bluuebunny/crossref_metadata_2025 huggingface dataset:
Rapid fluorescence method for screening Salmonella spp. from enteric differential agars
Authors: Aguirre P M , Cacho J B , Folgueira L , López M , García J , Velasco A C
Published: January 2020

Four hundred thirty-two lactose-negative colonies isolated from human feces on stool differential agar media were flooded with one drop of MUCAP Test reagent (Biolife Italiana S.r.I., Milan, Italy) and observed under a Wood lamp for the development of a blue fluorescence over or around the colony. On the basis of manual and automated conventional tests for the screening of Salmonella spp., the MUCAP Test yielded the following results: 79 true-positives, 314 true-negatives, 35 false-positives, and 4 false-negatives (sensitivity, 95%; specificity, 90%; positive predictive value, 69%; negative predictive value, 99%). The specificity of the test performed on colonies isolated on MacConkey agar (95%) was higher than that performed on colonies isolated on SS agar (88%; P less than 0.03). The MUCAP Test is an easy, rapid, and sensitive method for the screening of colonies suspected of being Salmonella spp., reducing the number of biochemical tests needed.
Rapid quantitation of DNA spot hybridization by flatbed scintillation counting
Authors: Hyypiä T , Auvinen E , Kovanen S , Ståhlberg T H
Published: January 2020

The flatbed scintillation counting system (Betaplate) was used for quantitative measurement of the radioactive hybridization signal in detection of adenovirus and papillomavirus DNA in clinical specimens. In this method, 96 samples on a nylon membrane can be handled as a single entity throughout the hybridization and counting procedure. The technique is sensitive, rapid, and convenient in routine use when compared with conventionally applied methods for the numerical analysis of hybridization results. The assay principle allows simultaneous testing of large numbers of specimens.
Antibiotic resistance patterns of group B streptococci in pregnant women
Authors: Berkowitz K , Regan J A , Greenberg E
Published: January 2020

This study examined the antibiotic resistance patterns of group B streptococci (GBS) isolated from gravid women. A total of 156 vaginal and cervical isolates of GBS were examined for resistance to penicillin, ampicillin, clindamycin, cefoxitin, gentamicin, and erythromicin. No resistance to penicillin or ampicillin was found, nor was penicillinase production demonstrated. A high level of resistance to gentamicin was noted (91%). Of the isolates examined, 9, 9.5, and 15.3% exhibited either resistance or intermediate susceptibility to erythromycin, clindamycin, and cefoxitin, respectively. Thirty strains (19%) exhibited a multiple antibiotic resistance pattern. Given the high penicillin and ampicillin treatment failure rates when attempting to eradicate vaginal GBS colonization and our findings of higher and multiple drug resistance patterns of GBS, the selection of an alternative antibiotic regimen is of considerable clinical importance. We recommend that routine reporting of GBS susceptibilities by clinical laboratories be adopted.
Use of molecular typing to study the epidemiology of Serratia marcescens
Authors: McGeer A , Low D E , Penner J , Ng J , Goldman C , Simor A E
Published: January 2020

Although Serratia marcescens is a well-known nosocomial pathogen, investigation of its hospital ecology has been limited by the lack of available typing techniques. During an investigation of the occurrence of this organism in a neonatal intensive care unit, we evaluated a number of such techniques. Using a selective medium, we conducted prospective surveillance of neonatal rectal colonization and environmental contamination with S. marcescens. In 8 months of surveillance, 5.1% (20 of 394) of the infants admitted to the unit became colonized. Most sink surfaces and drains were also culture positive. Differences between isolates could not be detected in biotypes from a commercial identification system (MicroScan) or by antibiograms, total protein fingerprints, or plasmid profiles. Serogrouping and genomic DNA restriction endonuclease analysis revealed the presence of six strains that colonized infants and a similar number of environmental strains. These two methods were concordant, with the exception that genomic DNA analysis demonstrated lack of relatedness between some strains within the same serogroup. DNA restriction endonuclease analysis was practical and reliable. The differences this method detected between environmental and neonatal strains provided strong evidence that the environment was not an important reservoir for S. marcescens in our neonatal intensive care unit.
Pasteurella multocida and Bordetella bronchiseptica infections in rabbits
Authors: Deeb B J , DiGiacomo R F , Bernard B L , Silbernagel S M
Published: January 2020

The natural history of infection with Pasteurella multocida and Bordetella bronchiseptica in domestic rabbits was studied prospectively at a commercial rabbitry. At weaning, about 25% of rabbits had nasal infections with P. multocida and 75% had infections with B. bronchiseptica. Infection of weanling rabbits paralleled nasal infections of their dams. The proportion of rabbits with both infections increased with age. At 2 to 4 months old, about 50% of rabbits with P. multocida or P. multocida and B. bronchiseptica infections had upper respiratory disease (URD), whereas rabbits with B. bronchiseptica infection had no disease. In rabbits about 10 months old, 75% with P. multocida or P. multocida and B. bronchiseptica infections had URD, whereas virtually none with B. bronchiseptica infection had disease. Disease of the nares, paranasal sinuses, middle ears, and lungs was associated with P. multocida and not B. bronchiseptica infection. In adult rabbits with nasal P. multocida infection, with or without signs of URD, about 80% had concurrent infection of the paranasal sinuses and middle ears and 20% had infection of the bronchi and lungs. In rabbits without nasal P. multocida infection, 20 to 35% had P. multocida infection of the paranasal sinuses and middle ears. Weanling rabbits with and without P. multocida infection had similar immunoglobulin G (IgG) levels. In rabbits observed prospectively, the only antibody differences between those transiently and persistently infected with P. multocida were a diminished IgA response in nasal lavages and an earlier IgM response in sera of transiently infected rabbits. IgG levels increased with the duration of infection. There was no relationship between immunoglobulin levels and freedom from P. multocida infection.
Native valve endocarditis caused by an organism resembling Corynebacterium striatum
Authors: Markowitz S M , Coudron P E
Published: January 2020

