bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparison of microbiologic assay methods for hemodialysis fluids
Authors: Arduino M J , Bland L A , Aguero S M , Carson L , Ridgeway M , Favero M S
Published: January 2020

To help prevent pyrogenic reactions and bacteremia in hemodialysis patients, the Association for the Advancement of Medical Instrumentation and the Centers for Disease Control recommend microbiologic assay of hemodialysis fluids at least monthly. Five commercially available assay systems were evaluated by using the membrane filtration technique with standard methods agar and trypticase soy agar as the standards for comparison. Each assay system was challenged with dialysate and reverse-osmosis water from local dialysis centers, aqueous suspensions of eight laboratory strains of gram-negative bacilli and nontuberculous mycobacteria, and a mixed microbial flora inoculated into reverse-osmosis water and laboratory-prepared dialysate. Mean viable counts from triplicate samples were obtained after incubation at 37 degrees C for up to 72 h. The efficiency of recovery varied with the specific type of microbial challenge. The SPC water sampler (Millipore Corp., Bedford, Mass.) was the most consistent in obtaining the highest viable counts. Other commercial systems were comparable to each other in overall performance. All assay systems tested provided an acceptable balance between microbial recovery and required sampling time, equipment, and expertise.
Molecular epidemiology of rotaviruses associated with pediatric diarrhea in Bangkok, Thailand
Authors: Pipittajan P , Kasempimolporn S , Ikegami N , Akatani K , Wasi C , Sinarachatanant P
Published: January 2020

Rotavirus diarrhea in 453 pediatric patients (29.8% of 1,518) was studied in greater Bangkok during 1985 to 1987. The disease persisted all year, increasing in incidence from August to January (30 to 50%). Polyacrylamide gel electrophoresis of rotavirus RNA from these patients and from an additional 46 patients of a 1982 to 1983 epidemic revealed 26 electropherotypes, 4 with short (S) and 22 with long (L) RNA profiles. Of the analyzed specimens, 85.5% were L forms. Only one or a few electropherotypes predominated in each epidemic, whereas others appeared sporadically at low frequencies. Shifts in the predominant electropherotypes were observed in every epidemic. Of these, 126 strains were tested for subgroup and serotype by monoclonal antibody enzyme immunoassay. Serotype 4 prevailed from 1982 to 1983, while serotype 1 was encountered more frequently than serotypes 2 and 4 from 1985 to 1987. A complete correlation was found between the electrophoretic migration of segments 10 and 11 and the serologically defined subgroup specificity. Distinct electropherotypes occurred within the same serotype, and strains with the identical electropherotype always showed the same serotype specificity. No specific electropherotype or serotype correlated with patient age. In this study, atypical rotaviruses and mixed infections with different rotaviruses were identified.
Modification of RPMI 1640 for use in vitro immunological studies of host-parasite interactions in giardiasis
Authors: Guy R A , Bertrand S , Faubert G M
Published: January 2020

Incubation of trophozoites for 6 h in RPMI 1640 affected the viability of the parasite; however, RPMI 1640 supplemented with L-cysteine did not affect trophozoite viability, ability to grow when transferred to fresh TYI-S-33, or ability to infect gerbils. Similarly, incubation of murine spleen cells in modified medium did not affect the viability of the cells or proliferative responses to mitogens. RPMI 1640 supplemented with 11.4 mM L-cysteine is a suitable maintenance medium for in vitro studies in immunoparasitology because it maintains viability as well as some of the physiological functions of both trophozoites and lymphocytes.
Use of a new synthetic-peptide-derived monoclonal antibody to differentiate between vaccine and wild-type Venezuelan equine encephalomyelitis viruses
Authors: Roehrig J T , Bolin R A , Hunt A R , Woodward T M
Published: January 2020

We have prepared a murine monoclonal antibody (MAb) capable of distinguishing between wild-type Venezuelan equine encephalomyelitis (VEE) virus and the TC-83 vaccine derivative. This MAb, 1A2B-10, was derived from immunization with a synthetic peptide corresponding to the first 19 amino acids of the E2 glycoprotein of Trinidad donkey VEE virus. The MAb reacts with prototype viruses from all naturally occurring VEE subtypes except subtype 6 in an enzyme-linked immunosorbent assay. It does not react with TC-83 virus or members of the western and eastern equine encephalitis virus complex or with Semliki Forest virus. This antibody will also differentiate between TC-83 and Trinidad donkey VEE virus in indirect immunofluorescence assays with virus-infected Vero cells.
Comparison of two rapid methods for detection of respiratory syncytial virus (RSV) (Testpack RSV and ortho RSV ELISA) with direct immunofluorescence and virus isolation for the diagnosis of pediatric RSV infection
Authors: Thomas E E , Book L E
Published: January 2020

The ability of two commercial immunoassays to detect respiratory syncytial virus (RSV) in respiratory specimens was evaluated as follows: 152 specimens were tested by TestPack RSV (Abbott), and 72 were tested by Ortho RSV ELISA (Ortho). Test outcomes were compared with those of virus isolation alone, direct immunofluorescence assay (DFA) alone, or virus isolation and/or DFA. TestPack RSV versus virus isolation showed 91% sensitivity, 96% specificity, 93% positive predictive value (PPV), and 95% negative predictive value (NPV). TestPack RSV versus DFA showed 89% sensitivity, 97% specificity, 96% PPV, and 93% NPV. When TestPack RSV performance was compared with that of virus isolation and DFA, the sensitivity was 87% and the specificity was 100%. Ortho RSV ELISA versus virus isolation showed 88% sensitivity, 87% specificity, 79% PPV, and 93% NPV. Ortho RSV ELISA versus DFA showed 91% sensitivity, 88% specificity, 81% PPV and 95% NPV. When Ortho RSV ELISA performance was compared with that of virus isolation and DFA, the sensitivity was 86%, the specificity was 89%, the PPV was 86%, and the NPV was 89%. The accuracy of the TestPack RSV in combination with ease of performance and no need for specialized equipment or special skills make it an attractive alternative to DFA for rapid direct detection of RSV.
Detection of Giardia lamblia antigen in children living in a Peruvian periurban shantytown (Pueblo Joven)
Authors: Vidal M F , Gilman R H , Ungar B L , Verástegui M R , Benel A C , Marquis G , Penny M , Lanata C , Miranda E
Published: January 2020

