bluuebunny/crossref_metadata_2025 huggingface dataset:
Direct dilution sampling, quantitation, and microbial assessment of open-system ventilation circuits in intensive care units
Authors: Malecka-Griggs B , Reinhardt D J
Published: January 2020

In a systematic approach, 37 duplicate samples of open system circuits (Bennett MA-1 ventilators) of patients in medical and surgical intensive care units were processed by direct and serial (APHA guidelines) dilutions. The paired difference test on 15 of the in-use circuitry solution samples indicated no difference between the direct and serial dilution methods (P less than 0.001). Seventy-seven additional respiratory therapy circuitry samples from similar intensive care patients were analyzed via a direct dilution method alone and processed microbiologically. The direct dilution procedure was a rapid and accurate means of evaluation of microbial contamination in the range of greater than or equal to 10 to less than or equal to 10(6) CFU/ml. High densities of organisms frequently were found. Sites of contamination included the proximal or patient end of the circuitry (heaviest), the nebulizer trap, and the distal or humidifier portions of the circuitry. The contaminants found were predominantly gram-negative nonfermenters: Acinetobacter calcoaceticus var. antitratus, Pseudomonas aeruginosa, Pseudomonas maltophilia, and Flavobacterium meningosepticum. Fermenters were Klebsiella pneumoniae, Proteus sp., Enterobacter cloacae, Citrobacter diversus, and Enterobacter agglomerans. Infrequently, gram-positive Streptococcus spp. and Staphylococcus spp. were noted.
Detection of antibody to Staphylococcus aureus teichoic acid by enzyme-linked immunosorbent assay
Authors: Yamada J K , Inderlied C B , Porschen R K
Published: January 2020

A sensitive, specific, and rapid enzyme-linked immunosorbent assay has been developed for the detection of immunoglobulin G to Staphylococcus aureus teichoic acid in human sera. Detection of S. aureus teichoic acid antibody is at least 800 times more sensitive than a double diffusion in gel assay, and positive titers of 1:25,600 and greater were observed with this assay. Results with the enzyme-linked immunosorbent assay can be obtained within 3.5 h by using antigen-coated cuvettes. Quantitation of S. aureus teichoic acid antibody by this enzyme-linked immunosorbent assay may be useful in the initial as well as the follow-up diagnosis of serious S. aureus infections.
Sepsis and empyema caused by Yersinia enterocolitica
Authors: Clarridge J , Roberts C , Peters J , Musher D
Published: January 2020

Yersinia enterocolitica is the cause of gastrointestinal infection in the overwhelming majority of recognized cases, although extraintestinal sites are occasionally involved. We report a case of Y. enterocolitica septicemia and empyema complicated by the adult respiratory distress syndrome. The organism was also recovered from the patient's feces by alkaline enrichment and persisted through at least 19 days of antibiotic treatment.
Typing of herpes simplex virus isolates by enzyme-linked immunosorbent assay: comparison between indirect and double-antibody sandwich techniques
Authors: Gerna G , Battaglia M , Revello M G , Gerna M T
Published: January 2020

Indirect and double-antibody sandwich enzyme-linked immunosorbent assay techniques were compared for serotyping 42 herpes simplex virus isolates. Both procedures gave similar results, which were in complete agreement with those obtained by inhibition of the indirect hemagglutination test. The indirect technique proved to be as specific as the double-antibody sandwich enzyme-linked immunosorbent assay, but simpler and cheaper.
Comparison of Phadebact coagglutination, Bactogen latex agglutination, and counterimmunoelectrophoresis for detection of Haemophilus influenzae type b antigens in cerebrospinal fluid
Authors: Collins J K , Kelly M T
Published: January 2020

Cerebrospinal fluid specimens from patients with suspected meningitis were screened with the Phadebact Haemophilus Test (Pharmacia Diagnostics), with Bactogen (Wampole Laboratories), and by counterimmunoelectrophoresis. With culture-positive fluids, Phadebact coagglutination detected 95%, Bactogen latex agglutination detected 91%, and counterimmunoelectrophoresis detected only 79%. Both agglutination techniques were 25-fold more sensitive than counterimmunoelectrophoresis when tested with dilutions of positive fluids. To obtain specific reactions with the Phadebact reagents it was necessary to heat treat (95 degrees C, 5 min) the fluid; with Bactogen and counterimmunoelectrophoresis this was not necessary.
Quality control of moxalactam susceptibility disks
Authors: Barry A L , Preston D A , Jones R N , Gavan T L , Thornsberry C
Published: January 2020