An organism resembling Corynebacterium striatum was isolated from the blood of a patient with acute aortic valvular insufficiency and no history of valvular heart disease. At autopsy, histopathologic examination of the aortic valve revealed pleomorphic gram-positive bacilli and destruction of valvular tissue. Our isolate differed from other nondiphtherial corynebacteria, including the type strain of C. striatum (ATCC 6940), in its ability to reduce nitrite. Nitrite reduction may be useful for distinguishing strains of corynebacteria.
Monoclonal antibodies to Nocardia asteroides and Nocardia brasiliensis antigens
Authors: Jiménez T , Díaz A M , Zlotnik H
Published: January 2020

Nocardia asteroides and Nocardia brasiliensis whole-cell extracts were used as antigens to generate monoclonal antibodies (MAbs). Six stable hybrid cell lines secreting anti-Nocardia spp. MAbs were obtained. These were characterized by enzyme-linked immunosorbent assay, Western blot (immunoblot), and immunofluorescence assay. Although all the MAbs exhibited different degrees of cross-reactivity with N. asteroides and N. brasiliensis antigens as well as with culture-filtrate antigens from Mycobacteria spp., they have the potential for use as reagents in the purification of Nocardia antigens.
Comparison of indirect immunofluorescent-antibody assay, enzyme-linked immunosorbent assay, and Western immunoblot for the diagnosis of Lyme disease in dogs
Authors: Lindenmayer J , Weber M , Bryant J , Marquez E , Onderdonk A
Published: January 2020

Enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescent-antibody assay (IFA), and Western immunoblot were used to test serum samples from 128 dogs for the presence of antibody to Borrelia burgdorferi. Sera included 72 samples from dogs suspected of having Lyme disease, 32 samples from dogs residing in areas in which Lyme disease was not considered endemic, and 24 samples from dogs with clinical and serologic evidence of immune-mediated disease (n = 10), Rocky Mountain spotted fever (n = 5), or leptospirosis (n = 9). Results of Western immunoblotting were used as the standard against which performances of ELISA and IFA were measured. ELISA was significantly more sensitive than IFA (84.8 versus 66.7%), although both tests were equally specific (93.5%). Eight samples that were positive by Western immunoblot were simultaneously negative by ELISA and IFA. Of these eight, four were from dogs suspected of having immune-mediated disease, two were from dogs suspected of having leptospirosis, and two were from dogs suspected of having Lyme disease. These results may indicate that sera from dogs with immune-mediated disease, and to a lesser extent sera from those with leptospirosis, cross-react with B. burgdorferi antigens. Alternatively, Western immunoblot results may not truly reflect Lyme disease status, particularly in the case of dogs with immune-mediated diseases. At present, however, the use of Western immunoblotting as a diagnostic standard for dogs offers the best alternative to a clinical definition of disease.
Mechanism of plasma clotting by Erysipelothrix rhusiopathiae
Authors: Takahashi T , Takahashi I , Tamura Y , Sawada T , Yoshida T , Suzuki S , Muramatsu M
Published: January 2020

Experiments were performed to determine the mechanism by which Erysipelothrix rhusiopathiae clots plasma. Detection of plasma-clotting activity in four strains of E. rhusiopathiae was carried out by mixing a 24-h broth culture of a tested bacterial strain with rabbit plasma (tube coagulation test). Sodium citrate, sodium oxalate, EDTA, and heparin were used as anticoagulants in preparing the rabbit plasma. E. rhusiopathiae strains clotted solely citrated plasma in 18 to 24 h. A known coagulase-positive strain of Staphylococcus aureus clotted all of the plasma preparations within 1 h. Various constituents of the organisms, such as cell-free culture filtrates, sonicated extracts, and Formalin-killed bacteria, were also checked for their ability to clot citrated plasma. No constituents of any strain of E. rhusiopathiae clotted the plasma. Only culture filtrates of S. aureus clotted the plasma under these conditions. The spectrophotometric assay demonstrated that two plasma-clotting strains of E. rhusiopathiae consumed the citrate in the plasma just before clotting. Of 301 veterinary clinical isolates of E. rhusiopathiae, 267 (88.7%) were positive in the tube coagulation test. On the basis of these results, it was concluded that plasma clotting by E. rhusiopathiae was due not to extracellular factors such as staphylocoagulase but to consumption of the citrate in the plasma.
Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting analysis of smooth-lipopolysaccharide heterogeneity among Brucella biovars related to A and M specificities
Authors: Garin-Bastuji B , Bowden R A , Dubray G , Limet J N
Published: January 2020