Stool microscopy and an enzyme-linked immunosorbent assay (ELISA) for Giardia lamblia antigen detection were compared for detecting G. lamblia in 30 Peruvian infants. Of 1,131 fecal specimens, G. lamblia was detected by ELISA alone in 44, by microscopy alone in 17, and by both methods in 91. In another group of 17 children negative for G. lamblia by stool microscopy, 6 had G. lamblia detected by ELISA or duodenal aspiration: 2 only by ELISA, 1 only by duodenal aspirate examination, and 3 by both examinations. The ELISA is useful for the detection of G. lamblia in fecal specimens but compared to stool microscopy does not significantly increase the detection of cases.
Detection of Helicobacter pylori by using the polymerase chain reaction
Authors: Valentine J L , Arthur R R , Mobley H L , Dick J D
Published: January 2020

A 1.9-kb cloned fragment of chromosomal DNA randomly selected from a Helicobacter pylori cloned library was evaluated as a potential probe. The probe detected 19 of 19 H. pylori strains and yielded a specificity of 98.7% when tested against 306 other bacterial strains representing 32 different species. False-positive results with non-H. pylori strains were due to the presence of contaminating vector sequences. A polymerase chain reaction (PCR) assay was developed by using 20-base oligonucleotide primers homologous to a portion of the 1.9-kb fragment. The PCR assay amplified a 203-nucleotide-pair product which was analyzed by agarose gel electrophoresis and Southern hybridization by using a third 20-base 32P-labeled oligonucleotide complementary to a region of DNA between the primers. The PCR assay was 100% sensitive, detecting all 35 H. pylori strains tested, and did not amplify sequences in several closely related species. The assay was sensitive for as little as one copy of the cloned plasmid DNA or 100 H. pylori bacterial cells. To evaluate the PCR assay for clinical samples, gastric biopsy and aspirate specimens were tested by PCR, and the results were compared with those of microbiologic culture and histologic examination. In fresh biopsy specimens, H. pylori sequences were detected by PCR in 13 of 14 (93%) positive tissues and 0 of 19 negative tissues. In gastric aspirate specimens, 11 of 13 (85%) positive tissues were positive by PCR. H. pylori DNA was detected in 1 of 14 aspirate specimens negative by culture, histology, and PCR of the accompanying biopsy tissue. PCR is a rapid, accurate, and sensitive method for the detection of H. pylori.
Evaluation of the Baxter-MicroScan 4-hour enzyme-based yeast identification system
Authors: Land G A , Salkin I F , el-Zaatari M , McGinnis M R , Hashem G
Published: January 2020

A new 4-h Yeast Identification Panel (YIP; Baxter-MicroScan, W. Sacramento, Calif.) was compared with the API 20C Yeast Identification System (Analytab Products, Inc., Plainview, N.Y.) in the identification of recent clinical yeast isolates. The YIP had a 94% correlation (288 of 306) in identifying 22 species within the genera Candida, Hansenula, Pichia, Rhodotorula, Saccharomyces, and Torulopsis. Correlation dropped to 65% for those species within the genera of slower growing yeasts, i.e., Blastoschizomyces spp., Crpytococcus spp., Geotrichum spp., Hyphopichia spp., Phaeococcomyces spp., Prototheca spp., and Trichosporon spp. Overall correlation with the API 20C was 92% (365 of 401) for those taxa included in the data base and 85% (373 of 437) for all yeasts encountered in the study. There were 36 (8.2%) discrepant identifications, which were due in part to the limited data base. Expansion of the data base plus the easy inoculation, reading, and rapid results of the YIP should make it an excellent method for yeast identification.
Chronic conjunctivitis caused by oral anaerobes and effectively treated with systemic metronidazole plus amoxicillin
Authors: van Winkelhoff A J , Abbas F , Pavicic M J , de Graaff J
Published: January 2020

In this study, we report on a case of refractory, unilateral anaerobic conjunctivitis. The predominant anaerobic flora consisted of Prevotella intermedia (formerly Bacteroides intermedius) and Peptostreptococcus micros. By using the technique of restriction endonuclease fingerprinting of genomic DNA, it was shown that the P. intermedia likely originated from the oral cavity. Topically applied antibiotics had failed to suppress the infection in the past. Successful treatment was achieved after systemic administration of metronidazole plus amoxicillin.
Comparison of in vitro culture and polymerase chain reaction for detection of Borrelia burgdorferi in tissue from experimentally infected animals
Authors: Lebech A M , Hindersson P , Vuust J , Hansen K
Published: January 2020

