bluuebunny/crossref_metadata_2025 huggingface dataset:
Phosphatidylinositol-specific phospholipase C, a possible virulence factor of Staphylococcus aureus
Authors: Marques M B , Weller P F , Parsonnet J , Ransil B J , Nicholson-Weller A
Published: January 2020

Phosphatidylinositol-specific phospholipase C (PIPLC), an enzyme that can specifically release phosphatidylinositol-linked proteins from host cells, is one of the extracellular enzymes produced by Staphylococcus aureus. To investigate whether PIPLC might be a virulence factor, we assessed PIPLC production by S. aureus strains that had been isolated from healthy carriers and from infected patients with or without toxic shock syndrome. Although none of five vaginal isolates from healthy women was a PIPLC producer, only 10 of 32 selected pathogenic strains that caused significant infections or toxic shock syndrome elaborated PIPLC enzyme activity. Seven of 24 toxic-shock-associated strains, compared with 3 of 8 non-toxic-shock-associated strains, were positive for PIPLC. The majority of strains that produced PIPLC were negative for toxic shock syndrome toxin 1 (P less than 0.05); this association between PIPLC production and strains negative for toxic shock syndrome toxin 1 was even stronger among strains isolated only from patients with toxic shock syndrome (P less than 0.01). Among all 32 pathogenic isolates, PIPLC-producing S. aureus strains were isolated from four of four patients developing adult respiratory distress syndrome and four of five patients with disseminated intravascular coagulation, suggesting a significant association between PIPLC production and adult respiratory distress syndrome and/or disseminated intravascular coagulation (P less than 0.002). On the basis of these results, we propose that PIPLC is a virulence factor of S. aureus and is implicated in the development of adult respiratory distress syndrome and disseminated intravascular coagulation.
Differences in antimicrobial susceptibility of pigmented and unpigmented colonial variants of Mycobacterium avium
Authors: Stormer R S , Falkinham J O
Published: January 2020

Unpigmented colonial variants were isolated from pigmented Mycobacterium avium isolates recovered from patients with acquired immunodeficiency syndrome and the environment. The variants were interconvertible: the rate of transition from unpigmented to pigmented type was 4.0 x 10(-5) variants per cell per generation. The unpigmented variants were more tolerant to antibiotics, especially beta-lactams, and Cd2+ and Cu2+ salts than were their pigmented parents. Both pigmented and unpigmented variants of the strains produced beta-lactamase, although beta-lactamase did not appear to be a determinant of beta-lactam susceptibility. Pigmented variants grew more rapidly in a number of commonly used mycobacterial media, were more hydrophobic, and had higher carotenoid contents than their unpigmented segregants.
Immunofluorescence assay for detection of antibodies to human immunodeficiency virus type 2
Authors: Kvinesdal B B , Nielsen C M , Poulsen A G , Højlyng N
Published: January 2020

A total of 215 serum samples were tested for antibodies against human immunodeficiency virus type 2 (HIV-2) with an immunofluorescence assay (IFA). Some samples originated from Denmark and some originated from Guinea-Bissau. The IFA results were compared with enzyme-linked immunosorbent assay (ELISA) and Western (immuno-) blot (WB) results. Twenty-nine serum samples were found to be true positive for HIV-2 antibodies as judged from WB and radioimmunoprecipitation results; all of these were also found to be positive in the HIV-2 IFA. Of 80 serum samples originating from HIV-1-infected persons, 60% showed reactivity in the HIV-2 ELISA, and 51% cross-reacted with at least one band in the HIV-2 WB. None of the sera cross-reacted in the HIV-2 IFA. A total of five serum samples (three African and two Danish) gave unspecific results in the HIV-2 IFA. It is concluded that the HIV-2 IFA is more specific and at least as sensitive as a first-generation ELISA and that IFA is superior to WB in discriminating between HIV-1 and HIV-2 infections.
Isoenzyme biotypes of Candida species
Authors: Lehmann P F , Kemker B J , Hsiao C B , Dev S
Published: January 2020

Isoenzyme profiles were obtained following polyacrylamide gel electrophoresis of crude extracts of Candida albicans and C. tropicalis. The two species were separated by distinct isoenzyme patterns. Within each species, variations were found for several isoenzymes. This allowed the development of a method for biotyping these fungi. Isoenzyme patterns of the sucrose-negative variants "C. stellatoidea" and "C. paratropicalis Baker, Salkin, Pincus, and D'Amato" were obtained and subjected to cluster analysis. This procedure failed to place the variants into clusters that were clearly distinct from the conventional sucrose-positive strains. All sucrose-negative strains were found to have alpha-glucosidase activity. There was an almost complete lack of heterogeneity in the isoenzyme patterns of 11 C. stellatoidea type I strains.
Development and evaluation of an in vitro virus isolation procedure as a replacement for the mouse inoculation test in rabies diagnosis
Authors: Rudd R J , Trimarchi C V
Published: January 2020

A tissue culture virus isolation procedure for rabies street strain virus on mouse neuroblastoma cells is described. Parameters for the optimum sensitivity of the procedure were determined to include a minimum 4-day incubation of virus in tissue culture and the use of diethylaminoethyl-dextran for increased cell susceptibility. The in vitro procedure performed well in a comparison with the fluorescent-antibody test and the mouse inoculation test (MIT) on weakly positive brain tissue. Decomposed specimens and virus inhibitors present in brain suspensions were found to interfere with the in vitro procedure. A Formalin-methanol fixative was found to be superior on plastic 96-well plates to previously used fixatives. A 2-year clinical trial of the procedure in parallel with the MIT demonstrated the practicality of the procedure. Accordingly, the New York State rabies diagnostic laboratory has replaced the MIT with the in vitro procedure as a backup for the fluorescent-antibody test in the routine diagnosis of rabies.
Fermentation of carbohydrates under aerobic and anaerobic conditions by intestinal microflora from infants
Authors: Ross L F , Shaffer G P
Published: January 2020