Smooth (S)-lipopolysaccharide (LPS) preparations from reference and field strains of several biovars of Brucella abortus, B. melitensis, and B. suis were prepared by (i) the hot phenol-water method, (ii) hot sodium dodecyl sulfate extraction and proteinase K digestion, or (iii) dimethyl sulfoxide extraction. These S-LPS-enriched fractions were further analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining after periodate oxidation. Immunoblots were developed by using either monoclonal antibodies specific for Brucella A or M antigens or polyclonal polyspecific or monospecific sera from rabbits, cattle, and goats. The specificity of monoclonal antibodies reactive with Brucella unique (A or M) epitopes was demonstrated by enzyme-linked immunosorbent assay, LPS latex agglutination, or agglutination inhibition. The most-represented subunits of S-LPS ranged in Mr from 30,000 to 70,000 relative to marker proteins. According to A or M immunodominance, two sodium dodecyl sulfate-polyacrylamide gel electrophoresis banding patterns were clearly distinguished among biovars, whatever the fraction tested: a close succession of regularly spaced narrow bands for A greater than M strains and regularly spaced triplets of bands including either (i) a first thin band followed by two thick bands for B. abortus M greater than A strains or (ii) one thick band between two thin bands for B. melitensis or B. suis M greater than A strains. Moreover, A and M specificities were reaffirmed by sandwich enzyme immunoassay and latex agglutination inhibition with monoclonal antibodies and polyclonal sera.
Detection of Epstein-Barr virus by polymerase chain reaction
Authors: Telenti A , Marshall W F , Smith T F
Published: January 2020

The polymerase chain reaction (PCR) was used to study DNA extracted from the blood of 25 transplant patients, 5 patients with infectious mononucleosis, and 13 healthy subjects and autopsy or biopsy tissue from 29 patients with lymphoproliferative disorders. Primers were directed to conserved regions of the Epstein-Barr virus (EBV) genome encoding capsid protein gp220 and Epstein-Barr nuclear antigen 1. Specific EBV amplification was found in the blood of 11 of 25 transplant patients and all patients with infectious mononucleosis. All patients with lymphoproliferative disorders occurring in the presence of immunosuppression (eight organ transplant patients and two patients with acquired immunodeficiency syndrome) had biopsies positive for EBV by PCR. Only 1 of 19 samples from lymphomas or leukemias unrelated to immunosuppression contained EBV. PCR confirmed the very close association of EBV and lymphoproliferative disorders occurring in the presence of immunosuppression. The significance of detecting EBV sequences in the blood of transplant patients, particularly in relationship to lymphoproliferation, requires further study.
Polymerase chain reaction for detection of Mycobacterium tuberculosis
Authors: Sjöbring U , Mecklenburg M , Andersen A B , Miörner H
Published: January 2020

A polymerase chain reaction for the specific detection of mycobacteria belonging to the Mycobacterium tuberculosis complex was developed. Using a single primer pair derived from the nucleotide sequence of protein antigen b of M. tuberculosis, we achieved specific amplification of a 419-base-pair DNA fragment in M. tuberculosis and M. bovis. After DNA was extracted from mycobacteria by using a simple, safe lysis procedure, we detected the 419-base-pair sequence in samples containing few mycobacteria. Preliminary data suggested that this technique could be applied to clinical specimens for early and specific diagnosis of tuberculosis.
Detection and quantitation by lysis-filtration of bacteremia after different oral surgical procedures
Authors: Heimdahl A , Hall G , Hedberg M , Sandberg H , Söder P O , Tunér K , Nord C E
Published: January 2020

Patients with bacteremia after dental extraction, third-molar surgery, dental scaling, endodontic treatment, and bilateral tonsillectomy were studied by means of lysis-filtration of blood samples with subsequent aerobic and anaerobic incubation. Samples were obtained before, during, and 10 min after treatment. Bacteremia was observed in 100% of patients after dental extraction, 55% of patients after third-molar surgery, 70% of patients after dental scaling, 20% of patients after endodontic treatment, and 55% of patients after bilateral tonsillectomy. Anaerobic microorganisms were isolated more frequently than aerobic microorganisms were, and viridans group streptococci were the most commonly isolated bacteria. Ten minutes after treatment, the frequency as well as the magnitude of bacteremia showed pronounced reduction.
Efficacies of selected disinfectants against Mycobacterium tuberculosis
Authors: Best M , Sattar S A , Springthorpe V S , Kennedy M E
Published: January 2020

The activities of 10 formulations as mycobactericidal agents in Mycobacterium tuberculosis-contaminated suspensions (suspension test) and stainless steel surfaces (carrier test) were investigated with sputum as the organic load. The quaternary ammonium compound, chlorhexidine gluconate, and an iodophor were ineffective in all tests. Ethanol (70%) was effective against M. tuberculosis only in suspension in the absence of sputum. Povidone-iodine was not as efficacious when the test organism was dried on a surface as it was in suspension, and its activity was further reduced in the presence of sputum. Sodium hypochlorite required a higher concentration of available chlorine to achieve an effective level of disinfection than did sodium dichloroisocyanurate. Phenol (5%) was effective under all test conditions, producing at least a 4-log10 reduction in CFU. The undiluted glutaraldehyde-phenate solution was effective against M. tuberculosis and a second test organism, Mycobacterium smegmatis, even in the presence of dried sputum, whereas the diluted solution (1:16) was only effective against M. smegmatis in the suspension test. A solution of 2% glutaraldehyde was effective against M. tuberculosis. This investigation presents tuberculocidal efficacy data generated by methods simulating actual practices of routine disinfection.
Electrophoretic analysis of the surface components of autoagglutinating surface array protein-positive and surface array protein-negative Aeromonas hydrophila and Aeromonas sobria
Authors: Kokka R P , Vedros N A , Janda J M
Published: January 2020

