM antibodies. Elevated IgG titers occurred in 80% of the patients at 1 to 2 weeks and in 100% at 4 weeks postinfection. IgM antibodies were detectable in all patient tested and were elevated in 60% of the patients at 1 to 2 weeks postinfection. The MV-specific CTL response was enhanced in 10 of the 13 patients tested, with a mean maximal lysis of 48.5% +/- 13.3%, compared with that of healthy peer controls who had had live-MV vaccinations during childhood (mean lysis, 14.6% +/- 12.9%; n = 6) and healthy adults with a remote history of natural measles (mean, 30.8% +/- 12.2%; n = 5). Three patients had low MV CTL levels at two time points following measles, with a mean lysis of 12% +/- 1.7%. It is concluded that while there is no evidence for a deficiency in the generation of cellular immunity to MV in the majority of patients with MV vaccine failure, a small number of individuals may fail to develop an enhanced T-cell response following infection.
Evaluation of a rapid enzyme immunoassay for detection of influenza A virus
Authors: Johnston S L , Bloy H
Published: January 2020

The Directigen FLU-A enzyme immunoassay for the detection of influenza A virus was compared with direct smear and culture in 211 clinical specimens. The FLU-A enzyme immunoassay proved to be a reliable, rapid screen for influenza A from symptomatic patients and was less dependent on technical expertise for interpretation than were direct smears.
Comparison of pooled formalin-preserved fecal specimens with three individual samples for detection of intestinal parasites
Authors: Aldeen W E , Shisenant J , Hale D , Matsen J , Carroll K
Published: January 2020

Triplicate preserved fecal samples from 265 patients were pooled into single samples, and the recovery rate of intestinal parasites from the pooled samples was compared with that from the individual samples. Parasites were detected in 109 patients when results from the individual samples were used compared with 108 patients when results from the pooled specimens were used. Pooling preserved fecal samples is an efficient and economical procedure for the detection of ova and parasites.
Evaluation of the polymerase chain reaction for diagnosis of herpes simplex virus encephalitis
Authors: Puchhammer-Stöckl E , Heinz F X , Kundi M , Popow-Kraupp T , Grimm G , Millner M M , Kunz C
Published: January 2020

Cerebrospinal fluid samples from 257 patients with suspected herpes simplex virus encephalitis were prospectively analyzed by herpes simplex virus polymerase chain reaction. The polymerase chain reaction indicated herpes simplex virus encephalitis in 9 serologically proven cases and in 14 additional patients. Increased polymerase chain reaction signals were observed together with more severe neurological symptoms (P < 0.01) and within the first days of acyclovir treatment (P < 0.05).
Sucrose-positive Edwardsiella tarda mimicking a biogroup 1 strain isolated from a patient with cholelithiasis
Authors: Walton D T , Abbott S L , Janda J M
Published: January 2020

An unusual strain of Edwardsiella tarda mimicking a biogroup 1 isolate was recovered in a mixed infection from a woman suffering from cholelithiasis. Rare biochemical characteristics (e.g., H2S negativity) originally detected were related to an unusual biochemical property in this species, sucrose fermentation; other points of interest regarding this strain included the site of isolation (bile) and the failure of this isolate to produce many of the in vitro virulence markers associated with E. tarda.
Human intestinal spirochetes are distinct from Serpulina hyodysenteriae
Authors: Lee J I , McLaren A J , Lymbery A J , Hampson D J
Published: January 2020

Twenty-nine intestinal spirochetes isolated from Australian aboriginal children and six strains from Italian adults (HRM1, -2, -4, -5, -7, and -14) were genetically examined at 15 enzyme loci by using multilocus enzyme electrophoresis. Results were compared with those previously obtained for 188 porcine intestinal spirochetes. DNA from human strain HRM7 and porcine strain Serpulina hyodysenteriae P18A were also radioactively labeled and hybridized with DNA from 12 other human and porcine intestinal spirochetes. Both the multilocus enzyme electrophoresis and hybridization techniques demonstrated that the human spirochetes were not S. hyodysenteriae. They belonged to another distinct genetic group of spirochetes that included P43/6/78, the bacterium recovered from the first recorded case of porcine intestinal spirochetosis. Bacteria in this distinct group also differed from Serpulina spp. in possessing only four, five, or occasionally six axial filaments, being slightly thinner, and having more pointed ends. These findings add further weight to the possibility that human intestinal spirochetes may act as enteric pathogens.
Rapid diagnosis of enterovirus infection by magnetic bead extraction and polymerase chain reaction detection of enterovirus RNA in clinical specimens
Authors: Muir P , Nicholson F , Jhetam M , Neogi S , Banatvala J E
Published: January 2020