A polymerase chain reaction (PCR) was developed for identification of Borrelia burgdorferi in biological specimens. The diagnostic efficiency was compared with that of in vitro culture. A primer set specifying a 791-bp DNA fragment of the B. burgdorferi B31 flagellin gene was used. Amplified DNA sequences were analyzed by agarose gel electrophoresis, and the identity of amplified DNA was confirmed by restriction enzyme cleavage and Southern blot hybridization with a 32P-labeled probe. By using purified B. burgdorferi DNA, the detection limit of the assay was approximately 0.002 pg of DNA, corresponding to one copy of the B. burgdorferi genome. By using in vitro-cultivated B. burgdorferi without prior DNA purification as the template DNA, 2 to 20 organisms could be detected. A 791-bp DNA fragment was amplified from all of 18 different B. burgdorferi strains tested, as well as from Borrelia hermsii and Borrelia anserina but not from Treponema pallidum. The efficacy of the PCR assay was evaluated on spleen, renal, and urinary bladder tissue specimens from eight experimentally infected gerbils. Specimens from the same organs were cultured in BSK medium in parallel. Of 24 organs, 21 (88%) were PCR positive and 17 (71%) were culture positive. All culture-positive specimens were also PCR positive. Compared with B. burgdorferi cultivation, PCR had at least a comparable diagnostic sensitivity, it was less laborious, and results were available within 1 to 2 days.
Identification of Anaplasma marginale long-term carrier cattle by detection of serum antibody to isolated MSP-3
Authors: McGuire T C , Davis W C , Brassfield A L , McElwain T F , Palmer G H
Published: January 2020

Rapid and accurate detection of Anaplasma marginale-infected cattle would enhance anaplasmosis control procedures and evaluation of vaccines. Current tests based on detection of antibodies in serum are not widely used for several reasons, including the occurrence of either false-positive or false-negative results. We evaluated binding of antibodies in serum to a subunit antigen isolated from A. marginale initial bodies--major surface protein 3 (MSP-3). MSP-3 was detected in lysates of eight geographically different isolates of A. marginale and purified by affinity chromatography with monoclonal antibody AmG75C2. Antibodies from cattle infected with any of five geographically different isolates of A. marginale reacted in immunoblots with MSP-3. Sera from uninfected cattle and cattle infected with another rickettsial organism and two hemoprotozoal organisms failed to react with MSP-3. Six carrier cattle infected with the Florida isolate of A. marginale had antibody titers to MSP-3 ranging from 10(3) to 10(6) during a 5-year evaluation period. Since specific antibodies to isolated MSP-3 persist in high titers in long-term carrier cattle sera and MSP-3 is common among A. marginale isolates, it is recommended as a subunit antigen for an anaplasmosis test.
Rapid method to extract DNA from Cryptococcus neoformans
Authors: Varma A , Kwon-Chung K J
Published: January 2020

A rapid and easy method for the extraction of total cellular DNA from Cryptococcus neoformans is described. This procedure modifies and considerably simplifies previously reported methods. Numerous steps were either eliminated or replaced, including preincubations with cell wall permeability agents such as beta-mercaptoethanol and dithiothreitol. The commercially available enzyme preparation Novozyme 234 was found to contain a potent concentration of DNases which actively degrade DNA. Degradation and loss of DNA was prevented by maintaining a high concentration of EDTA in the lysing solution. This procedure resulted in high yields (150 to 200 micrograms of DNA from 100 ml of culture) of good-quality (undegraded), high-molecular-weight DNA which was readily digested by restriction endonucleases, making it suitable for use in various molecular applications.
Pulsed field electrophoresis of genomic restriction fragments for the detection of nosocomial Legionella pneumophila in hospital water supplies
Authors: Ott M , Bender L , Marre R , Hacker J
Published: January 2020

Ten Legionella pneumophila strains isolated from different sources were analyzed according to their restriction fragment patterns obtained by cleavage of genomic DNA with NotI and SfiI and separation by pulsed field electrophoresis. Three L. pneumophila isolates from a nosocomial outbreak in Lübeck (Germany) and three other L. pneumophila strains independently isolated from a water tap located in the care unit where the patients were hospitalized exhibited identical restriction fragment profiles. Therefore, we concluded that these environmental specimens were the source of the Legionnaires disease. Another two isolates from patients and two strains from the environment, all unrelated to the outbreak described, showed different cleavage patterns.
Prevalence of antibody to Chlamydia pneumoniae TWAR in japan
Authors: Kanamoto Y , Ouchi K , Mizui M , Ushio M , Usui T
Published: January 2020

Chlamydia pneumoniae TWAR is a newly recognized Chlamydia species that is a pathogen of respiratory tract infection. To clarify the endemic status of C. pneumoniae in Japan, we evaluated the incidence of C. pneumoniae antibody in 1,330 serum samples (660 from outpatients, 600 from normal individuals, and 70 from cord blood). The antibody titer was determined by a microimmunofluorescence test by using the elementary body of C. pneumoniae TW-183 as the antigen. Immunoglobulin G antibody titers of 1:32 or higher were regarded as evidence of past infection. The detection rate of C. pneumoniae antibody rapidly increased in subjects between the ages of 4 and 7 years, reached 44% in subjects between the ages of 8 and 11 years, and was about 50% in older subjects. The rate did not differ between healthy subjects and outpatients. These results suggest that C. pneumoniae infection is highly endemic in Japan as it is in Western countries. However, the antibody prevalence was high in the low age groups in Japan compared with that in Western countries.
Egg yolk emulsion agar, a new medium for the cultivation of Helicobacter pylori
Authors: Westblom T U , Madan E , Midkiff B R
Published: January 2020