The protein and lipopolysaccharide (LPS) compositions of 10 autoagglutinating Aeromonas hydrophila and Aeromonas sobria strains were studied; one group consisted of five serogroup O:11 strains that contained an S layer, while a second group was composed of diverse serogroups that were S layer negative by transmission electron microscopy. All serogroup O:11 strains were found to contain a predominant 52,000- to 54,000-molecular-weight protein that was present on both whole-cell and outer membrane protein profiles; this protein was found to be glycine extractable under low-pH (pH 4) conditions and was identified as the surface array protein. LPS analysis revealed that all O:11 strains exhibited homogeneous-length O-polysaccharide side chains characterized primarily by two or three major bands. In contrast, S-layer-negative autoagglutinating strains of other serogroups lacked this predominant surface array protein, and silver stain analysis of LPS indicated that such profiles mainly consisted of core antigens and were deficient in or devoid of O-polysaccharide side chains. These collective results offer potential explanations for observed differences between these two groups in virulence, disease spectrum, and pathogenic properties.
Characterization by DNA restriction endonuclease analysis of Listeria monocytogenes strains related to the Swiss epidemic of listeriosis
Authors: Nocera D , Bannerman E , Rocourt J , Jaton-Ogay K , Bille J
Published: January 2020

Listeria monocytogenes strains responsible for outbreaks of listeriosis were studied by using serotyping and phage typing. An additional approach based on restriction endonuclease analysis (REA) of the chromosomal DNA was used to characterize L. monocytogenes strains collected from various sources during and after a Swiss outbreak of listeriosis (1983 to 1987). Among the 169 wild-type strains of Listeria spp. that were examined, 161 (95%) belonged to the species L. monocytogenes, of which 109 were of human origin. Ten different REA profiles were obtained from the 120 L. monocytogenes serotype 4b strains tested. All 57 serotype 4b strains that were identified as Swiss epidemic strains by phage typing clustered in two closely related REA profiles. In particular, 10 L. monocytogenes 4b strains isolated from the brand of soft cheese responsible for the outbreak and from its direct environment were indistinguishable from isolates from 40 patients by both phage typing and REA analysis. However, 5 of the 17 non-phage-typeable L. monocytogenes strains and 18 L. monocytogenes strains with a phage type different from those of the Swiss epidemic types showed the same profile. REA enabled the characterization of non-phage-typeable strains and, thus, seems a promising tool for L. monocytogenes typing, especially during epidemiological investigations.
Biochemical and serological characterization of Bacteroides intermedius strains isolated from the deep periodontal pocket
Authors: Dahlén G , Wikström M , Renvert S , Gmür R , Guggenheim B
Published: January 2020

Fifty-one fluorescence-positive black-pigmented Bacteroides strains obtained from 51 patients with deep periodontal pockets (greater than 6 mm) were identified and characterized. Fifty of these strains were presumptively identified as Bacteroides intermedius according to the indole reaction. This was confirmed by further biochemical characterization. The 50 strains from diseased sites were then compared with 16 B. intermedius strains isolated from periodontally healthy individuals with no signs of destructive periodontal disease. Tests for antimicrobial susceptibility showed similar patterns for all 50 pocket-derived strains, except for one beta-lactamase-positive strain that was resistant to penicillin G and ampicillin. Forty-seven strains were tested for binding of three monoclonal antibodies defining three distinct serogroups of B. intermedius. Thirty-one strains belonged to serogroup I, three to serogroup II and thirteen to serogroup III. In comparison to the strains from the shallow periodontal pockets, serogroup I was significantly overrepresented in the patient group with periodontal disease. We conclude that saccharolytic black-pigmented Bacteroides species from deep periodontal pockets constituted, with very rare exceptions, a biochemically homogeneous but antigenically heterogeneous group of B. intermedius and that serogroup I is predominantly found in deep periodontal lesions.
Identification of a double-stranded RNA virus by using polymerase chain reaction and magnetic separation of the synthesized DNA segments
Authors: Rimstad E , Hornes E , Olsvik O , Hyllseth B
Published: January 2020

A double-nested polymerase chain reaction assay (PCR), followed by magnetic separation of the PCR-synthesized DNA segments, was developed to detect a double-stranded RNA virus, infectious pancreatic necrosis virus from salmonid fish. Viral RNA was extracted from cell cultures and used for cDNA synthesis. The cDNA produced was used as a template in a double PCR. The sensitivity of this double PCR was approximately 0.8 pg of template double-stranded RNA. The DNA segment produced from the first PCR was also used as a template in a second PCR with a set of two 5'-labeled primers, one with biotin and the other with 32P. The PCR segment that was then synthesized was separated from the solution by using streptavidin-coated, superparamagnetic beads. The levels of radioactivity measured in the magnetically separated fractions were significantly higher in the positive samples than they were in the negative samples.
Rapid prenatal diagnosis of congenital Toxoplasma infection by using polymerase chain reaction and amniotic fluid
Authors: Grover C M , Thulliez P , Remington J S , Boothroyd J C
Published: January 2020