We describe a rapid method for extraction and detection of enterovirus RNA in clinical samples. By using magnetic bead technology, enterovirus RNA was efficiently and rapidly extracted from cerebrospinal fluid, stool, saliva, blood, pericardial fluid, urine, and cryopreserved or formalin-fixed solid tissue. Enterovirus RNA was then detected by reverse transcription followed by polymerase chain reaction amplification with primers designed to allow detection of most enterovirus serotypes. For detection of enteroviruses in specimens from patients with acute enteroviral disease, the overall sensitivity of enzymatic RNA amplification was greater than that of cell culture isolation, especially in blood specimens and in stool specimens from patients with acute cardiac disease. Enterovirus RNA was also detected in cryopreserved and archival formalin-fixed myocardial tissue from patients with acute myocarditis and chronic dilated cardiomyopathy. The ability to study archival specimens is of particular value in conducting retrospective investigation. The RNA extraction procedure used was considerably faster than extraction methods using organic reagents, used less hazardous reagents, and was of similar sensitivity. This detection protocol may therefore be useful both for the diagnosis of enterovirus infection and in studying the pathogenesis of acute and chronic enterovirus-induced disease.
Genetic similarity of Candida albicans strains from vaginitis patients and their partners
Authors: Schmid J , Rotman M , Reed B , Pierson C L , Soll D R
Published: January 2020

The moderately repetitive sequence Ca3 was used to fingerprint strains of Candida albicans isolated from vulvovaginal infections of 10 women and strains isolated from their male partners. The Dendron software package was then used to compare the DNA fingerprints of these strains with those of vaginal commensals from women from the same geographical locale, vaginal commensals from women from a different geographical locale, and commensals from male partners of asymptomatic women from the same geographical locale. The results demonstrate that, in the majority of cases (8 of 10), strains from symptomatic patients and their partners are either identical or more similar to each other than to other strains, infecting strains do not represent a group genetically distinguishable from vaginal commensal isolates from women from the same geographical locale, and both infecting strains and commensals from individuals in the test locale can be distinguished from commensals obtained in another geographical locale. The results also suggest that women with vaginal infections are responsible for strain replacement in their male partners.
Evaluation of an enzyme immunoassay for detection of cryptococcal capsular polysaccharide antigen in serum and cerebrospinal fluid
Authors: Frank U K , Nishimura S L , Li N C , Sugai K , Yajko D M , Hadley W K , Ng V L
Published: January 2020

The Premier enzyme immunoassay (Meridian Diagnostics, Inc., Cincinnati, Ohio) was compared with a latex agglutination assay (CALAS; Meridian) for the ability to detect cryptococcal capsular polysaccharide antigen (CrAg) in serum and cerebrospinal fluid (CSF). A total of 594 specimens (471 serum samples and 123 CSF samples) obtained from 430 patients, most of whom were at risk for or had AIDS, were tested in parallel by both systems. Both tests were independently evaluated for their ability to (i) detect CrAg when used as a screening test and (ii) quantitate the CrAg present when used as a titration assay. Chart review to assess clinical outcome after the time of specimen collection was conducted for all patients. When both assays were used as screening assays, 103 serum samples and 18 CSF samples were positive and 356 serum samples and 104 CSF specimens were negative by both assays (97.8% concordance). Thirteen specimens (12 serum samples, 1 CSF sample) gave discrepant screening results. When the tests were used as semiquantitative assays for titer determinations, the CrAg titers determined by the enzyme immunoassay were generally higher than those obtained with the latex agglutination assay. In summary, results obtained with the enzyme immunoassay correlated well with those obtained with the latex agglutination test for screening for the presence of CrAg and for determining the titer of CrAg in serum or CSF.
Antibodies against papillomavirus antigens in cervical secretions from condyloma patients
Authors: Dillner L , Fredriksson A , Persson E , Forslund O , Hansson B G , Dillner J
Published: January 2020

Samples of cervical secretions and serum from 30 women with genital condylomas and 30 age-matched controls were tested for the presence of immunoglobulin A (IgA) and IgG antibodies against a panel of papillomavirus-derived antigens. The same cervical samples were also analyzed for presence of human papillomavirus (HPV) DNA by Southern blotting and polymerase chain reaction. By Southern blotting HPV DNA was detected in 8 of 30 patients with condylomas and 2 of 30 controls, and by the polymerase chain reaction HPV DNA was detected in 14 of 30 patients with condylomas and 5 of 30 controls. A total of 18 of 29 patients with condylomas and 8 of 28 controls had IgA antibodies in cervical secretions to an E2 synthetic peptide, and 17 of 29 patients with condylomas and 5 of 28 controls had local IgA antibodies to an E7 peptide (P < 0.025 and P < 0.005, respectively). The results suggest that measurement of local antibody production against selected HPV antigens may be useful in the study of HPV immunology and, possibly, for the diagnosis of HPV infection.
Epidemiological analysis of Candida albicans strains by multilocus enzyme electrophoresis
Authors: Caugant D A , Sandven P
Published: January 2020