To determine how factors involved in infant carbohydrate metabolism could be effectively assayed in vitro, fecal materials from healthy infants were obtained and tested with fermentation mixtures incorporating glucose, fructose, and lactose incubated under aerobic and anaerobic conditions. The amounts of organic acids, hydrogen, carbon dioxide, and methane produced were determined. Principal component analysis indicated that products of the three sugar systems produced very similar factor patterns and, therefore, all three sugars may not be required to adequately characterize intestinal microbial fermentation processes. Moreover, the fermentation processes under aerobic and anaerobic conditions produced essentially identical cluster patterns of the factors from the three sugar systems. Collectively, the results indicated that two sugar systems, glucose and lactose, measured under aerobic fermentation conditions sufficed in accounting for most of the variation among babies.
Rapid identification of serotypes of Mycobacterium avium-M. intracellulare complex by using infected swine sera and reference antigenic glycolipids
Authors: Ikawa H , Oka S , Murakami H , Hayashi A , Yano I
Published: January 2020

The species of 136 strains of acid-fast bacteria isolated from swine with mycobacteriosis were identified by numerical taxonomy and chemotaxonomy on the basis of mycolic acid subclass composition as members of the Mycobacterium avium-M. intracellulare (MAI) complex. The isolates were further classified by using both thin-layer chromatography of the antigenic glycopeptidolipids (GPL) obtained from the bacteria by the method of Tsang et al. (A. Y. Tsang, I. Drupa, M. Goldberg, J. K. McClatchy, and P. J. Brennan, Int. J. Syst. Bacteriol. 33:285-292, 1983) and the seroagglutination test devised by W. B. Schaefer (Am. Rev. Respir. Dis. 92[Suppl.]:85-93, 1965). For the reference standard, purified antigenic GPL of serotypes 4, 8, and 9 were isolated and their structures were analyzed by negative fast-atom bombardment-mass spectrometry. The fast-atom bombardment-mass spectrometric spectra of the intact GPL antigens of serotypes 4, 8, and 9 agreed with the structures reported earlier by Brennan et al. (P. J. Brennan and M. B. Goren, J. Biol. Chem. 254:4205-4211, 1979; P. J. Brennan, G. O. Aspinall, and J. E. Nam Shin, J. Biol. Chem. 256:6817-6822, 1981). With these antigenic GPL, the thin-layer chromatographic behaviors of the alkali-stable lipids of the above-described isolates were examined. These MAI complex isolates fell into the serotype 8 (85 strains), 4 (33 strains), and 9 (7 strains) and untypeable (11 strains) categories. Furthermore, an enzyme-linked immunosorbent assay (ELISA) based on type-specific glycolipid antigens and infected swine sera was used to diagnose the serological types of the MAI complex isolates. Of 14 cases typed by both the seroagglutination reaction and thin-layer chromatography, 13 showed clear agreement with the ELISA results. The results demonstrated that ELISA using infected sera was especially useful, and it can be recommended on the basis of simplicity, sensitivity, and specificity as an adjunct to the seroaggulutination test and thin-layer chromatography for identification of mycobacteria belonging to the MAI complex.
Evaluation of the API 20C yeast identification system for the differentiation of some dematiaceous fungi
Authors: Espinel-Ingroff A , McGinnis M R , Pincus D H , Goldson P R , Kerkering T M
Published: January 2020

Ninety-seven isolates of Cladosporium spp., Exophiala spp., Fonsecaea spp., Lecythophora hoffmannii, Phaeoannellomyces werneckii, Phialophora spp., Wangiella dermatitidis, and Xylohypha bantiana were used to evaluate the API 20C Yeast Identification System for the differentiation of dematiaceous fungi. Using the API 20C system, we were able to distinguish most species of Phialophora and Cladosporium and to separate L. hoffmannii from the species of Phialophora tested; X. bantiana from C. carrionii, C. resinae, and C. sphaerospermum; and W. dermatitidis from Exophiala jeanselmei and Exophiala spinifera. Ninety-two (60.1%) of 153 possible species-pair combinations were separated.
Immunoblot analysis of serum immunoglobulin G response to surface proteins of Clostridium difficile in patients with antibiotic-associated diarrhea
Authors: Pantosti A , Cerquetti M , Viti F , Ortisi G , Mastrantonio P
Published: January 2020

We examined by immunoblot analysis the serum immunoglobulin G antibody response to EDTA-extracted surface proteins of Clostridium difficile in 16 patients with antibiotic-associated diarrhea. For each patient, paired serum samples were tested against proteins of the infecting strain and of a collection strain (C253) known to belong to the electrophoretic group 2 pattern. Eight patients, all harboring group 2 C. difficile strains, exhibited responses to the proteins of the infecting strain; six patients showed increases in the level of antibodies between acute-phase and convalescent-phase sera. A great variability in the antigens recognized was found; however, seven patients possessed antibodies directed against an antigen of about 35 kilodaltons, corresponding to the major protein of group 2 strains. The sera of these seven patients cross-reacted also with the 35-kilodalton and other proteins of strain C253. Our data show that C. difficile proteins other than toxins can elicit an immune response in patients with C. difficile-associated disease; in this group of patients, the major surface protein of the group 2 strains was the antigen most often recognized.
Isolation, acid proteinase secretion, and experimental pathogenicity of Candida parapsilosis from outpatients with vaginitis
Authors: De Bernardis F , Lorenzini R , Verticchio R , Agatensi L , Cassone A
Published: January 2020