Infection of pregnant women with Toxoplasma gondii places the developing fetus at risk for congenital infection. We report a prospective study of 43 documented cases of acute maternal Toxoplasma infections acquired during gestation in which the polymerase chain reaction (PCR) was evaluated for diagnosis of fetal infection and compared with the current standard methods. On the basis of direct lysis of pelleted amniotic fluid cells followed by amplification of a gene sequence specific for T. gondii, PCR correctly identified the presence of T. gondii in five of five samples of amniotic fluid from four proven cases of congenital infection. PCR also detected three of five positive cases from a nonprospective group. The two diagnostic methods of comparable speed, detection of specific immunoglobulin M from fetal blood and and inoculation of amniotic fluid into tissue culture, correctly identified only 3 and 4 of the 10 positive samples, respectively. The considerably more time-consuming methods of mouse inoculation of amniotic fluid and fetal blood both detected 7 of 10 positive samples. There were no false-positive diagnoses by any of the methods. Therefore, detection of T. gondii by PCR appears to be the most promising method for prenatal diagnosis of congenital Toxoplasma infection, since it is both extremely rapid and highly sensitive.
Serodiagnosis of infectious mononucleosis by using recombinant Epstein-Barr virus antigens and enzyme-linked immunosorbent assay technology
Authors: Gorgievski-Hrisoho M , Hinderer W , Nebel-Schickel H , Horn J , Vornhagen R , Sonneborn H H , Wolf H , Siegl G
Published: January 2020

Four recombinant, diagnostically useful Epstein-Barr virus (EBV) proteins representative of the viral capsid antigen (p150), diffuse early antigen (p54), the major DNA-binding protein (p138), and the EBV nuclear antigen (p72) (W. Hinderer, H. Nebel-Schickel, H.H. Sonneborn, M. Motz, R. Kühbeck, and H. Wolf, J. Exp. Clin. Cancer Res. 7[Suppl.]:132, 1988) were used to set up individual enzyme-linked immunosorbent assays (ELISAs) for the qualitative and quantitative detection of immunoglobulin M (IgM) and IgG antibodies. In direct comparison with results obtained by standard immunofluorescence or immunoperoxidase assays, it was then shown that the recombinant EBV ELISAs provide the means for specific and sensitive serodiagnosis of infectious mononucleosis (IM) caused by EBV. The most useful markers in sera from such patients proved to be IgM antibodies against p54, p138, and p150. Additional positive markers for recent or ongoing IM apparently were IgG antibodies against p54 and p138. In contrast, anti-p72 IgG had a high preference for sera from healthy blood donors and, therefore, can be considered indicative of past exposure to the virus. Altogether, the individual ELISAs proved to be as specific and at least as sensitive for the diagnosis of IM as the currently available standard techniques are. Moreover, our findings suggest that, by combining individual test antigens, a workable ELISA system consisting of three assays (IgM against p54, p138, and p150; IgG against p54 and p138; and IgG against p72) can be established for the standardized rapid diagnosis of acute EBV infections.
Production of hemolysin and other extracellular enzymes by clinical isolates of Pseudomonas pseudomallei
Authors: Ashdown L R , Koehler J M
Published: January 2020

One hundred clinical isolates of Pseudomonas pseudomallei from humans were tested for their ability to produce extracellular, biologically active substances which are thought to contribute to the virulence of Pseudomonas species. All isolates produced at least on extracellular enzyme; 91 strains were positive for lecithinase, lipase, and protease; but none was positive for elastase. Ninety-three strains produced a hemolysin which was detectable around the heavy growth on saline-washed sheep erythrocyte brain heart infusion agar but not demonstrable around individual colonies or in broth culture filtrate. In contrast, a hemolysin which was cytolytic around individual colonies of P. pseudomallei on the assay plate and in broth culture filtrate was exhibited by four strains. By using one of these four isolates as the test strain, the latter hemolysin was characterized further. It was heat labile, most active in an acid environment (pH 5.5), and cytolytic in broth culture filtrate for a variety of animal and human erythrocytes. Sterols, particularly cholesterol and 7-dehydrocholesterol, inhibited its hemolytic activity, but the activity was not enhanced by reducing agents or suppressed by reagents which modify sulfhydryl-activated hemolysins. A nonhemolytic mutant of the test strain of P. pseudomallei retained the extracellular enzymes of its parent, indicating that the hemolysin was not a lecithinase, lipase, or protease.
Differentiation of Genes Coding for
Escherichia coli
Verotoxin 2 and the Verotoxin Associated with Porcine Edema Disease (VTe) by the Polymerase Chain Reaction
Authors: Johnson W. M. , Pollard D. R. , Lior H. , Tyler S. D. , Rozee K. R.
Published: January 2020

Two sets of synthetic oligonucleotide primers were used in a polymerase chain reaction adaptation to distinguish the closely related genes for type 2 verotoxin (VT2 or Shiga-like toxin [SLT-II]) and the verotoxin associated with porcine edema disease (VTe or SLT-II variant [SLT-IIv]) in
Escherichia coli.
Analysis with restriction endonucleases recognizing 4- or 5-base-pair sequences of human adenovirus type 3 isolated from ocular diseases in Sapporo, Japan
Authors: Itakura S , Aoki K , Sawada H , Shinagawa M
Published: January 2020