We developed a new agar, egg yolk emulsion (EYE) agar, for cultivation of Helicobacter pylori. EYE agar contains Columbia agar base (Oxoid), 10% EYE (Oxoid), 1% IsoVitaleX (BBL), and 40 mg of Triphenyleteraxolium chloride (Sigma) per liter. We compared EYE agar with the following agars: (i) brain heart infusion agar-7% horse blood-1% IsoVitaleX (GDW agar; C. S. Goodwin, E. D. Blincow, J. R. Warren, T. E. Waters, C. R. Sanderson, and L. Easton, J. Clin. Pathol. 38:1127-1131, 1985), (ii) brain heart infusion agar-10% horse serum-0.2% charcoal-1% yeast extract-40 mg of triphenyltetrazolium chloride per liter (GLU agar; Y. Glupczynski, M. Labbe, and F. Thiabaumont, p. 3-6, in F. Megraud and H. Lamouliatte, ed., Gastroduodenal Pathology and Campylobacter pylori, 1989), (iii) Columbia agar with 7% lysed horse blood (D&M agar; J. C. Dent and C. A. M. McNulty, Eur. J. Clin. Microbiol. Infect. Dis. 7:555-558, 1988), and (iv) brain heart infusion agar-10% EYE-1% IsoVitaleX (BHIE agar). H. pylori CFU counts, expressed as average percentages of maximum growth, were as follows: EYE agar, 96; GDW agar. 76; BHIE agar, 57; D&M agar, 52; and GLU agar, 23. Colony counts for EYE agar were significantly higher than for GDW agar (P = 0.027), BHIE agar (P = 0.005), D&M agar (P = 0.0001), and GLU agar (P less than 0.0001). EYE agar also had higher CFU counts than two commercial chocolate media; the EYE agar count was 80%, versus 33% for BBL chocolate medium and 63% for Remel chocolate medium.
Serratia marcescens meningitis
Authors: Theccanat G , Hirschfield L , Isenberg H
Published: January 2020

A case of Serratia marcescens meningitis in a 66-year-old man is reported. The infection occurred 4 weeks after apparently successful otic surgery, and a nidus of infection in the middle ear was established at autopsy. This is the second case of S. marcescens meningitis following ear surgery reported in the English-language literature.
Inguinal lymphadenitis associated with Capnocytophaga bacilli
Authors: Johnson C C , Poupard J
Published: January 2020

Capnocytophaga organisms are capnophilic, gram-negative bacilli that have been associated with infections deriving from the flora of the oropharynx. We report a case of inguinal adenitis caused by Capnocytophaga species that probably represents sexual transmission of the pathogen.
Serratia fonticola isolated from a leg abscess
Authors: Bollet C , Gainnier M , Sainty J M , Orhesser P , De Micco P
Published: January 2020

Serratia fonticola was isolated from a very large leg abscess in a patient following an accident. This is the first documented human infection due to S. fonticola.
Evaluation of a visual, rapid, membrane enzyme immunoassay for the detection of herpes simplex virus antigen
Authors: Zimmerman S J , Moses E , Sofat N , Bartholomew W R , Amsterdam D
Published: January 2020

We evaluated a 12-min, direct, monoclonal antibody-based enzyme immunoassay (EIA) (SureCell; Kodak, Rochester, N.Y.) which aids in the detection of herpes simplex virus infection; the assay system is also approved for culture confirmation. The test was evaluated from direct clinical samples and compared with conventional culture methodology by using a single swab. A total of 265 specimens from 180 female cervical-urogenital sites, 62 male urogenital sites, 4 rectal sites, 3 skin sites, 6 oral sites, and 10 colposcopy sites were collected on Dacron or cotton swabs and placed in viral transport medium (VTM). Within 6 h of receipt, 0.2 ml of the vortexed VTM was inoculated into each of two replicate cell cultures. Cell monolayers were observed daily for ten days, and cytopathic effect was confirmed by using an indirect immunoperoxidase reagent. The procedure for the SureCell assay conformed to the manufacturer's recommendations. When conventional culture was compared with EIA results, the overall sensitivity, specificity, positive predictive value, negative predictive value, and agreement were 64.4, 98.9, 96.7, 84.4, and 87.2%, respectively. Variables affecting the EIA sensitivity are the stage of the lesion and conventional culture methodologies. A review of culture results for 32 EIA false-negative tests indicated that 15 were detected after 48 h of incubation. Cytopathic effect observed at 48-, 72-, and 96-h cutoffs altered the sensitivity for the EIA. To ensure detection of SureCell herpes simplex virus-negative specimens, it is recommended that an unused aliquot of VTM be tested in cell culture.
First isolation of Yersinia enterocolitica serotype O:8 in Japan
Authors: Ichinohe H , Yoshioka M , Fukushima H , Kaneko S , Maruyama T
Published: January 2020

A 4-year-old Japanese boy was infected with Yersinia enterocolitica serotype O:8, H:befiv, biotype 1B, phage type Xz, restriction endonuclease analysis of plasmid DNA type B, restriction endonuclease analysis of chromosomal DNA type 8. He presented with acute gastroenteritis with elevated body temperature (40 degrees C), rigor, and shivers. The diagnosis was septicemia. This is apparently the first report of this serotype from a human infection outside North America.
Cloned DNA probes specific for the intracellular Campylobacter-like organism of porcine proliferative enteritis
Authors: Gebhart C J , Lin G F , McOrist S M , Lawson G H , Murtaugh M P
Published: January 2020