Candida parapsilosis was isolated from the vaginas of several nonpregnant, nondiabetic, mostly premenopausal outpatients who presented the characteristic signs and symptoms of a frank vulvovaginal candidiasis (heavy discharge with cottage cheese appearance and intense itching, with or without vulvar erythema and dyspareunia). All isolates conformed morphologically, biochemically, and serologically to the standard description of the species. They showed high acid proteinase-secretory activity in vitro and were appreciably pathogenic for cyclophosphamide-immunodepressed mice. Some isolates were also tested for their vaginopathic potential in ovariectomized rats under estradiol administration. In all cases, the rat vagina was colonized by C. parapsilosis to an extent and duration not different from those caused by a vaginopathic isolate of Candida albicans. Periodic acid-Schiff-stained vaginal smears taken at intervals during rat experimental infection showed C. parapsilosis yeasts adhering to exfoliated epithelial cells of rat vagina. Overall, these results emphasize the proteolytic and pathogenic potential of C. parapsilosis and suggest that this fungus may be an agent of clinical vaginitis.
Description of 14 new capsular types of Streptococcus suis
Authors: Gottschalk M , Higgins R , Jacques M , Mittal K R , Henrichsen J
Published: January 2020

Fourteen new capsular types of Streptococcus suis (types 9 to 22) are described. All reference strains are morphologically and biochemically similar to types previously described. Reference strain types 9 to 13, 15, 16, and 22 were isolated from diseased pigs, whereas types 17 to 19 and 21 came from clinically healthy pigs; type 14 was isolated from a human case of meningitis, and type 20 was isolated from a diseased calf. The group T streptococcus of de Moor has been included in the typing system as type 15. Two-way cross-reactions between types 6 and 16 and a one-way cross-reaction between types 2 and 22 have been demonstrated. In addition, several cross-reactions probably not due to capsular material were detected among different types by using the coagglutination test. This test should not be used alone; weak or multiple positive reactions must be confirmed by the capsular reaction test or the capillary precipitation test.
Systemic and local antibody responses in elderly subjects given live or inactivated influenza A virus vaccines
Authors: Powers D C , Sears S D , Murphy B R , Thumar B , Clements M L
Published: January 2020

Intranasal live attenuated cold-adapted (ca) influenza A/Kawasaki/9/86 (H1N1) reassortant virus and parenteral inactivated influenza A/Taiwan/1/86 (H1N1) virus were given alone or in combination to 80 ambulatory elderly subjects. An enzyme-linked immunosorbent assay was used to measure hemagglutinin-specific (HA) antibodies in serum and nasal wash specimens collected before vaccination and 1 and 3 months later. Serum immunoglobulin G (IgG) and nasal wash IgA HA responses were elicited in 56 and 20%, respectively, of 25 inactivated-virus vaccinees and in 67 and 48%, respectively, of 27 recipients of both vaccines but in only 36 and 25%, respectively, of 28 vaccinees given live virus alone. Inactivated virus, administered alone or with live virus vaccine, induced higher titers of serum antibody than did the live virus alone. In contrast, nasal IgA HA antibody was elicited more often and in greater quantity by the vaccine combination than by either vaccine alone. Despite these differences, the peak titers of local antibody mounted by each group of vaccinees were similar. By 3 months postvaccination, serum IgG and nasal IgA HA antibody titers remained elevated above prevaccination levels in 50 and 17%, respectively, of the inactivated-virus vaccinees and in 46 and 23%, respectively, of recipients of both vaccines but in only 19 and 7%, respectively, of the live-virus and systemic antibodies, if vaccinees. The finding that live ca influenza A virus induced short-lived local and systemic antibodies, if confirmed, suggests that live virus vaccination may not be a suitable alternative or adjunct to inactivated virus vaccination for the elderly.
Minitek urea disk test, a sensitive and cost-effective method to screen for Campylobacter pylori in gastric biopsies
Authors: Sweeney L , Garcia L P , Talbert M , Silverman M , Needham C A
Published: January 2020

One hundred fifty-five biopsy specimens from the gastric mucosa of 81 patients undergoing routine endoscopy procedures were tested for the presence of Campylobacter pylori by three methods: Gram stain, culture, and modified Minitek, a rapid urea disk test (BBL Microbiology Systems, Cockeysville, Md.). Twenty-nine patients were infected with C. pylori. Sensitivities and specificities of detection were 100 and 94% with the Minitek test and 93 and 100% with Gram stain, respectively. Rapid testing by the urea disk is a simple, cost-effective, and accurate method for detecting the presence of C. pylori in gastric biopsy specimens.
Probe directed at a segment of Rickettsia rickettsii rRNA amplified with polymerase chain reaction
Authors: Wilson K H , Blitchington R , Shah P , McDonald G , Gilmore R D , Mallavia L P
Published: January 2020