In vitro evaluation of two types of moxalactam disks revealed significant performance differences when Staphylococcus aureus was being tested. The differences were traced to the amount of decarboxylated moxalactam present in the disks. The decarboxylated analog was much more active than the parent compound against S. aureus, not active against Pseudomonas aeruginosa, and approximately as active as the parent compound against Escherichia coli. A nine-laboratory coordinated study was performed to establish quality control parameters for 30-micrograms moxalactam disks. Problems with the establishment of interpretive standards for moxalactam disk tests were evaluated in the light of differences between disks utilized in earlier studies and those that are now commercially available. The type of disk greatly influences standards for tests with S. aureus but has insignificant influence on testing gram-negative bacilli.
Providencia rustigianii: a new species in the family Enterobacteriaceae formerly known as Providencia alcalifaciens biogroup 3
Authors: Hickman-Brenner F W , Farmer J J , Steigerwalt A G , Brenner D J
Published: January 2020

The name Providencia rustigianii sp. nov. is proposed for a group of organisms previously known as Providencia alcalifaciens biogroup 3. By DNA hybridization, strains of P. rustigianii were 81 to 99% related to each other at 60 degrees C, but only 44 to 49% related to P. alcalifaciens biogroups 1 and 2 and 26 to 33% related to Providencia stuartii. P. rustigianii could be differentiated from P. alcalifaciens and P. stuartii by simple biochemical tests. P. rustigianii produced acid from D-galactose but not from trehalose; P. stuartii produced acid from both; and P. alcalifaciens produced acid from neither. P. rustigianii could be distinguished from Providencia rettgeri (formerly Proteus rettgeri) by urea hydrolysis and acid production from D-arabitol; P. rustigianii was negative for these two tests, but P. rettgeri was positive. Strains of P. rustiganii were 32 to 34% related to strains of P. rettgeri. Three of the 11 strains of P. rustigianii were isolated from stools, but the sources of the other isolates are unknown. Three strains (27%) were sensitive to colistin, and 82 to 100% were sensitive to ampicillin, carbenicillin, cephalothin, gentamicin, kanamycin, nalidixic acid, streptomycin, and tetracycline. Strain ATCC 33673 (CDC no. 0132-68) is the type strain for this species.
New York City medium for enhanced recovery of Mycoplasma pneumoniae from clinical specimens
Authors: Granato P A , Poe L , Weiner L B
Published: January 2020

Modified New York City (MNYC) medium and PPLO medium without methylene blue (PPLO agar) were compared for their ability to support the growth of Mycoplasma pneumoniae from clinical specimens. Pharyngeal specimens were collected from 1,070 college students who visited the Syracuse University Student Health Center. Of these patients, 623 were symptomatic for respiratory infection, and the remaining 447 were asymptomatic for respiratory illness. Throat swabs were inoculated into PPLO broths, and these broths were subcultured onto MNYC medium and PPLO agar after 3 and 14 days of incubation. A total of 222 (20.7%) clinical isolates of M. pneumoniae were recovered on these solid media, with the majority of the isolates (196) recovered from symptomatic patients. All isolates grew on MNYC medium, whereas five isolates failed to grow on PPLO agar. All isolates of M. pneumoniae recovered from symptomatic patients were detected on MNYC medium within 1 to 5 days of incubation, whereas 5 to 7 days of incubation were required before mycoplasmal growth was detected on PPLO agar. Over 86% of these mycoplasma isolates were detected on MNYC medium within 3 days of incubation and before the detection of any mycoplasmal growth on PPLO agar. A similar pattern of recovery times was observed for mycoplasmas isolated from asymptomatic patients. The results of this study have shown that MNYC medium is better than PPLO agar in supporting the rapid growth of M. pneumoniae from clinical specimens after 72-h blind subculture in PPLO glucose broth.
Rapid detection of respiratory syncytial virus with a monoclonal antibody
Authors: Bell D M , Walsh E E , Hruska J F , Schnabel K C , Hall C B
Published: January 2020

We developed an indirect fluorescent-antibody test employing a mouse monoclonal antibody directed against the nucleoprotein of RSV for rapid detection of respiratory syncytial virus (RSV). This test demonstrated distinctive fluorescent inclusions in HEp-2 cells infected with 24 RSV isolates collected during 6 previous years but not in cells infected with 13 other respiratory viruses. Examination of nasal cells of 100 infants with acute respiratory illness showed that the indirect fluorescent-antibody test employing the monoclonal antibody was 79% sensitive and 100% specific, as compared with the combination of both culturing and a similar indirect fluorescent-antibody test with commercial anti-RSV serum. This monoclonal antibody is an easily produced, well-characterized, sensitive, and specific reagent for the rapid detection of RSV antigen.
Numerical taxonomy of Vibrio cholerae and related species isolated from areas that are endemic and nonendemic for cholera
Authors: McNicol L A , De S P , Kaper J B , West P A , Colwell R R
Published: January 2020