Although the etiology of porcine proliferative enteritis is not understood, the consistent presence of intracellular Campylobacter-like organisms (CLOs) in proliferating pig intestinal epithelial cells suggests that the organism is involved in the disease process. In order to obtain information about this organism, we generated and characterized specific DNA probes to the intracellular CLO which was purified without culturing. Intracellular CLOs were isolated from mucosa by homogenization, filtration, and absorption to wheat germ agglutinin-Sepharose. The DNA was purified, and a CLO genomic library was constructed. The specificity of recombinant plasmids was confirmed by both dot blot hybridization and Southern analysis of normal and diseased mucosa, as well as of a variety of Campylobacter species. Several of the CLO-specific probes hybridized with porcine mucosa obtained from pigs with proliferative enteritis but not with nondiseased mucosa. The probes hybridized equally with mucosa or DNA obtained from each of the two clinical forms of proliferative enteritis, i.e., proliferative hemorrhagic enteropathy and porcine intestinal adenomatosis. The CLO-specific probes failed to hybridize with any of the commonly isolated porcine Campylobacter species, including Campylobacter hyointestinalis, C. mucosalis, and C. coli. Therefore, the intracellular CLO of porcine proliferative enteritis may be an as yet unidentified or uncultured species.
Enzyme-linked immunosorbent assay for products of the 60-megadalton plasmid of Escherichia coli serotype O157:H7
Authors: Toth I , Barrett T J , Cohen M L , Rumschlag H S , Green J H , Wachsmuth I K
Published: January 2020

Eighty strains of pathogenic Escherichia coli, representing each of the major diarrheal disease-causing groups, were examined by direct enzyme-linked immunosorbent assay (ELISA) for the presence of proteins associated with a 60-MDa plasmid from E. coli serotype O157:H7. Antiserum specific for plasmid-encoded proteins was prepared by immunizing a rabbit with a wild-type E. coli O157:H7 strain (strain 7785) and absorbing the serum with a plasmid-cured derivative (strain 2-45). Use of this antiserum in Western immunoblot analysis detected two proteins of 82 and 92 kDa in strain 7785 but not in strain 2-45. All 16 wild-type E. coli O157:H7 strains and all 10 Shiga-like toxin (SLT)-producing E. coli strains of serotypes other than O157 were ELISA positive. Thirteen of 14 enterotoxigenic and all of 24 enteroinvasive E. coli strains were ELISA negative, as were all of 16 E. coli strains isolated from healthy persons. Of 16 traditional enteropathogenic E. coli (EPEC) serotypes, 10 were ELISA positive, including 10 of 12 strains carrying the EPEC adherence factor gene. Absorption of the serum with an EPEC adherence factor-positive EPEC eliminated EPEC reactivity. This study demonstrates that two plasmid-mediated proteins are common to E. coli O157:H7 strains and to SLT-producing strains of other serotypes. Detection of these proteins by ELISA provides a sensitive and specific screening test for identifying SLT-producing E. coli of both O157 and non-O157 serotypes. Identification of the cross-reactive proteins found in EPEC could provide the basis for a single assay to detect both EPEC and SLT-producing E. coli.
Aspergillus fumigatus contamination of lymphokine-activated killer cells infused into cancer patients
Authors: Arnow P M , Houchins S G , Richards J M , Chudy R
Published: January 2020

Lymphokine-activated killer (LAK) cells, prepared by incubating autologous lymphocytes in cell culture medium with interleukin-2, selectively lyse tumor cells and are effective immunotherapy of some cancers. During a 3-month period, two patients at our center were infused with LAK cells subsequently found to have been contaminated by Aspergillus fumigatus. Each case was investigated by obtaining environmental cultures and assessing aseptic practices during LAK cell preparation. Investigation of the first case demonstrated a malfunction of the laminar air flow hood, under which interleukin-2 and the patient's lymphocytes had been added to cell culture medium, and showed heavy A. fumigatus contamination of the hood, adjacent countertop, and cell culture incubator. Despite repair of the laminar air flow hood and cleaning of the laboratory, a second case occurred, and cultures at that time implicated the humidified cell culture incubators as the source of A. fumigatus. Following incubator sterilization and removal of the humidification apparatus from the incubators, weekly environmental cultures in the LAK cell laboratory were negative, and none of the LAK cell cultures from the 20 patients treated during the ensuing 15 months grew A. fumigatus. Our findings show that growth of fungi in humidified incubators, which previously has caused contamination problems in tissue culture and clinical microbiology laboratories, can result in patient infections when humidified incubators are used to prepare cells for reinfusion.
Identification of dye-sensitive strains of Brucella melitensis
Authors: Corbel M J
Published: January 2020

Strains of Brucella melitensis biovars 1, 2, and 3 isolated in India, Italy, Kuwait, Saudi Arabia, the Federal Republic of Germany, and Zimbabwe were inhibited by thionin (20 micrograms/ml) but not by basic fuchsin (20 micrograms/ml) or safranin O (200 micrograms/ml). In other respects the strains were typical of their species and biovars. The existence of dye-sensitive strains of B. melitensis suggests that the present system of classification requires modification.
Comparison of immunofluorescence and enzyme immunoassay for detection of measles-specific immunoglobulin M antibody
Authors: Rossier E , Miller H , McCulloch B , Sullivan L , Ward K
Published: January 2020

During a measles outbreak, 283 serum specimens from 221 suspected cases of measles were tested by immunofluorescence and enzyme immunoassay for the presence of measles-specific immunoglobulin M (IgM) antibodies by using commercially available reagents. There was 97% agreement between the two assays; thus, the choice of the method for diagnostic testing is a matter of convenience and experience. In all 62 cases of measles from which a single blood sample was available, measles IgM-specific antibodies were detectable by both methods. Fifty percent of the 62 cases were positive within 3 days after onset of the rash. This increased to 91% 10 days after onset of the rash.
Survival of feline mycoplasmas in urine
Authors: Brown M B , Stoll M , Maxwell J , Senior D F
Published: January 2020