In an effort to explore a sensitive taxon-specific detection system for bacteria, we sequenced the 16S rRNA from two strains of Rickettsia rickettsii, five other rickettsiae, and Coxiella burnetti to find a probe site unique to R. rickettsii. We then synthesized a 16-mer that hybridized only to the rRNA of R. rickettsii. Using a primer complementary to a sequence found only in rickettsial rRNA, we then generated a cDNA. We amplified the probe site in a 180-base segment of the cDNA by using the cDNA primer and a second primer also unique to rickettsiae in a polymerase chain reaction. The segments of rRNA from each of the rickettsiae were amplified 10(6)- to 10(7)-fold, and the R. rickettsii probe hybridized only to the amplified segment from R. rickettsii. The rRNAs from Staphylococcus aureus, C. burnetii, and Neisseria meningitidis were not amplified and did not hybridize with the probe. The approach detailed below may prove clinically useful in the direct detection of pathogens that are difficult to cultivate.
Species-specific cloned DNA probes for the identification of Campylobacter hyointestinalis
Authors: Gebhart C J , Ward G E , Murtaugh M P
Published: January 2020

Conventional microbiological methods for isolating and identifying Campylobacter species are laborious, tedious, and subjective. Because of the increasing importance of Campylobacter species in human and animal diseases and the recent emergence of many new species and atypical strains, we are developing chromosomal DNA probes for rapid and simple identification of Campylobacter species, especially those of veterinary importance. We report the cloning and characterization of chromosomal DNA fragments from Campylobacter hyointestinalis, an organism isolated from pigs with proliferative enteritis. To obtain C. hyointestinalis-specific probes, chromosomal DNA fragments from C. hyointestinalis were cloned into plasmid vector pGEM-3Z. Recombinant plasmids were screened for C. hyointestinalis-specific inserts by DNA hybridization, using chromosomal DNA from either C. hyointestinalis or C. fetus which had been 32P labeled. Recombinants which hybridized to C. hyointestinalis, but not C. fetus, DNA were 32P labeled and screened further for sensitivity and specificity. Three probes were identified that were species specific and capable of detecting 10(4) C. hyointestinalis organisms by bacterial spot blotting in 48 h. We anticipate that these probes will be useful for routine species identification and for epidemiological studies.
Single primer pair for amplifying segments of distinct Shiga-like-toxin genes by polymerase chain reaction
Authors: Karch H , Meyer T
Published: January 2020

By using a single synthetic oligonucleotide primer pair in the polymerase chain reaction, we amplified specific Shiga-like-toxin (SLT) gene segments from DNAs of 20 clinical Escherichia coli isolates, irrespective of whether they produce SLT-I, SLT-II, or heretofore uncategorized SLTs. These segments were not detectable in any of 20 nontoxigenic E. coli strains. The primers deduced from a conserved region among SLT genes are so-called degenerate-sequence primers; i.e., they contain intentionally introduced sequence ambiguities to overcome minor sequence variations within different SLT genes. In direct gel hybridization with genomic DNA, both primers recognized SLT-I and SLT-II DNA sequences. Amplified sequences of target DNA obtained by polymerase chain reaction were visualized after gel electrophoresis by ethidium bromide staining, and definitive identification of the amplification product as an SLT gene segment was achieved by hybridization to SLT-I- and SLT-II-specific 20-base oligonucleotide probes complementary to a portion of the amplified sequences but not to the primers. The detecting oligonucleotide probes shared only 30% base homology and were shown to recognize specifically SLT-I or SLT-II sequences within genomic DNA. Moreover, they were used to distinguish whether the amplified sequence originated from SLT-I or SLT-II genes. The PCR system with the primers described here is a powerful technique to amplify SLT sequences in E. coli strains that produce serologically distinct SLTs and will facilitate identification of these pathogens, particularly among a multitude of nonpathogenic E. coli strains.
Thiol proteinase expression and pathogenicity of Entamoeba histolytica
Authors: Reed S L , Keene W E , McKerrow J H
Published: January 2020

Expression of the 56-kilodalton (kDa) neutral thiol proteinase has been shown to correlate with the potential of clinical isolates of Entamoeba histolytica to produce invasive disease. A 56-kDa band was identified by gelatin substrate gel electrophoresis in 10 of 10 isolates from patients with colitis or amebic liver abscesses, but in only 1 of 10 isolates from asymptomatic patients. Pathogenic isolates appear capable of releasing significantly larger quantities of the proteinase, as measured by cleavage of a synthetic peptide substrate, ZRR-AMC (benzyloxy-carbonyl-arginine-arginine-4-amino-7-methylcoumarin). We have also shown that the proteinase is released during the course of clinical invasive amebic disease, as demonstrated by the presence of circulating antibodies detectable by enzyme-linked immunosorbent assay. These studies support the importance of the 56-kDa thiol proteinase in the pathogenesis of invasive amebiasis.
Rapid in situ generation of DNA restriction endonuclease patterns for Neisseria gonorrhoeae
Authors: Poh C L , Ocampo J C , Sng E H , Bygdeman S M
Published: January 2020