A total of 165 strains of vibrios isolated from clinical and environmental sources in the United States, India, and Bangladesh, 11 reference cultures, and 4 duplicated cultures were compared in a numerical taxonomic study using 83 unit characters. Similarity between strains was computed by using the simple matching coefficient and the Jaccard coefficient. Strains were clustered by unweighted average linkage and single linkage algorithms. All methods gave similar cluster compositions. The estimated probability of error in the study was obtained from a comparison of the results of duplicated strains and was within acceptable limits. A total of 174 of the 180 organisms studied were divided into eight major clusters. Two clusters were identified as Vibrio cholerae, one as Vibrio mimicus, one as Vibrio parahaemolyticus, three as Vibrio species, and one as Aeromonas hydrophila. The V. mimicus cluster could be further divided into two subclusters, and the major V. cholerae group could be split into seven minor subclusters. Phenotypic traits routinely used to identify clinical isolates of V. cholerae can be used to identify environmental V. cholerae isolates. No distinction was found between strains of V. cholerae isolated from regions endemic for cholera and strains from nonendemic regions.
In vitro evaluation of the BACTEC resin-containing blood culture bottle
Authors: Smith S M , Eng R H
Published: January 2020

The BACTEC resin-containing blood culture bottle (16B; Johnston Laboratories) was designed to bind antibiotics and to support bacterial growth. However, only modest increases in the recovery rates of pathogens have been found in clinical studies. This in vitro study evaluated the ability of the 16B to recover organisms from human serum containing clinically achievable concentrations of antibiotics. Escherichia coli, Enterobacter cloacae, Klebsiella pneumoniae, Pseudomonas aeruginosa, Haemophilus influenzae, Staphylococcus aureus, Streptococcus faecalis, and a viridans streptococcus were added to serum containing antibiotic(s), and at various time intervals of antibiotic exposure, portions were removed and inoculated into both the 16B and the 6B (conventional aerobic) bottles. The studies of the kinetics of killing of the bacterial strains by the various antibiotics showed a good correlation between those combinations of bacteria-antibiotic(s) which produced slow killing and the combinations of bacteria-antibiotic(s) which were recovered preferentially in the 16B bottles. Low recovery rates were noted when the antibiotics killed the organisms rapidly. The indications for use of the resin-containing blood culture bottle should be limited to those situations in which the patient is receiving antibiotics and the bacteremia is suspected to involve a pathogen which is killed slowly by the administered drug(s) or when the bacteremia is continuous. The failure of the BACTEC 16B blood culture bottle to recover organisms may in part reflect the bactericidal activity of the antibiotics administered.
Selective media for recovery of Haemophilus influenzae from specimens contaminated with upper respiratory tract microbial flora
Authors: Chapin K C , Doern G V
Published: January 2020

Isolation of Haemophilus influenzae from specimens contaminated with upper respiratory tract microbial flora was attempted with three different media: enriched chocolate agar, chocolate agar plus vancomycin, and chocolate agar plus vancomycin, bacitracin, and clindamycin. Recovery rates of H. influenzae from 852 pediatric pharyngeal swab specimens were 6.0, 28.5, and 59.9%, respectively.
Ganglioside GM1 enzyme-linked immunospot assay for simple identification of heat-labile enterotoxin-producing Escherichia coli
Authors: Czerkinsky C C , Svennerholm A M
Published: January 2020

A new method has been developed for demonstration of heat-labile (LT) enterotoxin produced by Escherichia coli. This method is based upon the release of LT from bacteria grown directly onto agar plates which have been coated with ganglioside GM1. Toxin bound to the GM1 solid state is subsequently demonstrated by means of a three-step immunoenzymatic procedure in which enzyme-substrate reactions are visualized as dark spots in agarose. When analyzing LT production from 105 E. coli strains, results obtained by this procedure (GM1-ELISPOT) correlated well with those of the GM1 enzyme-linked immunosorbent assay (GM1-ELISA); in no instance were any false-positive reactions observed when highly specific monoclonal antibodies against LT were used. Easy to perform, the GM1-ELISPOT allows demonstration of LT within 24 h after inoculation of the plates, and large numbers of specimens can be screened at the same time without the need of any special equipment. Thus, this new method should meet the requirements of any diagnostic laboratory.
Diagnosis of acute toxoplasmosis by an enzyme immunoassay for specific immunoglobulin m antibodies
Authors: Wielaard F , van Gruijthuijsen H , Duermeyer W , Joss A W , Skinner L , Williams H , van Elven E H
Published: January 2020