The effects of length of incubation and urine osmolality on the survival of feline mycoplasmas and ureaplasmas and representative gram-positive and gram-negative bacteria in synthetic urine which approximated the osmolality of normal cat urine were investigated. Both Escherichia coli and Staphylococcus aureus withstood the effects of increasing osmotic pressure. In the most concentrated urine, significant decreases (P less than 0.001) in CFU were observed for E. coli at exposure times of 30 min and longer. S. aureus was not affected by longer exposure or increased osmotic strength. Both Mycoplasma felis and Mycoplasma gateae were affected adversely by longer exposure times and high osmotic strength (P less than 0.001). A Ureaplasma sp. was not adversely affected except at very high (greater than or equal to 2,980 mosM) osmotic strengths or after prolonged incubation (120 min) at relatively high (1,976 mosM) osmotic strengths (P less than 0.001). The failure of both M. felis and M. gateae to survive under osmotic conditions present in normal feline urine suggests that it is unlikely that these mycoplasmas are involved in urinary disorders in cats.
Kingella kingae intervertebral disk infection
Authors: Amir J , Shockelford P G
Published: January 2020

Disk inflammation in children is believed to result from infection, and Staphylococcus aureus is reported to be the organism most commonly isolated from cases of intervertebral disk infection. A case of disk inflammation caused by the unusual pathogen Kingella kingae is described. The antibiotic susceptibility of other K. kingae isolates and the clinical features of 11 other previously reported cases of disk infection caused by this microorganism are reviewed.
Aeromonas spp. and their association with human diarrheal disease
Authors: Deodhar L P , Saraswathi K , Varudkar A
Published: January 2020

Between January 1988 and December 1989 Aeromonas species were isolated from 45 (1.8%) of 2,480 patients with acute gastroenteritis. No other bacterial enteric pathogens were found in any of these 45 patients. Of the 45 Aeromonas isolates, 35 strains (77.8%) were Aeromonas hydrophila, 7 (15.5%) were Aeromonas sobria, and 3 (6.7%) were Aeromonas caviae. Most of the patients were under 5 years of age. No bacterial enteric pathogens, including Aeromonas species, were isolated from 512 age- and sex-matched control subjects. Examination of the Aeromonas isolates for exotoxin production (enterotoxin and hemolysin) indicated that all strains, irrespective of species, were enterotoxin positive (rabbit ileal loop model) and hemolysin positive (rabbit erythrocyte model). These results suggest that Aeromonas species are potential enteric pathogens in our geographical region.
Neutralizing antibodies to heterologous animal rotavirus serotypes 5, 6, 7, and 10 in sera from Ecuadorian children
Authors: Brüssow H , Offit P A , Sidoti J
Published: January 2020

Serum samples from 870 Ecuadorian children who underwent natural rotavirus exposure were tested for neutralizing serum antibody to heterologous animal rotavirus (RV) serotypes. Six percent of the sera neutralized porcine RV OSU (serotype 5), 10% neutralized bovine RV NCDV (serotype 6), 4% neutralized avian RV Ch-2 (serotype 7), and 8% neutralized bovine RV V1005 (serotype 10). Neutralization was defined as a 90% reduction in infectious virus at a 1:100 serum dilution. The prevalence of antibody to all four heterotypic viruses increased with the age of the children and the number of human RV serotypes neutralized, but prevalences did not differ significantly between children from rural and urban areas of Ecuador. No serum sample that specifically neutralized bovine RV NCDV was identified. We inferred from the seroepidemiological analysis that human RVs contain immunorecessive neutralization epitopes that can stimulate cross-neutralizing antibody to heterotypic animal RVs. This occurs increasingly with age and with the number of human serotypes recognized by a child's neutralizing antibody. Thus, it appears that a broadened immune response to the heterotypic strains occurs with repetitive RV infections.
Controlled evaluation of BACTEC Plus 26 and Roche Septi-Chek aerobic blood culture bottles
Authors: Weinstein M P , Mirrett S , Wilson M L , Harrell L J , Stratton C W , Reller L B
Published: January 2020

Blood culture bottles that accept up to 10 ml of blood have recently been introduced for the BACTEC nonradiometric blood culture system. The new formulation, designated BACTEC Plus, contains 25 ml of tryptic soy broth, 0.05% sodium polyanetholesulfonate, and mixed resins. In a collaborative study conducted at three university hospitals, we evaluated the BACTEC Plus 26 (BP26) aerobic bottle and the Roche Septi-Chek aerobic bottle with its agar slide paddle in 5,293 paired blood cultures. Significantly more microorganisms (P less than 0.001), especially Staphylococcus aureus (P less than 0.001), Staphylococcus epidermidis (P less than 0.01), enterococci (P less than 0.005), and members of the family Enterobacteriaceae (P less than 0.005), were detected by the BP26 bottle. When both bottles detected growth, BP26 did so earlier (P less than 0.001). In particular, S. epidermidis (P less than 0.001), streptococci (P less than 0.005), enterococci (P less than 0.05), and members of the family Enterobacteriaceae (P less than 0.001) were detected earlier by the BP26 bottle. We conclude that the BP26 bottle provides a yield and speed of detection of microorganisms superior to those of the Roche Septi-Chek aerobic blood culture bottle.
Analysis of serotypes and electropherotypes of equine rotaviruses isolated in the United States
Authors: Hardy M E , Woode G N , Xu Z C , Williams J D , Conner M E , Dwyer R M , Powell D G
Published: January 2020