A total of 49 adenovirus type 3 (Ad3) strains from patients with epidemic keratoconjunctivitis at an eye clinic in Sapporo, Japan, from 1983 to 1986 were classified into either genome type Ad3f (26 strains) or Ad3g (23 strains). By the use of Hinf1 and TaqI restriction endonucleases, they were classified into 11 and 3 subgenome types, respectively. During this observation period, two epidemics of Ad3 infection relating to epidemic keratoconjunctivitis occurred. The prevalent Ad3g strains isolated during the epidemic in 1983 belonged to a single subgenome type. However, in the course of the Ad3f epidemic in 1986, alteration of one subgenome type of the prevalent strain to a different subgenome type which was presumed to be a derivative of the former was observed. In the years in which only a few Ad3 strains were isolated, such Ad3f strains were classified into different subgenome types. However, all Ad3g strains except two belonged to the same subgenome type.
Campylobacter-like organisms are uncommon pathogens in patients infected with the human immunodeficiency virus
Authors: Wilcox C M , Byford B A , Forsmark C E , Hadley W K , Cello J P , Jacobson M A
Published: January 2020

Over a 25-month period, we prospectively evaluated 36 patients with symptomatic human immunodeficiency virus disease (including 27 with unexplained chronic diarrhea) by flexible sigmoidoscopy for the presence of Campylobacter-like organisms. No Campylobacter-like organisms were isolated. Campylobacter-like organisms appear to be an uncommon cause of idiopathic chronic diarrhea in symptomatic human immunodeficiency virus disease.
Clostridium septicum as a cause of pericarditis and mycotic aneurysm
Authors: Brahan R B , Kahler R C
Published: January 2020

Clostridium septicum is a bacterial species associated with gas gangrene in both humans and animals. Although not usually a pathogen in humans, it has been implicated in some cases of abscesses and bacteremia. We now report the first case of pericarditis with mycotic aneurysm due to C. septicum.
High-level production of Escherichia coli STb heat-stable enterotoxin and quantification by a direct enzyme-linked immunosorbent assay
Authors: Urban R G , Pipper E M , Dreyfus L A , Whipp S C
Published: January 2020

A convenient and sensitive enzyme-linked immunosorbent assay (ELISA) for the STb heat-stable enterotoxin of Escherichia coli was developed and used to quantify STb production by strains with a high level of expression. Based on an antigenic profile of the secreted form of STb, a synthetic peptide (STb3-27) spanning the major predicted epitope was synthesized, coupled to keyhole limpet hemocyanin, and used to immunize rabbits. Anti-STb3-27 antibodies were affinity purified on a synthetic peptide-Sepharose 4B column and used in a direct-binding STb ELISA. Based on a highly purified form of toxin as a standard, the ELISA detected as little as 1 to 2 ng of STb from crude culture filtrates. ELISA data revealed that natural STb-producing strains elaborate little STb in defined-medium cultures relative to that elaborated by a recombinant strain harboring a cloned copy of the estB gene. Replacement of the endogenous STb promoter with any of several highly active promoters, including a bacteriophage T7 promoter, a beta-galactosidase promoter, and a tryptophan-beta-galactosidase hybrid (tac) promoter, increased the yield of STb 10- to 20-fold over levels obtained by an E. coli strain harboring the recombinant estB gene. The high level of STb antigen detected by the ELISA correlated with intestinal secretory activity. The combination of a convenient assay and effective hyperproduction of STb will serve as a basis for a large-scale toxin purification strategy.
Evaluation of direct immunofluorescence, enzyme immunoassay, centrifugation culture, and conventional culture for the detection of respiratory syncytial virus
Authors: Johnston S L , Siegel C S
Published: January 2020

Four methods of detecting respiratory syncytial virus (RSV) from clinical specimens were evaluated. A total of 410 specimens consisting of nasopharyngeal washes, aspirates, and swabs were simultaneously tested for the presence of RSV by direct immunofluorescence assay (DFA), enzyme immunoassay (EIA) (Kallestad Pathfinder), shell vial centrifugation culture (SVC), and conventional culture. DFA identified 146 (83%) of the 175 positive cases, EIA detected 153 (87%), SVC detected 127 (73%), and conventional culture detected 70 (40%). Conventional culture isolated an additional 19 respiratory viruses other than RSV. DFA and EIA were able to detect nonviable virus not isolated by a culture method, and SVC isolated low-titer virus not detected by conventional culture. DFA and EIA gave similar results; however, the EIA system was less dependent on technical expertise. The use of SVC enhanced the conventional culture system with 63 RSV isolates not recovered from the tube culture. We recommend complementary use of both culture and nonculture methods in the detection of RSV.
Hemolysins and other characteristics that help differentiate and biotype Staphylococcus lugdunensis and Staphylococcus schleiferi
Authors: Hébert G A
Published: January 2020

Reference strains and clinical isolates representing the newly defined species Staphylococcus lugdunensis and Staphylococcus schleiferi were examined with the battery of tests previously recommended (G.A. Hébert, C.G. Crowder, G.A. Hancock, W.R. Jarvis, and C. Thornsberry, J. Clin. Microbiol. 26:1939-1949, 1988) for other species of coagulase-negative staphylococci (CNS). The Staph-Ident system (Analytab Products, Plainview, N.Y.) supplemented with tests for synergistic hemolysis, adherence to glass, pyroglutamyl-beta-naphthylamide hydrolysis, and susceptibility to a set of five antimicrobial disks differentiated each of these species from other species of CNS and separated strains within each species into several biotypes. Most strains (95%) of S. lugdunensis produced a delta hemolysin like that seen with nine other species of CNS. Most strains (91%) of S. schleiferi produced a beta hemolysin, which is a unique characteristic among CNS. Most (95%) of the S. schleiferi but very few (12%) of the S. lugdunensis were adherence positive. Both hemolysins and adherins are potential virulence factors among CNS. Some (29%) of the S. lugdunensis were beta-lactamase positive. The S. lugdunensis were resistant to polymyxin B and bacitracin (10 U), but the S. schleiferi were susceptible to both disks. Clinical isolates of S. lugdunensis were aligned in 18 biotypes because of eight biochemical profiles and eight physiologic subtypes; isolates of S. schleiferi were in 8 biotypes because of three biochemical profiles and subtypes. These tools for correctly identifying and then biotyping two more clinical species of CNS should enhance both epidemiologic and ecologic investigations.
Adherence measured by microtiter assay as a virulence marker for Staphylococcus epidermidis infections
Authors: Deighton M A , Balkau B
Published: January 2020