A recently developed enzyme-linked immunosorbent assay for detection of immunoglobulin M (IgM) class antibodies to Toxoplasma gondii was evaluated with respect to specificity and sensitivity. By using an antibody capture principle and F(ab')2 conjugates, interference of rheumatoid factors was absent. No cross-reactions with anti-toxoplasma IgG occurred, and no interference with antinuclear antibodies was found. A large-scale study with about 1,500 clinical specimens revealed a 100% specificity. By testing 79 sera from patients with acute-phase acquired toxoplasmosis, sensitivity was found to be 97%. In routine clinical practice, the IgM-enzyme-linked immunosorbent assay proved to be a more sensitive tool for diagnosis than the immunofluorescent-antibody test. The course of IgM-enzyme-linked immunosorbent assay antibodies in acute patients was studied; IgM reached peak levels within 1 month after onset of illness, and could be demonstrated up to an average of 8 months after onset.
Serology of capsulated streptococci pathogenic for pigs: six new serotypes of Streptococcus suis
Authors: Perch B , Pedersen K B , Henrichsen J
Published: January 2020

A total of 138 capsulated strains of streptococci were isolated from pigs with various infections, notably septicemia and bronchopneumonia. Biochemically all strains were similar, and 31 strains were serologically identical to the previously described groups R, S, and RS streptococci, provisionally designated Streptococcus suis types 1, 2, and 1/2 by Elliot. The remaining 107 strains could be divided into six new capsular serotypes of S. suis.
Enzyme-linked immunosorbent assay that uses labeled antigen for detection of immunoglobulin M and A antibodies in toxoplasmosis: comparison with indirect immunofluorescence and double-sandwich enzyme-linked immunosorbent assay
Authors: van Loon A M , van der Logt J T , Heessen F W , van der Veen J
Published: January 2020

A direct enzyme-linked immunosorbent assay (ELISA) is described that uses horseradish peroxidase-labeled antigen for detection of immunoglobulin M (IgM) and IgA antibodies to toxoplasma. In this assay, polystyrene microtiter plates were sensitized with anti-human IgM or IgA antibody to separate IgM or IgA from other classes of antibody. The presence of IgM or IgA antibodies to toxoplasma (Tox-IgM, Tox-IgA) was then detected by sequential addition of soluble horseradish peroxidase-labeled toxoplasma antigen and substrate. As judged by examining sucrose gradient-fractionated sera, the assay was specific for IgM or IgA classes of antibody. In contrast to the indirect immunofluorescence for IgM antibodies to toxoplasma, no inhibition of IgM reactivity by specific IgG antibodies could be detected. Furthermore, rheumatoid factor did not cause false-positive results. Of 80 single sera with high antibody titer to toxoplasma in indirect immunofluorescence and complement fixation, 40 were positive in the direct ELISA for Tox-IgM, 36 were positive in the double-sandwich ELISA, and only 21 were positive in the indirect immunofluorescence for Tox-IgM when whole serum was used. In the indirect immunofluorescence, another 13 sera became positive after sucrose gradient fractionation. The direct ELISA for IgA antibodies to toxoplasma was positive in 43 sera, of which 39 were positive in the direct ELISA for Tox-IgM. High levels of IgM antibodies were found within 3 months after the onset of symptoms, slowly decreasing thereafter. Tox-IgM may persist for more than 1 year after infection.
Serum antibodies in Campylobacter enteritis
Authors: Kaldor J , Pritchard H , Serpell A , Metcalf W
Published: January 2020

A solid-phase enzyme immunoassay method for the measurement of total and class-specific serum antibodies to Campylobacter jejuni was developed. The test was found to be both sensitive and specific. Immunoglobulin M, G, and A antibodies were detected in about 90% of sera collected from patients recovering from Campylobacter enteritis, with the pattern of appearance and decline of these antibodies conforming to that usually seen in an acute infection. A poor antibody response was noted in patients with intermittent or prolonged enteritis. The clinical implications and diagnostic value of this test are discussed.
Detection of two Blastomyces dermatitidis serotypes by exoantigen analysis
Authors: Kaufman L , Standard P G , Weeks R J , Padhye A A
Published: January 2020