Genotypic diversity in a collection of 98 isolates of Candida albicans was assessed by multilocus enzyme electrophoresis. Four of the 10 enzyme loci studied were polymorphic. The electrophoretic patterns observed were compatible with those expected for a diploid organism. The 98 isolates were assigned to 14 electrophoretic types, each of which was represented by from 1 to 21 isolates. Samples from various clinical sites of seven bone marrow transplant patients treated in the same unit within a 13-month period were obtained repeatedly. Three patients were found to be colonized with more than one strain. In one patient, flucytosine-resistant strains were isolated after systemic antifungal treatment was started. These isolates had electrophoretic types different from those of the strains that colonized the patient before treatment. There was no evidence that cross-infection between these patients occurred in the hospital. The population structure of C. albicans is discussed in regard to the multilocus genotype data.
Typing of Staphylococcus aureus by pulsed-field gel electrophoresis, zymotyping, capsular typing, and phage typing: resolution of clonal relationships
Authors: Schlichting C , Branger C , Fournier J M , Witte W , Boutonnier A , Wolz C , Goullet P , Döring G
Published: January 2020

Sixty-nine Staphylococcus aureus isolates from two epidemiologically unrelated sources were typed by pulsed-field gel electrophoresis after SmaI digestion of chromosomal DNA (genome typing), and the results were compared with those obtained by other typing methods: phage typing with the international set of phages, capsular serotyping with monoclonal antibodies against capsular polysaccharides type 5 and 8, and zymotyping by polyacrylamide agarose electrophoresis for esterase polymorphism. A good correlation of S. aureus types was found by these four typing methods. Differentiation increased in the order capsular typing < zymotyping < phage typing < genome typing, yielding 2, 10, 20, and 26 different S. aureus types, respectively. Five of the 26 genome types were further divided into several subtypes revealing clonal relationships. When 36 French S. aureus isolates were compared with 33 German S. aureus isolates, 3 strains representing clonal populations were identical in both groups. S. aureus isolates from patients with cystic fibrosis were also typed at the beginning and the end of a 4-week summer camp for these patients. The results suggested a possible strain transmission during the summer camp. We conclude that genome typing by pulsed-field gel electrophoresis is a powerful tool not only for strain identification but also for the resolution of the clonal relationships of S. aureus strains.
Comparison of DNA fingerprinting and serotyping for identification of avian Pasteurella multocida isolates
Authors: Wilson M A , Morgan M J , Barger G E
Published: January 2020

The DNA fingerprint profiles and serotypes of 63 avian Pasteurella multocida field isolates, 13 attenuated vaccine isolates (propagated from vaccines manufactured by five companies), and 16 somatic reference strains were compared. DNA fingerprinting established the relationship of isolates that could not be distinguished by serotyping. Of the 76 isolates, 28 DNA fingerprint profiles and 12 somatic types were recognized. One isolate was nonreactive with 16 reference somatic and 5 reference capsule-type antisera. Thirty-one field isolates and seven vaccine isolates were identified as capsule type A. Twenty-nine field isolates and six vaccine isolates were nonencapsulated. Three field isolates were capsule type F. Isolates of capsule types B, D, and E were not found. One field isolate, identified as somatic type 7, had a DNA fingerprint identical to that of the somatic reference type 6 profile. Twelve field isolates had profiles identical to the somatic reference type 3 strain profile; 11 of these were identified as somatic type 3, 4, and 1 was identified as somatic type 3. The DNA fingerprint profiles of 50 field isolates and 13 attenuated vaccine isolates did not match profiles of the 16 somatic type reference strains. Twenty-five DNA fingerprint profiles were recognized from 30 of these field isolates. The DNA fingerprint profiles of 20 field isolates and 13 attenuated vaccine isolates were identical. Three somatic types (3; 3,4; and 4,16) were identified from the field isolates, and two somatic types (3 and 3,4) were identified from the attenuated vaccine isolates. DNA fingerprinting is useful for accurate identification and epidemiologic study of P. multocida isolates.
Detection of antibodies against equine herpesvirus types 1 and 4 by using recombinant protein derived from an immunodominant region of glycoprotein B
Authors: Sinclair R , Binns M M , Chirnside E D , Mumford J A
Published: January 2020

The N-terminal fragment comprising residues +1 to +50 (gB1-50) of equine herpesvirus type 1 (EHV-1) glycoprotein B was expressed as a glutathione S-transferase fusion protein in Escherichia coli. Recombinant gB1-50 (rgB1-50) was recognized in immunoblots by sera from rabbits immunized with EHV-1 and by convalescent-phase sera from horses with natural EHV-1 infections. An enzyme-linked immunosorbent assay (ELISA) for monitoring antibody levels against EHV-1 was developed by using rgB1-50, and its specificity was assessed with a panel of reference antisera against other equine viruses. A specific cross-reaction was detected with EHV-4, which was confirmed by inhibition ELISA. Convalescent-phase sera from horses with natural EHV-1 or EHV-4 infections possessed antibody titers against rgB1-50 ranging from 1:2,000 to 1:64,000, indicating the presence of an immunodominant antigenic site. The study demonstrated the potential application of rgB1-50 as a diagnostic antigen and highlights the glutathione S-transferase fusion system as a simple and effective method of producing purified milligram quantities of antigen.
An OspA antigen-capture enzyme-linked immunosorbent assay for detecting North American isolates of Borrelia burgdorferi in larval and nymphal Ixodes dammini
Authors: Burkot T R , Wirtz R A , Luft B , Piesman J
Published: January 2020