Restriction endonuclease (RE) digestion patterns of 26 isolates of Neisseria gonorrhoeae representing different serovars of serogroups WI, WII, and WIII were generated by agarose pellet entrapment and in situ digestion with HinfI and BglII. The method was fast, simple, and reproducible, and stable RE patterns were produced from subsequent in vitro passages. The cost of culture materials was reduced considerably, and no toxic or flammable solvents needed to be used. Excellent resolution of DNA fragments of higher molecular weights was obtained as a result of minimal mechanical shearing of the DNA. The REs HinfI and BglII were discriminative in the fragment length ranges of 2 to 6.5 and 2.5 to 21.5 kilobases, respectively. On the basis of densitometric scanning of electrophoretograms generated by HinfI digestion, the 26 isolates representing 12 serovars were divided into seven groups. BglII was found to be more discriminative; 15 RE patterns were established among the 26 isolates. Patterns generated by both REs showed that there was no correlation between a particular RE pattern and a serovar, since strains with identical RE patterns were from different serovars. With the exception of two strains (D3 and D14), which demonstrated positive correlation when both enzymes were used, all strains with identical serovar patterns had different RE patterns.
DNA polymorphism in Mycobacterium paratuberculosis, "wood pigeon mycobacteria," and related mycobacteria analyzed by field inversion gel electrophoresis
Authors: Lévy-Frébault V V , Thorel M F , Varnerot A , Gicquel B
Published: January 2020

Mycobacterium paratuberculosis strains, mycobacteria from patients suffering from Crohn's disease, "wood pigeon mycobacteria," and representatives of Mycobacterium avium-Mycobacterium intracellulare were compared by restriction endonuclease DraI digestion and field inversion gel electrophoresis. Characteristic profiles were seen for M. paratuberculosis, including isolates from patients suffering from Crohn's disease, for wood pigeon mycobacteria, and for M. avium-M. intracellulare serotypes 2, 16, 18, and 19. Two M. paratuberculosis strains used for vaccine production (St 18 and 316 F) presented patterns different from those of the other M. paratuberculosis strains. Strains St 18 yielded a pattern identical to that of the M. avium type strain serotype 2, whereas 316 F gave a unique pattern. The method developed in this study represents a useful taxonomic tool for the identification and classification of mycobacteria.
Nonreactivity of American avian group A rotaviruses with subgroup-specific monoclonal antibodies
Authors: Theil K W , McCloskey C M
Published: January 2020

Nine group A rotavirus isolates recovered from commercially reared poultry in the United States did not react with subgroup 1 (255/60) or subgroup 2 (631/9) specific monoclonal antibodies in an enzyme-linked immunosorbent assay.
Temafloxacin disk potency and tentative interpretive criteria for susceptibility tests
Authors: Barry A L , Jones R N
Published: January 2020

Temafloxacin disk susceptibility test criteria were evaluated by testing 697 bacterial isolates. Either 5- or 10-micrograms disks could be used satisfactorily. A 5-micrograms temafloxacin disk with zone size breakpoints of less than or equal to 12 mm for resistance (MIC, greater than 4.0 micrograms/ml) and greater than 16 mm for susceptibility (MIC, less than or equal to 2.0 micrograms/ml) is recommended.
Comparative recovery of cytomegalovirus from saliva, mucolysed induced sputum, and bronchoalveolar lavage fluid from patients at risk for or with acquired immunodeficiency syndrome
Authors: Rush J D , Ng V L , Hopewell P C , Hadley W K , Mills J
Published: January 2020

The recovery rates of cytomegalovirus from mucolysed induced sputum samples and bronchoalveolar lavage fluid obtained from individuals at risk for or with acquired immunodeficiency syndrome were compared. It was demonstrated that cytomegalovirus could be reliably recovered from mucolysed induced sputum, and such recovery was highly predictive of recovery from bronchoalveolar lavage samples obtained from the same individual.
Detection of Rickettsia rickettsii DNA in clinical specimens by using polymerase chain reaction technology
Authors: Tzianabos T , Anderson B E , McDade J E
Published: January 2020

A polymerase chain reaction (PCR) procedure for detecting rickettsial DNA was developed and shown to be specific for Rickettsia rickettsii and R. conorii, the etiologic agents of Rocky Mountain spotted fever (RMSF) and Boutonneuse fever, respectively. Blood clots were obtained from nine confirmed RMSF patients and six controls and analyzed for the presence of rickettsial DNA by the PCR method. A defined region of the rickettsial genome was successfully amplified from seven of the nine clinical specimens tested; all six control specimens gave negative results. These findings indicate that R. rickettsii can be detected early after the onset of RMSF, possibly facilitating the decision regarding appropriate antibiotic therapy for some patients. Further refinement of PCR technology could make this procedure a mainstay in the clinical laboratory.
Prospective survey of colonization and infection caused by expanded-spectrum-beta-lactamase-producing members of the family Enterobacteriaceae in an intensive care unit
Authors: De Champs C , Sauvant M P , Chanal C , Sirot D , Gazuy N , Malhuret R , Baguet J C , Sirot J
Published: January 2020

The occurrence of expanded-spectrum-beta-lactamase-producing strains was prospectively studied in 56 patients in an intensive care unit. Ten patients were infected or colonized; some of these patients had asymptomatic intestinal colonization. Four different expanded-spectrum beta-lactamases were identified and used as epidemiological markers to survey nosocomial infections.
Use of a cutoff range in identifying mycobacteria by the Gen-Probe Rapid Diagnostic System
Authors: Sherman I , Harrington N , Rothrock A , George H
Published: January 2020