We evaluated the diagnostic value of antibody to Blastomyces dermatitidis A precipitin arcs for identifying B. dermatitidis cultures by the exoantigen technique with 88 isolates from the United States and Canada, 12 from Africa, and 1 each from India and Israel. In addition, we studied 190 mycelial-form isolates of other fungi which could be confused antigenically or morphologically with B. dermatitidis. Antigen extracts from all of the North American isolates, the Indian and Israeli isolates, and one of the African isolates reacted with the B. dermatitidis A antibody. The anti-A serum did not react with antigens from the other 11 African isolates or with antigens from any of the 190 heterologous fungi. Studies with unadsorbed and adsorbed antisera to selected North American and African isolates of B. dermatitidis showed that the North American isolates not only produced the distinct A antigen, but also formed another antigen that was common to all 11 of the antigenically less complex African isolates. The results of this study indicate that at least two serotypes of B. dermatitidis exist. All of the North American, Indian, and Israeli isolates and one of the African isolates contained the A antigen and were designated serotype 1. Most of the African isolates (11 of 12) constituted a second serotype that was deficient in A but showed an antigen designated K.
Analysis of eight cases of neonatal meningitis and sepsis due to Enterobacter sakazakii
Authors: Muytjens H L , Zanen H C , Sonderkamp H J , Kollée L A , Wachsmuth I K , Farmer J J
Published: January 2020

Eight cases of neonatal meningitis due to Enterobacter sakazakii (formerly known as yellow-pigmented Enterobacter cloacae) which occurred in The Netherlands during the last 6 years were investigated retrospectively. Two patients had necrotizing enterocolitis and meningitis simultaneously. Despite treatment (in most cases with ampicillin and gentamicin), the fatality rate was 75%. Strains were much more susceptible to some of the new beta-lactam antibiotics than to ampicillin. A mode of transmission other than passage through the birth canal was likely, at least for some patients. A cluster of four patient strains in one hospital had almost identical plasmid DNA profiles. However, two strains isolated from formula at the same hospital 2 days after the onset of one case had different profiles, as did the strains from patients in other hospitals.
Comparison of different antigen preparations as substrates for use in passive hemagglutination and enzyme-linked immunosorbent assays for detection of antibody against bovine enteric coronavirus
Authors: Crouch C F , Raybould T J
Published: January 2020

Purified coronavirus, detergent extracts of purified coronavirus, and virus-infected Madin-Darby bovine kidney cells were evaluated as antigen substrates in enzyme-linked immunosorbent assay (ELISA) and passive hemagglutination systems. Only detergent-extracted and -unextracted, purified viruses were reactive as antigen substrates in ELISA, whereas all three antigen preparations could be used for sensitization of erythrocytes in the passive hemagglutination assay. The passive hemagglutination system with infected cell extracts exhibited a similar level of sensitivity and specificity to the ELISA system employing purified coronavirus but enabled 300 times more tests to be performed per volume of virus-infected cell culture.
Separation of bacterial ubiquinones by reverse-phase high-pressure liquid chromatography
Authors: Moss C W , Guerrant G O
Published: January 2020

A procedure was developed for the separation of ubiquinones by high-pressure liquid chromatography on a reverse-phase C18 column. Ubiquinones Q6 through Q14 were resolved in 20 min and were distinguished from menaquinones by comparing UV spectra at 248 and 275 nm.
Diagnosis of bacterial vaginosis by direct gram stain of vaginal fluid
Authors: Spiegel C A , Amsel R , Holmes K K
Published: January 2020

To determine whether bacterial vaginosis (BV), also known as nonspecific vaginitis, could be diagnosed by evaluating a Gram stain of vaginal fluid, we examined samples from 60 women of whom 25 had clinical evidence of BV and 35 had candidal vaginitis or normal examinations. An inverse relationship between the quantity of the Lactobacillus morphotype (large gram-positive rods) and of the Gardnerella morphotype (small gram-variable rods) was noted on Gram stain (P less than 0.001). When Gram stain showed a predominance (3 to 4+) of the Lactobacillus morphotype with or without the Gardnerella morphotype, it was interpreted as normal. When Gram stain showed mixed flora consisting of gram-positive, gram-negative, or gram-variable bacteria and the Lactobacillus morphotype was decreased or absent (0 to 2+), the Gram stain was interpreted as consistent with BV. Gram stain was consistent with BV in 25 of 25 women given a clinical diagnosis of BV and in none of 35 women with candidal vaginitis or normal examinations. Duplicate slides prepared from 20 additional specimens of vaginal fluid were stained by two methods and examined by three evaluators. Interevaluator interpretations and intraevaluator interpretations of duplicate slides were in agreement with one another and with the clinical diagnosis greater than or equal to 90% of the time. We concluded that a microscopically detectable change in vaginal microflora from the Lactobacillus morphotype, with or without the Gardnerella morphotype (normal), to a mixed flora with few or no Lactobacillus morphotypes (BV) can be used in the diagnosis of BV.
Monoclonal antibodies for serotyping Leishmania strains
Authors: Greenblatt C L , Slutzky G M , de Ibarra A A , Snary D
Published: January 2020