An antigen-capture enzyme-linked immunosorbent assay (ELISA) was developed for detecting North American isolates of Borrelia burgdorferi in larval, nymphal, and adult ticks. The assay uses an anti-OspA monoclonal antibody (H5332) for antigen capture and biotin-labelled polyclonal sera with streptavidinhorseradish peroxidase for signal generation. The assay recognized 15 of 15 North American B. burgdorferi isolates and did not cross-react with spirochete antigens of Borrelia hermsii, Borrelia turicatae, Borrelia coriaceae, or Borrelia parkeri, or with tick antigens of Ixodes dammini, Ixodes scapularis, Ixodes pacificus, Ixodes cookei, Ixodes angustus, or Ambylomma americanum. The assay, with a sensitivity of less than 150 spirochetes, can detect infections in larval, nymphal, and adult ticks. In addition to fresh ticks, B. burgdorferi infections in ticks stored frozen, dried, or in 70% ethanol can be determined with the assay.
Use of monoclonal antibodies specific for the a determinant of K88 pili for detection of enterotoxigenic Escherichia coli in pigs
Authors: Westerman R B , Fortner G W , Mills K W , Phillips R M , Greenwood J M
Published: January 2020

Monoclonal antibodies directed against the a determinant of K88 pili from porcine enterotoxigenic Escherichia coli which react with all three K88 variants have been produced. These antibodies have been used for diagnosis of porcine enterotoxigenic E. coli in a direct enzyme-linked immunosorbent assay with sensitivity to 50 ng of pilus protein per ml.
Mammalian epithelial cell line kit for detection of Clostridium difficile toxin
Authors: Walpita P , Billman G F , Krous H F
Published: January 2020

The performance characteristics of a mammalian epithelial (MEP) cell line kit (Cytotoxi Test; Advanced Clinical Diagnostics, Toledo, Ohio) for the detection of Clostridium difficile toxin was compared with that of conventional tissue culture assays with human embryonic lung (HEL) cells in shell vials and human foreskin fibroblasts (HFFs) in test tubes. One hundred forty-nine stool samples were tested. The MEP cells were at least as sensitive as the HEL cells for use in C. difficile toxin detection. Results for the MEP cells were also obtained considerably more rapidly than those for HEL cells when the cells were examined at 4 and 24 h and then every 24 h for up to 5 days. Approximately one-third of all positive MEP cells were detected at 4 h and 95% were detected by 48 h. In comparison, in the HEL shell vial monolayers, only 6% of the positive cells were detectable at 4 h and 76% were detectable at 48 h. The times for C. difficile toxin-induced cytotoxicity in HFF cells were similar to those in HEL cells. Shell vials carrying HEL cell monolayers (ViroMed Laboratories Inc., Minnetonka, Minn.) are a sensitive and reliable commercial source for the detection of C. difficile toxin, although they cannot detect C. difficile as rapidly as the Cytotoxi test with the MEP cell monolayers.
Colonization by enteroaggregative Escherichia coli in travelers with and without diarrhea
Authors: Cohen M B , Hawkins J A , Weckbach L S , Staneck J L , Levine M M , Heck J E
Published: January 2020

Enteroaggregative Escherichia coli (EAggEC) has been found to be associated with pediatric diarrhea in developing countries. In order to determine the role of EAggEC as an agent of traveler's diarrhea, we used a sensitive and specific DNA probe for EAggEC to screen bacterial colony blots from 278 volunteers before and after travel. Colonization with EAggEC was infrequent (2.5%) prior to travel but rose to 27 to 33% after travel in volunteers who took either placebo or trimethoprim-sulfamethoxazole. Travelers who took trimethoprimsulfamethoxazole were colonized with organisms that were uniformly resistant to that antimicrobial agent; when volunteers received ciprofloxacin, colonization with EAggEC was prevented (2.0%). Although colonization rates were high in the placebo and trimethoprim-sulfamethoxazole groups, only a minority of travelers who were colonized with EAggEC experienced diarrhea. On the basis of our data, we suggest that colonization with EAggEC alone is not sufficient to cause traveler's diarrhea.
Analysis of homotypic and heterotypic serum immune responses to rotavirus proteins following primary rotavirus infection by using the radioimmunoprecipitation technique
Authors: Richardson S C , Grimwood K , Bishop R F
Published: January 2020