Staphylococcus epidermidis strains isolated from clinical sources showed a wide range of abilities to adhere to glass and plastic materials. The degree of adherence depended on a number of factors, most notably, the composition of the growth medium. Adherence was enhanced by the addition of glucose or oleic acid to the growth medium and inhibited by serum. We have demonstrated a statistically significant association between the quantitative assessment of adherence to polystyrene tissue culture plates and clinical relevance. No such association was found when adherence was assessed by the qualitative adherence assay. Possible new approaches for assessing the clinical relevance of coagulase-negative staphylococcal isolates are discussed.
Polymerase chain reaction amplification of the constant and variable regions of the Bacteroides nodosus fimbrial gene
Authors: John G H , Carlson J O , Kimberling C V , Ellis R P
Published: January 2020

Serogrouping of Bacteroides nodosus is based on antigenic differences in fimbriae of the different New Zealand prototype strains. Because of the time needed to isolate and grow pure cultures of B. nodosus and the difficulty in distinguishing between different serogroups because of cross-agglutination, a new DNA-based diagnostic approach based on the fimbrial gene sequence of B. nodosus was developed. Published nucleotide sequences of the fimbrial genes for serogroups A, G, D, and H showed conservation at the 5' end, coding for the N terminus, and variability at the 3' end, coding for the C terminus. The polymerase chain reaction was used to amplify both the constant and variable regions of the fimbrial genes. Constant-region oligonucleotide primers were used to amplify a 100-base-pair fragment from the constant regions of the fimbrial genes of 10 New Zealand serogroups. Serogroup-specific oligonucleotide primers for serogroups A and H allowed amplification of a 282-base-pair fragment from serogroup A and a 363-base-pair fragment from serogroup H. Thus, amplification of the constant and variable regions of the fimbrial gene allows rapid detection and grouping of B. nodosus.
Detection of the aerolysin gene in Aeromonas hydrophila by the polymerase chain reaction
Authors: Pollard D R , Johnson W M , Lior H , Tyler S D , Rozee K R
Published: January 2020

Synthetic oligonucleotide primers were used in a polymerase chain reaction (PCR) technique to detect the gene for aerolysin in strains of Aeromonas hydrophila and to screen for identical genes in A. caviae, A. sobria, and A. veronii isolated from patients with diarrheal disease. Primers targeted a 209-bp fragment of the aer gene coding for the beta-hemolysin and detected template DNA only in the PCR using nucleic acid (NA) from hemolytic strains of A. hydrophila which were also cytotoxic to Vero and CHO cells and enterotoxic in suckling-mouse assays. PCR amplification of NA from hemolytic A. sobria or nonhemolytic A. hydrophila and A. caviae strains was consistently negative. Primer specificity was determined in the PCR by using NA extracted from 56 strains of bacteria, including hemolytic Escherichia coli and Listeria monocytogenes as well as several recognized enteric pathogens defined in terms of their toxigenicity. The detection limit for the aerolysin gene by PCR amplification was 1 ng of total NA. The PCR clearly identified aerolysin-producing strains of A. hydrophila and may have application as a species-specific virulence test because other hemolytic Aeromonas species tested were negative.
Rapid screening for B19 parvovirus DNA in clinical specimens with a digoxigenin-labeled DNA hybridization probe
Authors: Zerbini M , Musiani M , Venturoli S , Gallinella G , Gibellini D , Gentilomi G , La Placa M
Published: January 2020

A rapid dot blot hybridization assay for the detection of B19 parvovirus DNA in human sera was developed. Small portions of four serum samples were mixed, filtered onto a nylon membrane, and hybridized with a digoxigenin-labeled DNA probe; for each membrane, 380 serum samples could be tested. When a dot was positive by the hybridization assay, the four serum samples dotted together were separately tested to identify the sample positive for B19 DNA. A total of 10,150 serum samples submitted for viral serological and laboratory investigation with no specific requests for B19 testing were analyzed. Nine serum samples were positive for B19 DNA by dot blot hybridization assay, and the results were confirmed by electron microscopy. This method has proven to be reliable, economical in terms of time and costs, and useful for large-scale screening of clinical specimens, both for diagnostic work and for a source of antigen.
Herpes simplex virus antigen direct detection in standard virus transport medium by Du Pont Herpchek enzyme-linked immunosorbent assay
Authors: Verano L , Michalski F J
Published: January 2020