Mouse monoclonal antibodies raised against Leishmania tropica major were found to precipitate with the excreted factor produced by this and other leishmanial species. We suggest that a classification system for Leishmania, based on selective precipitation reactions between monoclonal antibodies and the excreted factor, would remove many ambiguities that currently exist.
Evaluation of a latex agglutination test for detection of antibodies to rubella virus in selected sera
Authors: Freeman S , Clark L , Dumas N
Published: January 2020

A new latex agglutination test for rubella virus was used to test sera with a hemagglutination inhibition titer of less than or equal to 8 (97 specimens) or greater than or equal to 256 (158 specimens). Maximum latex agglutination test sensitivity was achieved when low-titer sera were tested undiluted and high-titer sera were diluted (1:10). A modification of the protocol of the manufacturer resulted in a latex agglutination-hemagglutination inhibition agreement of 96.7%.
Rapid biochemical tests for the identification of groups A, B, C, F, and G streptococci from throat cultures
Authors: Slifkin M , Gil G M
Published: January 2020

A test employing three fluorogenic 4-methylumbelliferyl substrates and the lectin of Dolichos biflorus was developed for the identification of beta-hemolytic streptococcal colonies associated with throat cultures. This non-serological method is unique in that it permits the accurate identification of groups C, F, and G streptococci, as well as groups A and B streptococci. The method is rapid, simple, and specific and appears to be a useful means to identify groups A, B, C, F, and G streptococci.
Accuracy of calibrated-loop transfer
Authors: Albers A C , Fletcher R D
Published: January 2020

The accuracy of the 0.001-ml calibrated platinum loop was tested by two methods: visible absorption spectroscopy and weight determinations. By both methods it was demonstrated that the volume of the loop delivery is related to the angle at which the loop is withdrawn from the solution being sampled and the diameter of the container, provided that the volume in the container is adequate to cover the loop. Vertical sampling from small-diameter containers (less than or equal to 7-mm inside diameter) delivered approximately 50% of 0.001 ml, and sampling at a 45 degree angle from larger containers (greater than or equal to 22-mm inside diameter) gave a delivery volume of greater than 150% of 0.001 ml, resulting in a threefold difference in the amount delivered and an error rate of +/- 50%.
Relationships between virulence and morphological or serological properties of variants dissociated from serotype 1 Haemophilus paragallinarum strains
Authors: Sawata A , Kume K
Published: January 2020

Relationships between morphological or serological properties and virulence were investigated in seven Haemophilus paragallinarum variants. The variants, originated from five serotype 1 strains, were classified into seven types on the basis of colony morphology and iridescence and the presence of variant-specific antigens. Smooth (S) and encapsulated organisms having variant-specific antigens and forming highly iridescent (ir+) colonies were highly virulent in vivo; slightly encapsulated organisms having variant-specific antigens and forming slightly iridescent (ir +/-) colonies were moderately virulent; and nonencapsulated or slightly encapsulated organisms with or without variant-specific antigens and forming noniridescent (ir-) or ir +/- colonies were avirulent. Virulence was well correlated with the amount of capsule substance containing hyaluronic acid. The evidence suggests that the presence of variant-specific agglutinogen L and hemagglutinin HA-L seem to be responsible for adherence or colonization, but not for virulence, of the organisms in chickens.
Comparison of methods for quantitating antigen-specific immunoglobulin M antibody with a reverse enzyme-linked immunosorbent assay
Authors: Siegel J P , Remington J S
Published: January 2020

We compared two methods for quantitating antigen-specific antibody by a reverse enzyme-linked immunosorbent assay. In the serial-dilution method, the result was determined to be the highest dilution of a serum yielding an absorbance above an established threshold. In the single-dilution method, the result was determined by comparing the absorbance yielded by the test serum at a standard dilution to that yielded by positive and negative reference sera at the same dilution. The results in the single-dilution method reflected the antigen-specific immunoglobulin M antibody activity as a proportion of total immunoglobulin M antibody in a serum, i.e., the immune load, whereas the results in the serial-dilution method reflected the absolute concentration of antigen-specific immunoglobulin M antibody activity. Compared with results in the serial-dilution method, results in the single-dilution method had considerably greater reproducibility on a day-to-day basis and under various test conditions. The single-dilution method was more useful in discriminating between sera from patients in an early stage of clinical infection due to Toxoplasma gondii and sera from patients in a later stage of infection.
ompA gene in the detection of Escherichia coli and other Enterobacteriaceae by nucleic acid sandwich hybridization
Authors: Palva A M
Published: January 2020