Three sequential serum samples collected from each of 20 young children with a naturally acquired primary rotavirus infection were assayed by the radioimmunoprecipitation technique for immunoglobulin G antibodies to rotavirus structural and nonstructural proteins of the four major human rotavirus serotypes G1, P1A; G2, P1B; G3, P2; and G4, P2. Fourteen children were infected with a serotype G1 rotavirus strain and six children were infected with a serotype G4 rotavirus strain. Sera were collected from each child in the acute and convalescent periods postinfection and also approximately 4 months later. Serum immune responses to rotavirus core antigens VP2 and VP3, to the major inner capsid antigen VP6, to nonstructural proteins NS35, NS28, and NS26, and to the outer capsid neutralization antigen VP4 of all test strains were detected in the majority of patients. These responses do not appear to be influenced by the G type or P type of the rotavirus strain used in the reactions. Homologous responses to the main neutralization antigen VP7 were detected in 93% of patients with serotype G1 infections and 50% of patients with serotype G4 infections. Heterologous VP7 responses were less frequently detected and were restricted to G1, G3, and G4 serotype rotavirus strains. No responses to VP7 of the serotype G2 rotavirus strain were detected in any patients. Heterotypic immune responses to the neutralization antigens, at least following serotype G1 and G4 infections, therefore appear to consist primarily of responses to VP4 rather than to VP7.
Evaluation of the fluorescence actin staining test for detection of enteropathogenic Escherichia coli
Authors: Shariff M , Bhan M K , Knutton S , Das B K , Saini S , Kumar R
Published: January 2020

Enteropathogenic Escherichia coli (EPEC) strains designated on the basis of their serotypes are epidemiologically associated with diarrhea. They adhere to the intestinal mucosa, producing the characteristic attaching and effacing (AE) lesion in an in vitro organ culture system. EPEC manifest localized adherence (LA) in the HEp-2 cell assay, and this is commonly used for clinical diagnosis. Recently, the fluorescence actin staining (FAS) test was proposed for the identification of E. coli causing the AE lesion. We therefore compared the FAS test with the HEp-2 cell assay and the EPEC adherence factor (EAF) probe assay for the detection of EPEC strains. Among 240 stool samples from children with diarrhea examined, 176 yielded E. coli and 14 of these strains showed the LA pattern in the HEp-2 cell assay; 11 of these were positive by both the EAF and the FAS tests. By using the HEp-2 cell assay as the "gold standard," the FAS test gave a sensitivity of 78.5% and a specificity of 100%. The three localized adherent FAS-negative strains tested subsequently were positive by the enteroaggregative E. coli DNA Probe and failed to produce the AE lesions characteristic of EPEC. When these strains were not considered, the sensitivity of the FAS test for detecting isolates that manifest LA was 100%. Against the EAF probe, the sensitivity and specificity of the FAS test were 91.6 and 100%, respectively. The FAS test avoids infrequent but nevertheless important phenotypic misclassifications in the HEp-2 cell assay, and it may therefore serve as a confirmatory test for EPEC.
Isolation of seven respiratory viruses in shell vials: a practical and highly sensitive method
Authors: Olsen M A , Shuck K M , Sambol A R , Flor S M , O'Brien J , Cabrera B J
Published: January 2020

The isolation of respiratory viruses in shell vials was compared with isolation in tube cultures in order to determine the sensitivity of the former, rapid method. Twenty of 21 influenza virus and 15 of 15 parainfluenza virus isolates were recovered in shell vials. One hundred twenty-seven of 138 respiratory syncytial virus isolates were detected in shell vials, but only 10 of 21 adenovirus isolates were positive by the rapid method. Shell vials are very effective for the diagnosis of respiratory viral infections, except for those caused by adenovirus.
Cervical adenitis caused by Staphylococcus epidermidis
Authors: Ryan-Poirier K , Patrick C C
Published: January 2020

Staphylococcus epidermidis, a human commensal, is a common cause of bacteremia in immunocompromised patients with indwelling medical devices. We report a case of isolated cervical adenitis caused by S. epidermidis in an immunocompetent patient and comment on the presumed pathogenesis.
Protein and antigenic analysis of Borrelia burgdorferi isolated in northern Italy: computerized analysis of phenotypic characteristics
Authors: Cinco M , De Giovannini R
Published: January 2020

Four Borrelia burgdorferi strains isolated in the same restricted geographic area share different protein patterns on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The use of polyclonal rabbit antisera and a battery of monoclonal antibodies directed toward the immunodominant proteins OspA, OspB, and pC also revealed different epitope distributions and specificities of these antigens on the strains examined. For the first time, a computerized analysis of these phenotypic characters was done mainly by cluster analysis. The computerized analysis revealed the levels of similarities among the strains and indicated, on a quantitative basis, that one of them is much closer to the American strain B31 than to the other strains.
Genus-specific polymerase chain reaction for the mycobacterial dnaJ gene and species-specific oligonucleotide probes
Authors: Takewaki S , Okuzumi K , Ishiko H , Nakahara K , Ohkubo A , Nagai R
Published: January 2020