A commercial 5-h direct herpes simplex virus (HSV) antigen detection enzyme immunoassay kit (Du Pont Herpchek) was compared with a cell culture isolation system by using primary rabbit kidney and MRC-5 cells with 779 clinical specimens received in virus transport medium and with stock tissue culture preparations of HSV types 1 and 2. In the first study of 422 specimens from symptomatic patients, Herpchek detected 110 of 111 HSV-positive specimens (26.3% of all specimens), with a sensitivity of 99% and a specificity of 100%. In the second study of 357 specimens primarily from asymptomatic pregnant women, however, Herpchek detected 70 of 119 HSV-positive specimens (33% of all specimens), with a sensitivity of 58.8% and a specificity of 99.5%. Stock virus dilution experiments showed that Herpchek was 10 to 100 times less sensitive than culture. Herpchek was found to be an acceptable test for symptomatic patients, but for asymptomatic patients shedding a low titer of HSV it was not as sensitive and cell culture of Herpchek-negative specimens is recommended for such cases.
Identification of restriction fragment length polymorphisms in DNA from Mycobacterium paratuberculosis
Authors: Whipple D , Kapke P , Vary C
Published: January 2020

DNAs from 34 mycobactin-dependent isolates of Mycobacterium paratuberculosis and 1 isolate of M. paratuberculosis 18 were digested with four restriction endonucleases. Southern hybridization experiments were performed with a 32P-labeled oligonucleotide DNA probe derived from the sequence of IS900, an insertion sequence present in 15 to 20 copies per M. paratuberculosis chromosome. The probe hybridized with DNA from each of the mycobactin-dependent isolates, and restriction fragment length polymorphisms were detected among the isolates with each restriction endonuclease used. Restriction fragment length polymorphism analysis may permit identification of various strains of M. paratuberculosis, which has not been possible with other techniques.
Osteomyelitis and synovitis produced by Mycobacterium marinum in a fisherman
Authors: Clark R B , Spector H , Friedman D M , Oldrati K J , Young C L , Nelson S C
Published: January 2020

We report a case of osteomyelitis and synovitis produced by Mycobacterium marinum in the left index finger of a fisherman. A combination of surgical intervention and antimicrobial therapy with minocycline, rifampin, and ethambutol was efficacious.
Reproducibility of API Staph-Ident system identifications of coagulase-negative staphylococci isolated from blood
Authors: Overman T L , Overley J K
Published: January 2020

Fifty-five isolates of coagulase-negative staphylococci from blood were identified four times each by using the API Staph-Ident (API-SI) system to determine the identification reproducibility of the system. Identifications were determined by using both Version 1 and Version 2 of the API-SI Profile Index. The 75 to 100% reproducibility levels, including reproducible nonidentifications, were 98.2% for Version 1 and 96.4% for Version 2. Version 1 gave an identification for 92.5% of the isolates with a 75 to 100% reproducibility, while Version 2 gave an identification for 100% of the isolates with a 75 to 100% reproducibility. The reproducibility of the API-SI identifications of coagulase-negative staphylococci from blood was good.
Outbreak of Salmonella typhimurium infection traced to contaminated chocolate and caused by a strain lacking the 60-megadalton virulence plasmid
Authors: Kapperud G , Gustavsen S , Hellesnes I , Hansen A H , Lassen J , Hirn J , Jahkola M , Montenegro M A , Helmuth R
Published: January 2020

We describe an outbreak of Salmonella typhimurium infection, caused by contaminated chocolate produced by one Norwegian company, which occurred in Norway and Finland in 1987. A total of 349 bacteriologically verified cases were recorded in Norway, and 12 cases were recorded in Finland. There was a predominance of young children among the patients (median age, 6 years), many of whom developed acute hemorrhagic diarrhea. The outbreak strain exhibited a rare phage lysis pattern and a characteristic plasmid profile lacking the 60-MDa virulence-associated plasmid. DNA hybridization failed to demonstrate any DNA sequence homology between the outbreak strain and the virulence plasmid. The outbreak strain was nonlethal for orally infected mice. The finding of only less than or equal to 10 S. typhimurium cells per 100 g of chocolate in about 90% of the positive samples obtained from retail outlets suggested that an inoculum of fewer than 10 organisms may have been sufficient to cause symptomatic disease.
Infectious but nonpathogenic isolate of Borrelia burgdorferi
Authors: Anderson J F , Barthold S W , Magnarelli L A
Published: January 2020

We document for the first time an infectious but nonarthritogenic variant of Borrelia burgdorferi. Strain 25015, previously isolated from an Ixodes dammini larva collected in upstate New York, was infectious but failed to produce arthritis or carditis in laboratory rats and mice. By contrast, pathogenic strain N40 invariably caused arthritis. This nonarthritogenic variant, with proteins with molecular weights different from those of the standard B31 strain, was frequently isolated from normal joint tissues of experimentally infected rats. Outer surface proteins A and B of strain 25015 have molecular weights of about 32,500 and 35,500, respectively, in contrast to molecular weights of approximately 31,000 and 34,000, respectively, for outer surface proteins A and B of strains B31 and N40. A prominent low-molecular-weight protein of about 23,500 also characterizes strain 25015. Test animals infected for 30 to 60 days had relatively high antibody titers (greater than or equal to 1:1,280). The nonarthritogenic variant will be useful, along with pathogenic strains, in providing comparative insight into the pathogenesis of Lyme borreliosis. Homologous immunoblotting of sera from rats and mice inoculated with both the arthritogenic and nonarthritogenic strains revealed antibody reactivities to proteins of B. burgdorferi different from those revealed in the heterologous tests.