Equine group A rotaviruses isolated over a 10-year period in New York State, New Jersey, Kentucky, and Texas were compared serotypically and electropherotypically. All isolates were determined to be serotype 3 by reaction with hyperimmune antiserum to the serotype 3 H-2 strain of equine rotavirus. All displayed RNA electrophoretic migration patterns related to that of the H-2 strain but distinct from that of serotype 5 strain H-1. A serologic survey of 184 mares in Kentucky, which was done to determine the incidence of H-1 and H-2 infections, showed geometric mean serum neutralizing titers to the H-2 strain of equine rotavirus to be significantly higher than those to the H-1 strain. These data suggest that the serotype 3 H-2 strain is the dominant equine rotavirus in Kentucky and perhaps elsewhere in the United States. We were unable to produce confirmational evidence that the H-1 strain occurs as a natural infection in the United States.
Application of a polymerase chain reaction for the detection of Mycobacterium leprae in skin tissues
Authors: de Wit M Y , Faber W R , Krieg S R , Douglas J T , Lucas S B , Montreewasuwat N , Pattyn S R , Hussain R , Ponnighaus J M , Hartskeerl R A
Published: January 2020

The polymerase chain reaction (PCR) based on the selective amplification of a 530-bp fragment of the gene encoding the proline-rich antigen of Mycobacterium leprae was applied on sections of fixed or frozen biopsy samples from leprosy patients. A simple procedure for the extraction of DNA from M. leprae in clinical specimens that provided suitable template DNA for amplification was developed. When PCR was applied on frozen sections, positive amplification in samples from all untreated acid-fast bacillus (AFB)-positive patients and in samples from 56% of the untreated AFB-negative patients could be detected, while biopsy samples from patients with skin diseases other than leprosy were all PCR negative. With neutral Formalin-fixed biopsy samples, positive amplification in 92% of the samples from untreated AFB-positive patients and in 61% of the samples from untreated AFB-negative patients could be detected by PCR. Biopsy samples exposed to mercuric chloride or nonbuffered formaldehyde containing fixatives were not suitable for application of PCR. This PCR holds promise as a tool for studies on M. leprae infection.
Molecular and epidemiological study of Salmonella clinical isolates
Authors: Rivera M J , Rivera N , Castillo J , Rubio M C , Gómez-Lus R
Published: January 2020

A survey of Salmonella infections was carried out over a 1-year period in the rural community covered by the Hospital Reina Sofía (Tudela, Spain). The 255 strains that were collected were studied by serotyping, antimicrobial resistance, and plasmid profile analysis. The predominant serotype was S. enteritidis (85.90%), followed by S. typhimurium (7.06%) and S. virchow (2.36%). Only 7.84% of the strains were resistant to antimicrobial agents. The most common resistance was to beta-lactam antibiotics. This resistance was due to the presence of one of two types of beta-lactamases, TEM-1 or TEM-2. Resistance to kanamycin was associated with the synthesis of a 3'-O-phosphotransferase. The resistance to streptomycin and chloramphenicol was either not enzymatic or was due to a 3"-O-phosphotransferase and a chloramphenicol acetyltransferase, respectively. Analysis of total plasmid DNA content revealed the presence of plasmids in 96.08% of the isolates. According to their plasmid profile, the strains could be classified into different groups. The three main groups, which accounted for 50.19, 20.78, and 4.70% of the isolates, respectively, corresponded to the antimicrobial-susceptible S. enteritidis serotype. These results suggested that plasmid profile analysis in conjunction with antimicrobial resistance determination can be useful for subtyping resistant Salmonella isolates.
Detection of flagella in 278 Legionella strains by latex reagent sensitized with antiflagellum immunoglobulins
Authors: Bornstein N , Marmet D , Dumaine M H , Surgot M , Fleurette J
Published: January 2020

To determine whether all Legionella species show common flagellum antigen properties, we developed a reagent using latex beads sensitized with flagellin-specific immunoglobulins that could be used in a simple and rapid agglutination reaction to identify Legionella colonies. A total of 278 strains (68 Legionella reference strains and 210 patient and environmental isolates) were tested. The results were compared with those obtained by a direct immunofluorescence assay using an antiflagellum serum and by morphological observations by electron microscopy. The immunological methods based on the use of a flagellum-specific serum have confirmed the presence of a common flagellum antigen for all Legionella species described to date. Flagella were detected for all the legionellae studied except four species: L. oakridgensis, confirmed as a nonflagellate species; L. brunensis; L. cincinnatiensis; and L. longbeachae serogroup 1. However, we noted a remarkable variability in flagellum expression, of greater or lesser degree, according to the species and their origin. A combination of all three methods of flagellum detection revealed that 86.3% of Legionella strains studied were flagellate. The latex test identified 89.6% of these strains, 97.5% of L. pneumophila, and 100% of L. pneumophila serogroup 1.
Comparison of restriction enzyme analysis and pulsed-field gradient gel electrophoresis as typing systems for Candida albicans
Authors: Vazquez J A , Beckley A , Sobel J D , Zervos M J
Published: January 2020

Candida species are an important cause of infection in immunocompromised hosts and the leading cause of nosocomial fungal infections. Study of the epidemiology of Candida infection has been difficult because of lack of a reliable typing system. We describe a typing system utilizing contour-clamped homogeneous electric fields (CHEF), which is a modified version of pulsed-field gradient gel electrophoresis, and compared it with restriction enzyme analysis (REA) of genomic DNA. The study was done with 35 Candida albicans clinical isolates from separate patients. CHEF and REA were performed on each isolate, and the patterns were compared. The REA procedure revealed 17 strain types while the CHEF procedure was able to distinguish 23 strain types of C. albicans. The CHEF technique yields unique patterns of chromosomal bands that can be used to distinguish clinical isolates and demonstrates greater sensitivity than REA.
Detection of transcripts of human papillomaviruses 16 and 18 in cancer-derived cell lines and cervical biopsies by enzyme immunoassay for DNA-RNA hybrids following solution hybridization
Authors: Coutlée F , Shah K V , Rader J S , Currie J L , Viscidi R P
Published: January 2020