Commercial DNA probes (Gen-Probe Corp., San Diego, Calif.) for Mycobacterium tuberculosis complex Mycobacterium avium, and Mycobacterium intracellulare were compared with conventional methods for accuracy, applicability, and speed for the identification of putative isolates of the M. tuberculosis and M. avium complexes. Results are expressed as percent hybridization. Values of greater than 15% were considered positive, and values of less than 5% were negative. Cultures having hybridization values within an indeterminate range of 5 to 15% were repeated. Mycobacterial isolates resembling M. tuberculosis and M. avium complex from cultures of 589 specimens, representing 432 patients, were entered into this study; 294 cultures were tested with the M. tuberculosis complex probe, and 326 cultures were tested with the M. avium probe. In all cases, probe results agreed with our biochemical identification of the isolates. The M. intracellulare probe was used with 117 isolates morphologically resembling M. avium complex, and one false-negative result was observed. Seventy-two cultures gave initial hybridization results that fell within the indeterminate range and were repeated. If the manufacturer's recommended 10% cutoff value had been used, the original hybridization values would have resulted in 27 misidentified cultures, 16 false-negatives and 11 false-positives.
Genetic diversity within Streptococcus mutans evident from chromosomal DNA restriction fragment polymorphisms
Authors: Caufield P W , Walker T M
Published: January 2020

Attempts to study the acquisition, transmission, and other aspects of the natural history of Streptococcus mutans infections in humans have been hampered by limitations and inconsistencies in methods by which phenotypic characteristics of individual isolates are examined. Because most mutans streptococci associated with human dental caries fall within the biotype I (serotypes c and f) grouping, designated S. mutans, these typing methods are of little value in distinguishing individual isolates. Here we show that strains of S. mutans obtained from over 30 individuals demonstrate unique "fingerprints" of chromosomal DNA digested with restriction endonuclease HaeIII. To demonstrate that this polymorphism in restriction fragments can be used to study the acquisition and transmission of this organism, we examined isolates of S. mutans from three mother-infant pairs obtained at the time the infant first became colonized by this organism. Results indicate that strains of S. mutans found in infants exhibit restriction fragment profiles identical to those of their mothers, strongly supporting the notion that mothers transmit this organism to their infants. Also, we show that strains of S. mutans with the same restriction fragment profile were stably maintained over a 3-year interval in the one mother-infant pair studied. Moreover, we found that mothers and their infants harbored only a few individual strains, suggesting that transmission of this organism is probably confined within discrete family cohorts. Collectively, these findings demonstrate the potential utility of genomic fingerprinting in studying the natural history of S. mutans infections in humans.
Detection and quantitation of Anaplasma marginale in carrier cattle by using a nucleic acid probe
Authors: Eriks I S , Palmer G H , McGuire T C , Allred D R , Barbet A F
Published: January 2020

Cattle which have recovered from acute infection with Anaplasma marginale, a rickettsial hemoparasite of cattle, frequently remain persistently infected with a low-level parasitemia and serve as reservoirs for disease transmission. To fully understand the role of these carriers in disease prevalence and transmission, it is essential that low levels of parasitemia can be accurately detected and quantitated. We have developed a nucleic acid probe, derived from a portion of a gene encoding a 105,000-molecular-weight surface protein, that can detect A. marginale-infected erythrocytes. The probe is specific for A. marginale and can detect 0.01 ng of genomic DNA and 500 to 1,000 infected erythrocytes in 0.5 ml of blood, which is equivalent to a parasitemia of 0.000025%. This makes the probe at least 4,000 times more sensitive than light microscopy. Hybridization of the probe with treated blood from animals proven to be carriers of anaplasmosis showed that parasitemia levels were highly variable among carriers, ranging from greater than 0.0025 to less than 0.000025%. Parasitemia levels of individual animals on different dates were also variable. These results imply that, at any given time, individuals within a group of cattle may differ significantly in their abilities to transmit disease.
Determination by DNA hybridization of Shiga-like-toxin-producing Escherichia coli in children with diarrhea in Thailand
Authors: Brown J E , Echeverria P , Taylor D N , Seriwatana J , Vanapruks V , Lexomboon U , Neill R N , Newland J W
Published: January 2020

Specific DNA probes were used to identify Shiga-like-toxin (SLT) I- and II-producing Escherichia coli from children less than 5 years of age with bloody diarrhea, in infants with diarrhea, and in controls of the same age without diarrhea in Thailand. At one hospital, SLT-producing E. coli was identified in 4 (7%) of 54 children with bloody diarrhea from whom other enteric pathogens were not identified and from 3 (6%) of 50 children without diarrhea. In the positive specimens, SLT-producing E. coli constituted only 0.3 to 4% of the 100 to 300 colonies on the replica blots. Non-toxin-encoding 933J and 933W bacteriophagelike DNA sequences were detected by colony hybridization with E. coli isolates from 18 (33%) of 54 children with bloody diarrhea and 23 (46%) of 50 controls. At another hospital, SLT-producing E. coli was not identified in 115 infants with diarrhea and 119 controls without diarrhea. One infant with diarrhea was infected with E. coli O76:H7 that hybridized with the enterohemorrhagic E. coli probe but not with the SLT probes. E. coli producing SLT I or SLT II was isolated in small numbers from a similar proportion of Thai children with bloody diarrhea in whom no other enteric pathogen was identified and from controls without diarrhea.
Respiratory syncytial virus-specific antibody responses in immunoglobulin A and E isotypes to the F and G proteins and to intact virus after natural infection
Authors: Welliver R C , Sun M , Hildreth S W , Arumugham R , Ogra P L
Published: January 2020