Identification of tuberculous and nontuberculous mycobacteria by biochemical methods is a long-term process that takes up to 8 weeks for completion and requires expertise to interpret the results. In order to detect and differentiate the major pathogenic mycobacterial species, we developed genus-specific primers that amplify the dnaJ gene from the broad spectrum of mycobacterial species and determined the nucleotide sequences within the dnaJ genes from 19 mycobacterial species (Mycobacterium tuberculosis, M. bovis, M. bovis BCG, M. africanum, M. microti, M. kansasii, M. marinum, M. gastri, M. simiae, M. scrofulaceum, M. szulgai, M. gordonae, M. avium, M. intracellulare, M. xenopi, M. fortuitum, M. chelonei, M. haemophilum, and M. paratuberculosis). On the basis of the dnaJ gene sequences, we developed dot blot hybridization analysis with species-specific oligonucleotide probes for the M. tuberculosis complex. M. avium, M. intracellulare, and M. kansaii, allowing a rapid identification of these species following polymerase chain reaction for the dnaJ gene. We conclude that polymerase chain reaction with the genus-specific primer that amplifies the dnaJ genes and subsequent dot blot analysis with species-specific oligonucleotide probes are most useful for differential diagnosis of tuberculosis and nontuberculous mycobacterial infections.
Immune response against lipopolysaccharide and invasion plasmid-coded antigens of shigellae in Vietnamese and Swedish dysenteric patients
Authors: Cam P D , Pál T , Lindberg A A
Published: January 2020

The peripheral immune responses of adult Vietnamese patients infected with Shigella dysenteriae 1 and Shigella flexneri 1b and 2a and those of S. flexneri-infected Swedish patients were studied against various lipopolysaccharide and invasion plasmid-coded antigens (Ipa-s) and compared with the titers for the corresponding local healthy populations. Both Vietnamese and Swedish patients reacted with significant (P, < 0.05) titer increases in the immunoglobulin A and G classes against the homologous lipopolysaccharide antigen. However, significant titer responses against the Ipa-s were seen among only the Swedish patients. We surmise that the weak-to-moderate responses against the Ipa-s in Vietnamese patients are due to the fact that the high level of titers induced by previous infections in the local population could not be considerably increased further by a recent infection.
Use of disinfectants to reduce microbial contamination of hubs of vascular catheters
Authors: Salzman M B , Isenberg H D , Rubin L G
Published: January 2020

The vascular catheter hub is a potential portal of entry for microorganisms that cause catheter-related sepsis. Thus, a reduction in catheter hub contamination might reduce the incidence of catheter-related sepsis. To develop a regimen suitable for reducing microbial contamination of the catheter hub, we experimentally contaminated catheter hubs and assessed the efficacies of disinfectant solutions. Catheter hubs were incubated overnight with suspensions of Staphylococcus epidermidis, Pseudomonas aeruginosa, or Candida parapsilosis. After removal of unattached microorganisms, the catheter hubs were swabbed by rotating cotton swabs dipped in 1% chlorhexidine, 1% chlorhexidine in 70% ethanol, 70% ethanol, 97% ethanol, or normal saline. Posttreatment swabs of the catheter hub were obtained and cultured quantitatively. The cleaning regimens containing ethanol were the most effective. Seventy percent ethanol was more effective than chlorhexidine and is likely to be the safest treatment. We conclude that cleaning of the catheter hub with disinfectant can dramatically reduce microbial contamination.
Enteric adenovirus infection among infants with diarrhea in rural Bangladesh
Authors: Jarecki-Khan K , Tzipori S R , Unicomb L E
Published: January 2020

A total of 4,409 stool specimens from infants less than 5 years of age seeking treatment for diarrhea in Matlab, Bangladesh, were tested for the presence of adenoviruses by using an enzyme immunoassay (EIA). EIA-positive stool samples were serotyped with monoclonal antibodies specific for adenovirus type 40 (Ad40) and Ad41 and group antigen, inoculated into Graham G293 cells, and retested by EIA. Of adenovirus-positive cultures, 125 (2.8%) specimens were confirmed as enteric adenoviruses (EAds), of which 51 (40.8%) were typed as Ad40 and 74 (59.2%) were typed as Ad41, and 12 of 4,409 (0.3%) were identified as nonenteric adenoviruses. A slight peak of incidence of EAd infection was observed in the cool, dry months, and an outbreak of Ad40 infections occurred in March 1988, when the detection rate of EAd reached 12.3%. Information on age, gender, and symptoms was available for 80 infants infected with adenovirus only. Age distribution was similar for types 40 and 41 and nonenteric adenovirus; the median ages were 11, 12, and 12 months, respectively. The ratio of males to females for the 80 infants varied according to serotype; Ad40 had the highest male/female ratio, 2.17. The symptoms experienced by the 80 children were similar for each adenovirus type. The most common clinical features of EAd infection were watery diarrhea (87.5%), more than eight loose bowel movements per day in the 24-h period prior to presentation (68.8%), with vomiting (80.0%), abdominal pain (76.3%), and low-grade fever (95.0%); these symptoms are significantly similar to symptoms of infants infected with group A rotavirus. EAd infection generally gave rise to mild to moderate dehydration, which is significantly similar to dehydration produced by infection with rotavirus.
Polymerase chain reaction-amplified nonradioactive probes for identification of Fusobacterium nucleatum
Authors: Bolstad A I , Jensen H B
Published: January 2020