The applicability of the DNA sandwich hybridization method to detection of bacterial DNA from crude samples is demonstrated using Escherichia coli as a model. In sandwich hybridization the sample DNA mediates the binding of a labeled probe DNA fragment to a second DNA bound on filter. For this study the DNA reagents were prepared by subclonings from a recombinant plasmid containing the E. coli K-12 ompA gene and an adjacent fragment of E. coli DNA. The 5' half of the ompA gene (738 base pairs) in pBR322 served as the filter reagent. The 3' half of the ompA gene (300 base pairs) and the adjacent 1,500 base pairs of E. coli DNA were cloned into the single-stranded phage vector M13mp7, and the resulting recombinant phage DNA was labeled with 125I and used as probe in the hybridizations. For maximal hybridization the DNA reagents had to be present in excess of the sample nucleic acid, which was preferably fragmented before testing. In the optimized test, 3 X 10(6) molecules of E. coli DNA from lysed cells were detected by an overnight reaction; the sensitivity of the test was not affected by the presence of 10(9) unrelated bacteria. With the ompA reagents, all members of the family Enterobacteriaceae tested were detected even if the sensitivity was decreased as compared with that for the homologous bacteria. With all other bacteria, including aerobic and anaerobic species of clinical importance, the test was negative. These findings suggest that it may be possible to find group-specific reagents to be used in diagnostic bacteriology.
Enzyme immunoassay for detection of antibody to toxins A and B of Clostridium difficile
Authors: Viscidi R P , Yolken R H , Laughon B E , Bartlett J G
Published: January 2020

An enzyme immunoassay (enzyme-linked immunosorbent assay [ELISA]) to detect hamster antibody to toxins A and B of Clostridium difficile was developed in which toxin preparations are used to coat the solid phase. The specificity of the assay was supported by blocking tests with the toxin preparations and proteins from a nontoxigenic strain. Sera from immunized and control hamsters were tested by this technique, and results were compared with those from a cytotoxicity neutralization assay. Antibody to toxins A and B assayed by ELISA showed a close quantitative correlation with antibody titers obtained by cytotoxicity neutralization. The ELISA assays described appear to provide a sensitive, specific, and practical method to define the prevalence of antibody to C. difficile toxins. These assays could be readily applied to human sera to examine and study the immune response of patients with C. difficile-induced disease.
Determination of serum bactericidal activity with the aid of luminous bacteria
Authors: Barak M , Ulitzur S , Merzbach D
Published: January 2020

Nonmarine luminous bacteria belonging to the genus Vibrio cholerae were extremely sensitive to the bactericidal activity of human serum. Luminous bacteria incubated in a medium containing serum showed a decrease in their in vivo luminescence that was directly proportional to the decrease in the viable count and was a function of the serum concentration. Both immunoglobulins and the complement system were required to exert the serum bactericidal activity. Serum lacking immunoglobulins or certain complement components, especially C3, did not affect the luminescence. The bactericidal effect of the serum on luminous bacteria was diminished by the presence of lipopolysaccharide or by pretreatment of the serum with different species of killed bacteria. As found in other systems, the bacteriolytic activity of serum was only augmented by lysozyme, but was not lysozyme dependent; although the luminous bacteria were converted into spheroplasts in serum containing 0.5 M sucrose, their in vivo luminescence was almost not affected. This system could easily distinguish between the C classical pathway and the properdin pathway. Ethylene glycol-bis (beta-aminoethyl ether)-N,N'-tetraacetic acid, which inhibits only the classical complement pathway, did not inhibit the decrease in luminescence as did EDTA. Thus, it was possible to distinguish between deficiencies in complement components participating in both pathways and complement components that were involved only in the classical pathway. This system could also be used as a substitute to the hemolytic system in complement fixation tests.
Detection of Antichlamydial Immunoglobulin G and M Antibodies by Enzyme-Linked Immunosorbent Assay
Authors: Mahony J. B. , Schachter J. , Chernesky M. A.
Published: January 2020

Chlamydia trachomatis
causes a wide range of infections in adults and conjunctivitis and pneumonia in neonates. The complement fixation test for chlamydial antibody is broadly reactive, but possesses low sensitivity, whereas the microimmunofluorescence test is highly sensitive, but technically difficult to perform. A simple, rapid enzyme-linked immunosorbent assay (ELISA) has been developed for the measurement of immunoglobulin G (IgG) and IgM antibodies to
C. trachomatis.
Wells of microtiter plates were coated with Renografin-purified elementary bodies (serotype L2) grown in cycloheximide-treated McCoy cells, and serum antibody was detected with peroxidase-labeled goat antihuman IgG and IgM antibody. Of 41 sera tested from patients with lymphogranuloma venereum, pelvic inflammatory disease, cervicitis, or urethritis there was a 90 and 63% correlation of positive results for IgG and IgM, respectively, by microimmunofluorescence and ELISA. Of the positive correlates, ELISA titers were up to 128 times higher than microimmunofluorescence titers for IgG and IgM. The ELISA detected no false-positive results, but missed two positive results for IgG. Both of these sera were reactive against serotypes C and J, suggesting that the ELISA with LGV L2 antigen may not measure antibodies to serotypes within the C serogroup. The IgM ELISA detected 7 negative and 4 positive results not detected by the microimmunofluorescence test. Of four paired sera examined by ELISA, three showed a fourfold rise in IgG antibody titer, and one showed a twofold rise. Further evaluation of this ELISA will be required to determine how useful it will be in seroepidemiological studies and as a diagnostic tool.
Extended scheme for serotyping Campylobacter jejuni: results obtained in Israel from 1980 to 1981
Authors: Rogol M , Sechter I , Braunstein I , Gerichter C B
Published: January 2020