mRNAs of human papillomaviruses (HPV) 16 and 18 were detected in cancer-derived cell lines and genital tract biopsy specimens by a novel hybridization assay. Biotinylated whole genomic HPV DNA probes were hybridized in solution to extracted total nucleic acids. Hybrids between the labeled probes and RNA transcripts were captured on a microplate coated with an antibiotin antibody. Bound hybrids were incubated with a beta-galactosidase-labeled monoclonal antibody to DNA-RNA hybrids and measured by the addition of a fluorogenic substrate. HPV 18 and HPV 16 mRNAs were detected in nucleic acids from 2.3 x 10(3) HeLa cells and 10(4) SiHa cells, respectively. The specificity of the assay for mRNA was demonstrated by the low reactivity of nucleic acids from SiHa cells after treatment with T1 RNase and by the selective reactivity of cellular nucleic acids which bound to an oligo(dT) column. With HPV 16 subgenomic probes, E6-E7 transcripts but not L1-L2 transcripts were detected in SiHa cells. Tests of 58 biopsy specimens from 31 patients showed that the detection of HPV 16 and HPV 18 transcripts in tissue specimens was feasible. Analysis of biopsy specimens with subgenomic probes revealed HPV 16 E6-E7 transcripts in all specimens that reacted with the whole genomic probe, while L1-L2 transcripts were found infrequently.
Production of species-specific murine monoclonal antibodies against Cryptococcus neoformans which recognize a noncapsular exoantigen
Authors: Hamilton A J , Bartholomew M A , Figueroa J , Fenelon L E , Hay R J
Published: January 2020

Three monoclonal antibodies (MAbs), designated 7C5, 7C9, and 5G8, against a cytoplasmic antigen of Cryptococcus neoformans were produced. MAbs 7C5 and 7C9 recognize culture filtrate antigen (exoantigen) of both encapsulated and nonencapsulated isolates of this pathogen, which suggests that they do not recognize capsular polysaccharide material. This is supported by immunofluorescence data which show reactivity of all 3 MAbs to cytoplasm and cell membranes only. MAb 7C9 also recognized C. neoformans var. gattii antigens but no other fungal pathogens tested in an enzyme-linked immunosorbent assay, while 7C5 and 5G8 recognized antigens of the cross-reactive pathogen Trichosporon beigelii but did not recognize either C. neoformans var. gattii isolates or any other fungal antigens. By Western blot (immunoblot), 7C9 detected antigen at 110 to 120, 65 to 70, 45 to 50, and 36 to 38 kDa; in addition to the latter band, the other two MAbs recognized a band at approximately 30 kDa. All three MAbs were of the immunoglobulin G1 subclass. The two MAbs which are capable of reacting with noncapsular culture supernatant antigen have possible uses in serodiagnosis, particularly in AIDS patients infected with C. neoformans, since in this group the present latex agglutination test has some limitations.
Isolation of Escherichia coli serotype O157:H7 and other Shiga-like-toxin-producing E. coli from dairy cattle
Authors: Wells J G , Shipman L D , Greene K D , Sowers E G , Green J H , Cameron D N , Downes F P , Martin M L , Griffin P M , Ostroff S M
Published: January 2020

We examined 1,266 fecal specimens from healthy cattle during the investigations of two sporadic cases of hemolytic uremic syndrome associated with raw milk consumption and an outbreak of gastroenteritis and hemolytic uremic syndrome caused by Escherichia coli serotype O157:H7. We collected specimens from heifers, calves, and adult cows on 22 farms, in a stockyard, and in a packing house. We also collected 3 raw hamburger specimens from a restaurant and 23 raw milk samples from two farms. All specimens were examined for E. coli O157:H7 by using sorbitol-MacConkey agar, H immobilization, O157 agglutination, and tissue culture cytotoxicity. E. coli O157:H7 was isolated from 16 heifers or calves and 1 adult cow on 22 farms, 1 stockyard calf, 2 beef specimens, and 1 raw milk sample. Selected fecal specimens were also examined for the presence of other Shiga-like-toxin-producing E. coli (SLTEC) by testing polymyxin B extracts of colony sweeps and then testing individual colonies for toxin production. SLTEC other than O157 was isolated from 8 of 10 farms investigated and from the stockyard; 8% of adult cows and 19% of heifers and calves were positive for SLTEC. Several animals were positive for SLTEC by colony sweep only. This investigation demonstrates that dairy cattle are a reservoir of E. coli O157:H7 and other SLTEC.
Isolation and characterization of Sporothrix schenckii from clinical and environmental sources associated with the largest U.S. epidemic of sporotrichosis
Authors: Dixon D M , Salkin I F , Duncan R A , Hurd N J , Haines J H , Kemna M E , Coles F B
Published: January 2020

The largest recorded epidemic of sporotrichosis in the United States occurred in 1988 and involved a total of 84 cases in 15 states. All cases were associated with Wisconsin-grown sphagnum moss. Twenty-one clinical isolates of Sporothrix schenckii and 69 environmental isolates of Sporothrix spp. from the epidemic were characterized and compared. The environmental isolates were recovered from 102 samples of sphagnum moss and other material by using direct plating techniques. Characteristics examined included macroscopic and microscopic morphology, conversion to a yeast phase, exoantigen reactions, and virulence in mice. On the basis of these studies, eight environmental isolates were identified as S. schenckii, five were identified as Ophiostoma stenoceras, and the remainder were identified as Sporothrix species. The environmental isolates of S. schenckii were recovered from moss samples from one Pennsylvania nursery and from three New York State Soil and Water Conservation districts, but none were recovered from moss directly from the bogs in Wisconsin.