A polymerase chain reaction probe with 100% sequence identity to 120 deoxyribonucleotides of Fusobacterium nucleatum Fev1, coding for a part of the 40-kDa major outer membrane protein, was labeled with the steroid hapten digoxigenin. The probe was compared with various degenerate oligonucleotide probes and found to tolerate much more stringent washing conditions. It was therefore superior in distinguishing, by means of Southern blots and slot blots, F. nucleatum from other oral gram-negative bacteria in the periodontal pocket and from other fusobacterial species and in distinguishing among different strains of F. nucleatum. F. periodonticum was found to be more similar to F. nucleatum than the other fusobacterial species tested.
Genetic diversity within isolates of mutans streptococci recognized by an rRNA gene probe
Authors: Saarela M , Alaluusua S , Takei T , Asikainen S
Published: January 2020

A total of 79 clinical isolates of mutans streptococci and five laboratory strains representing serotypes c, d, e, f, and g were genotyped by a nonradioactive hybridization method with the rrnB rRNA operon of the Escherichia coli chromosome as a probe. The hybridization patterns of chromosomal DNA fragments obtained by digestion with restriction endonucleases HindIII, SmaI, and BamHI revealed genotypic heterogeneity among the serotypes and among isolates of the same serotype recovered from unrelated subjects. Diversity also existed among isolates obtained from a single subject. For 5 of 13 subjects studied, two or three genotypes within serotypes were found, while eight subjects harbored the same number of genotypes as serotypes. The data show that the method utilizing the rRNA gene probe is of value in determining the molecular epidemiology of isolates of mutans streptococci.
High-level expression of a 56-kilodalton protein gene (bor56) of Rickettsia tsutsugamushi Boryong and its application to enzyme-linked immunosorbent assays
Authors: Kim I S , Seong S Y , Woo S G , Choi M S , Chang W H
Published: January 2020

The 56-kDa protein of Rickettsia tsutsugamushi, which is located on the rickettsial surface, has been shown to be an immunodominant antigen. The gene that encodes the 56-kDa protein of R. tsutsugamushi Boryong (bor56) was cloned. Sequencing revealed an open reading frame of 1,602 bp encoding 534 amino acids with a molecular weight of 56,803. The 56-kDa protein of R. tsutsugamushi Boryong (Bor56) was expressed as a fusion protein with the maltose-binding protein of Escherichia coli by deleting 252 bp from the 5' end of the open reading frame and subcloning it into the StuI site of pIH821. The recombinant fusion protein was purified by amylose column chromatography for application in an enzyme-linked immunosorbent assay to evaluate the ability of the method to detect the antibody to R. tsutsugamushi in human patient sera. By using sera from 100 patients with scrub typhus and 70 patients with other febrile diseases, a high diagnostic sensitivity (95%) and a high diagnostic specificity (100%) were demonstrated, suggesting the suitability of the recombinant antigen for use as an immunodiagnostic tool.
Hepatitis C virus in blood samples from volunteer donors
Authors: Zhang H Y , Kuramoto I K , Mamish D , Sazama K , Holland P V , Zeldis J B
Published: January 2020

The specificities of four assays for hepatitis C virus (HCV) were compared by using units from volunteer blood donors. Upon Food and Drug Administration licensure of the first immunoassay for anti-HCV, EIA-1, units previously deemed acceptable for transfusion and all subsequent blood donations were screened. EIA-1 repeat-reactive (RR) units were tested for HCV by a second-generation enzyme-linked immunoassay (EIA-2) and by a four-antigen recombinant immunoblot assay (RIBA II) and for HCV RNA by reverse transcriptase polymerase chain reaction. All HCV RNA-positive samples were reactive by both RIBA II and EIA-2. All RIBA II-reactive sera were EIA-2 RR. In EIA-1, 0.45% of the prescreened units and 0.59% of the prospectively screened donors were RR. Of these, 52.5 and 54%, respectively, were EIA-2 RR, 71.4 and 69% of the EIA-2 RR units were reactive on RIBA II, and 93 and 88% of the RIBA II-reactive samples were HCV RNA positive. When the sample/cutoff ratio for EIA-2 was greater than 5, 91% of the samples were RIBA II reactive and 82% of the samples were HCV RNA positive. None of EIA-2 RR units with a sample/cutoff ratio of < 5 was RIBA II reactive or HCV RNA positive. In conclusion, RIBA II and RT PCR results are highly concordant. A sample/cutoff ratio of > 5 in EIA-2 is a good discriminator for the likelihood of a true HCV infection on the basis of RT PCR and RIBA II assays.
Application of DNA probes for rRNA and vanA genes to investigation of a nosocomial cluster of vancomycin-resistant enterococci
Authors: Woodford N , Morrison D , Johnson A P , Briant V , George R C , Cookson B
Published: January 2020