We studied the antibody response to the fusion (F) and attachment (G) proteins of respiratory syncytial virus and to purified intact virus in the respiratory secretions of 29 infants and children. The goal of the study was to determine whether the immune response to either of the glycoproteins occurred predominantly in the immunoglobulin A (IgA) as opposed to the IgE isotype, which would indicate that one protein subunit would be a better candidate as a potential vaccine. Antibody responses were determined by using an enzyme-linked immunosorbent assay with purified F and G proteins and sucrose gradient-purified intact virus as targets. Infants and children were capable of developing an antibody response in both the IgA and IgE isotypes to each target antigen. The magnitude of the antibody response to the F protein was essentially similar to that to the intact virus, while responses to the G protein were diminished in infants. A slightly more favorable ratio of IgA to IgE responses was observed against the F protein in comparison to the G protein. While neither protein subunit had the ideal characteristics of inducing an IgA response in the absence of an IgE response, the F protein seems to be a better candidate for use as a vaccine, on the basis of better IgA/IgE ratios.
Rapid detection of group C streptococci from animals by latex agglutination
Authors: Inzana T J , Iritani B
Published: January 2020

A group C latex agglutination reagent, included as the negative control in the PathoDx Strep A latex agglutination test (Diagnostic Products Corp., Los Angeles, Calif), was compared with culture for rapid detection of group C streptococci (Streptococcus equi, S. equisimilis, S. zooepidemicus, and S. dysgalactiae) from swabs of veterinary specimens. The overall sensitivity of the latex test was 78%, and specificity was 97.6%. Only 25% of S. dysgalactiae isolates were detected, thereby accounting for the relatively low sensitivity. Ninety-three percent of the group C streptococci other than S. dysgalactiae were isolated from horses. When the latex test was evaluated for detection of group C streptococci other than S. dysgalactiae from horses, the sensitivity and specificity were 95.3 and 100%, respectively. We found the group C latex agglutination test to be a rapid and accurate method for the detection of the major pathogenic group C streptococci from swabs of equine specimens.
Clonal nature of Salmonella typhi and its genetic relatedness to other salmonellae as shown by multilocus enzyme electrophoresis, and proposal of Salmonella bongori comb. nov
Authors: Reeves M W , Evins G M , Heiba A A , Plikaytis B D , Farmer J J
Published: January 2020

Crude cell extracts of 26 isolates of Salmonella serotype typhi (S. typhi) and 48 other Salmonella isolates representing 28 serotypes and seven DNA hybridization subgroups were analyzed for electrophoretic variants of 24 metabolic enzymes by starch gel electrophoresis. All strains of S. typhi had identical isoenzyme patterns, indicating that they were a single clone. All of the enzymes detected in the remaining strains were polymorphic, and the degree of genetic variation was quite high. The average number of alleles per enzyme locus was 4.7, and the mean genetic diversity per locus was 0.556. Thirty-two distinct allele profiles, or electrophoretic types (ETs), were found in these 48 strains of Salmonella serotypes other than S. typhi. Analysis of the genetic relationships of the ETs to each other showed that, with one exception, the ETs formed subgroups that were consistent with the subgroupings based on DNA hybridization studies. ET profiles were not always linked to specific serologic patterns. These data show that multilocus enzyme electrophoresis has a potential application in epidemiologic and taxonomic studies of salmonellae, although it is not differential for S. typhi. We also propose a new species, Salmonella bongori comb. nov., a new combination base on the elevation of Salmonella choleraesuis subsp. bongori to the level of species.
Identification and classification of Campylobacter strains by using nonradioactive DNA probes
Authors: Chevrier D , Larzul D , Megraud F , Guesdon J L
Published: January 2020

Acetylaminofluorene-labeled genomic DNA probes were used for the identification and classification of Campylobacter strains. Relationships among 17 well-known strains of Campylobacter species and subspecies were studied by comparing acetylaminofluorene- or 32P-labeled probes. Results obtained with both methods were closely correlated and were in agreement with those already reported. In an identification experiment, hybridization with nonradioactive probes was performed on 60 strains isolated from stool samples after subculturing and quick DNA extraction; conventional biochemical tests were conducted in parallel. A good correlation was found between the results obtained by nonradioactive hybridization and by biochemical tests. Thus, the acetylaminofluorene-labeled genomic DNA probe method is an interesting alternative for laboratories without access to radioisotopes for the identification and classification of bacteria.
Examination of colonies and stool blots for detection of enteropathogens by DNA hybridization with eight DNA probes
Authors: Echeverria P , Taylor D N , Seriwatana J , Brown J E , Lexomboon U
Published: January 2020

We compared three methods for detecting enteropathogens in 416 children with diarrhea: (i) examination of 10 lactose-fermenting and all non-lactose-fermenting Escherichia coli (colony blots); (ii) examination of 300 colonies (replicate blots); and (iii) determination of the total bacterial growth of stools (stool blots). All specimens were spotted onto Whatman 541 filters and hybridized with specific radiolabeled DNA probes. Enterotoxigenic E. coli was detected in 38 patients by examining colony blots, in 52 patients by examining replicate blots, and in 45 patients by examining stool blots. Enteropathogenic E. coli adhesin factor was detected in 12 patients by colony blots, in 25 patients by replicate blots, and in 16 patients by stool blots. E. coli that hybridized with the enterohemorrhagic E. coli probe was detected in 2 patients by colony blots, in 11 patients by replicate blots, and in 0 patients by stool blots. Shiga-like toxin-producing E. coli was detected in 0 patients by colony blots, in 12 patients by replicate blots, and in 0 patients by stool blots. Shigella spp. were identified by standard bacteriological methods in 82 patients, and enteroinvasive E. coli was identified by colony blots in 11 patients (total, 93), by replicate blots in 56 patients, and by stool blots in 35 patients. Of 82 culture-confirmed Shigella infections, 45 were identified by examining replicate blots with the 17-kilobase-pair probe and 36 were identified by examination with the Ipa probe (P less than 0.05). Examining replicate blots with specific probes identified more enterotoxigenic E.coli (P < 0.005), enteropathogenic E.coli adhesion factor-producing E.coli (P < 0.001), and Shiga-like toxin-producing E.coli (P < 0.005) infections than examining colony blots. More Shigella and enteroinvasive E.coli infections were identified by standard bacteriological methods and examining colony blots with a specific probe than by examining replicate and stool blots (P < 0.001).
Evaluation of a reversed passive latex agglutination test for detection of Escherichia coli heat-labile toxin in culture supernatants
Authors: Scotland S M , Flomen R H , Rowe B
Published: January 2020