DNA probes specific for genes encoding rRNA and the glycopeptide resistance gene vanA were used to investigate a cluster of vancomycin-resistant (MICs, > 512 mg/liter) Enterococcus faecalis and Enterococcus faecium isolated from separate patients in a renal unit in a London hospital. When digested with BamHI, 12 of 13 vancomycin-resistant E. faecalis isolates exhibited a common restriction fragment length polymorphism pattern of rRNA genes (ribotype). A vanA probe hybridized with chromosomal DNA in these 12 isolates. The other isolate of vancomycin-resistant E. faecalis had a different ribotype and the vanA gene was located on plasmid DNA. These data suggest that cross-infection with a single strain of vancomycin-resistant E. faecalis occurred in most instances. In contrast, 23 vancomycin-resistant E. faecium isolates showed greater heterogeneity, comprising 8 ribotypes, suggesting that multiple strains were present in the unit. Twenty-one of these 23 isolates harbored a 24-MDa plasmid which hybridized with the vanA probe, implying that interstrain dissemination of a vancomycin resistance plasmid may have occurred among E. faecium isolates in the renal unit.
Immunological response to cell-free antigens of Paracoccidioides brasiliensis: relationship with clinical forms of paracoccidioidomycosis
Authors: Blotta M H , Camargo Z P
Published: January 2020

Sera from patients with the acute (AF) and chronic (CF) forms of paracoccidioidomycosis (PCM) were tested against Paracoccidioides brasiliensis cell-free antigens by Western blot (immunoblot). The CFA preparation contained components ranging in molecular mass from 18 to 102 kDa. The immunoglobulin G (IgG) reactivity profiles were similar for patients with both forms of the disease, and the 43-kDa component was recognized by 100% of the sera. IgM antibodies from the AF- and the CF-PCM sera recognized 21 and 20 components, respectively, the AF-PCM sera reacting preferentially with components with molecular masses above 50 kDa. None of the AF-PCM sera (IgM) reacted with the 43-kDa component, and only 10% of the CF-PCM sera recognized this molecule. The IgA response was more significant in the CF-PCM group than in the AF-PCM group, and the 43- and 74-kDa components were the most reactive ones (about 40% each). Our results showed that the cell-free antigen preparation is very appropriate for the immunoblotting analysis of PCM sera, and they also showed that the detection of IgG anti-gp43 is the best marker for the diagnosis and the following up of patients with the acute or the chronic form of the disease.
Enzyme immunoassay for detection of human immunodeficiency virus-specific immunoglobulin A antibodies
Authors: Connor E , Wang Z , Stephens R , Holland B , Palumbo P , McSherry G , Oleske J , Denny T
Published: January 2020

Early diagnosis of human immunodeficiency virus (HIV) infection may be difficult in adults with acute or recent HIV infection and in infants with perinatally acquired HIV. Detection of HIV-specific immunoglobulin A (IgA) antibodies in infant serum by Western blot (immunoblot) has been suggested as a reliable method to identify HIV-infected infants, especially those over the age of 6 months, and as an adjunct to diagnosis of acute HIV infection in adults. We developed a simple enzyme immunoassay for detection of HIV-specific IgA, using standard commercially available reagents. Enzyme immunoassay was comparable to Western blot for detection of HIV-specific IgA in sera from adults (n = 216), older children (n = 49), and infants born to HIV-infected mothers (n = 65). Specificity was 100% and sensitivity ranged from 80 to 92%. IgA-enzyme immunoassay is a simple, highly sensitive method for detection of HIV-specific IgA antibodies and is easily adapted to the standard clinical laboratory.
Correlation of typing methods for Acinetobacter isolates from hospital outbreaks
Authors: Dijkshoorn L , Aucken H M , Gerner-Smidt P , Kaufmann M E , Ursing J , Pitt T L
Published: January 2020

Four methods, namely, biotyping, cell envelope protein electrophoresis, ribotyping, and comparison of antibiograms, were used for strain identification of Acinetobacter isolates from five outbreaks in hospitals. There was good agreement among the methods for the identification of an index strain, but biotyping and the comparison of antibiograms were the least discriminatory.
Production of enterotoxins and toxic shock syndrome toxin by bovine mammary isolates of Staphylococcus aureus
Authors: Kenny K , Reiser R F , Bastida-Corcuera F D , Norcross N L
Published: January 2020

The production of staphylococcal enterotoxin A (SEA), SEB, SEC, SED, and SEE and toxic shock syndrome toxin 1 by bovine mammary isolates of Staphylococcus aureus was evaluated. Enterotoxin secretion was detected by immunodiffusion using specific polyclonal antisera. Of 262 isolates examined, 75 (28.6%) produced one or more toxins. The most common pattern was secretion of both SEC and SED and toxic shock syndrome toxin 1. No isolates secreted SEE, one produced SEA, and seven secreted SEB.