One hundred strains of Escherichia coli were tested for the production of the heat-labile enterotoxin by the Y1 adrenal cell test and a commercially available reversed passive latex agglutination test. The strains were grown in Casamino Acids-yeast extract broth, and filtered culture supernatants were tested for the presence of heat-labile enterotoxin. There was perfect correlation between the Y1 test and the reversed passive latex agglutination test, and the latter was simple to perform and completed within 48 h.
Identification of Campylobacter pylori by using the rapID NH system
Authors: Swiatlo A L , Attar B , Levendoglu H , Go B , Kocka F E
Published: January 2020

Campylobacter pylori has been associated with chronic active antral gastritis. The organism was isolated from 19 of 45 gastric mucosal biopsies on blood agar plates with increased CO2 at 35 degrees C. The RapID NH system, a set of dehydrated substrates for preformed enzymes, was used to assist in the identification of C. pylori. All C. pylori gave the same biochemical profile, and it was different from those of all other organisms in the profile index of the manufacturer. The RapID NH system is useful in the identification of C. pylori.
Chronic Shigella flexneri infection preceding development of acquired immunodeficiency syndrome
Authors: Simor A E , Poon R , Borczyk A
Published: January 2020

Shigella sp. is known to be an important cause of diarrhea in homosexual men, although chronic infection is infrequently recognized. We describe recurrent and relapsing symptomatic infection due to Shigella flexneri in a human immunodeficiency virus-infected homosexual man subsequently developed acquired immunodeficiency syndrome. Patients with acquired immunodeficiency syndrome may be prone to developing chronic shigellosis because of impaired intestinal cell-mediated immunity.
Seroepidemiological study of infectious mononucleosis in older patients
Authors: Kirov S M , Marsden K A , Wongwanich S
Published: January 2020

In southern Tasmania, Australia, primary Epstein-Barr virus infection occurs in adults greater than 30 years of age at a higher frequency (approximately 13% of all cases) than is generally reported for other parts of the world, and approximately 7% of the general population of the region have no antibodies to the virus. Epstein-Barr virus should not be overlooked as a possible cause of disease in older patients in similar populations elsewhere.
Rapid diagnosis of cytomegalovirus by indirect immunofluorescence assay with monoclonal antibody F6b in a commercially available kit
Authors: Lucas G , Seigneurin J M , Tamalet J , Michelson S , Baccard M , Delagneau J F , Deletoille P
Published: January 2020

Anti-human cytomegalovirus (CMV) monoclonal antibody (MAb) F6b produced by clone 95/12 was used for the rapid diagnosis (16 to 24 h) of CMV isolates in 308 clinical specimens and compared with classic isolation. MAb F6b gave 100% correlation with isolation. When this MAb was used for direct diagnosis with 212 urine and bronchoalveolar specimens, detection of CMV varied from 50 to 75% compared with viral isolation.
DNA probe for identification of Streptococcus pneumoniae
Authors: Pozzi G , Oggioni M R , Tomasz A
Published: January 2020

A total of 287 clinical isolates of Streptococcus pneumoniae (pneumococcus) were tested for their ability to undergo autolysis when treated with sodium deoxycholate. The test was positive for all but one isolate, strain DOC-1. This autolysis required the activity of an enzyme which is unique and characteristic of S. pneumoniae: a choline-dependent N-acetylmuramoyl-L-alanine amidase, the gene product of the lytA gene. We used lytA as a DNA probe to test the distribution of the autolysin gene among clinical isolates of S. pneumoniae. In dot blot hybridization experiments our probe reacted with the DNA of 60 of 60 strains tested, including the autolysis-deficient clinical isolate DOC-1. No hybridization occurred when strains of Streptococcus sanguis, Streptococcus mutans, Streptococcus pyogenes, Streptococcus (Enterococcus) faecalis, Streptococcus (Enterococcus) faecium, Streptococcus agalactiae, and Streptococcus bovis were tested. The lytA gene appears to be an ideal candidate for use as a DNA probe for the identification of S. pneumoniae.
Phialophora repens, an emerging agent of subcutaneous phaeohyphomycosis in humans
Authors: Hironaga M , Nakano K , Yokoyama I , Kitajima J
Published: January 2020

A 63-year-old Japanese man had phaeohyphomycosis that occurred as a solitary subcutaneous nodule on the dorsal aspect of his left hand. In the nodule there were foci of mixed granulomatous and suppurative infiltrations circumscribed by thick fibrous tissue reaction. The foci contained short septate hyphae and occasionally small rounded aggregates of irregularly branched septate hyphae, both of which were nonpigmented or rarely weakly pale brown. Fungal culture from the nodule was positive for a dematiaceous mold. The mycologic features of the mold were typical of Phialophora repens. The infection was successfully treated by excision of the nodule. This is the second reported case of infection due to P. repens.