The serotyping scheme for Campylobacter jejuni previously developed in the National Center for Campylobacter, Jerusalem, was extended by the use of 20 new sera and modified by the absorption of the sera, when necessary, with homologous boiled cultures or heterologous live cultures. The extended scheme is based on slide agglutination of live suspensions and is performed in two stages: pretesting with four pooled sera and final testing with monovalent sera. So far, 34 serotypes have been recognized. Among 442 isolates of C. jejuni, 86.4% could be typed with this scheme. Of the 382 cultures typed, 90% reacted with one single serum, and the remainder showed a complex antigenic structure. The frequent serotypes identified were: 11 (12.9%), 12 (8.2%), 18 (6.3%), and 3 (6.1%). When epidemiological data were available, the results of the serotyping corresponded with the epidemiological evidence.
Production and characterization of monoclonal antibodies specific for a glycosylated polypeptide of human cytomegalovirus
Authors: Kim K S , Sapienza V J , Chen C M , Wisniewski K
Published: January 2020

Nine hybrid cell lines producing antibodies specific for cytomegalovirus (CMV) antigen were obtained after fusion of P3/X63-Ag8 myeloma cells with spleen cells from BALB/c mice immunized with CMV complement-fixing antigen. By the immunoblot technique, five of nine antibodies (4D11, 7B4, 7D2, 8E3, and 8E10) were identified as being reactive to a CMV glycosylated polypeptide with molecular weight of 66,000 (GP66). Four other antibodies (1B8, 8E9, 4D2, and 7E2) appeared to be reactive with CMV antigen(s) only if the antigen was not denatured by sodium dodecyl sulfate. These remain unassigned until further studies are done. With the enzyme-linked immunosorbent assay (ELISA), competitive bindings were performed with a constant amount of horseradish peroxidase-conjugated antibody and various concentrations of unconjugated homologous and heterologous antibodies on CMV antigen-coated ELISA wells, and the antigenic determinant specific for each antibody was determined. The nine antibodies could be classified into six different groups, each group reacting with a different epitope or a different region with two or more antigenic determinants which are so close to each other that they cause binding inhibition. They are groups A (4D11), B (7B4, 8E10), C (7D2), D (4D2, 7E2, 8E9), E (8E3), and F (1B8). The extent of competition among antibodies within each group was the same. By using the two antibodies that reacted with different epitopes on GP66, a double-antibody sandwich ELISA method was developed. The method was sensitive enough to detect as little as 50% of the antigen present in one infected cell or 0.000245 U of CMV complement-fixing antigen per test well. Other strains of CMV (David, Kerr, Espilat, C-87, and five clinical isolates) gave positive results, whereas herpes simplex virus types 1 and 2, varicella-zoster virus and Epstein-Barr virus nuclear antigen preparations did not. By the indirect immunofluorescence assay, antibodies 4D11 and 8E3 were able to detect GP66 in the nucleus of CMV-infected F-5000 human embryonic fibroblasts as early as 2 h postinfection and were superior in this respect to the remaining seven antibodies tested. By the double-antibody sandwich ELISA, the presence of GP66 in CMV-infected cells was detected as early as 2 h postinfection.
Growth of Haemophilus influenzae in simulated blood cultures supplemented with hemin and NAD
Authors: Artman M , Domenech E , Weiner M
Published: January 2020

The mean generation time of Haemophilus influenzae in simulated blood cultures is 103 to 107 min. With 0.56 to 0.58 doublings per h, even large inocula of 256 cells per ml reach only 2 X 10(6) cells per ml and produce no visible evidence of growth after 24 h of incubation. Hemin and NAD added to simulated blood cultures triple the rate of growth of H. influenzae, so that even small inocula produce visible turbidity after overnight incubation. With a mean generation time of 36 min, a single cell of H. influenzae in simulated blood culture supplemented with hemin and NAD undergoes 30 doublings in 18 h, producing 2(30) (1.07 X 10(9] cells and visible turbidity.
Preliminary investigation of Candida albicans biovars
Authors: Román M C , Linares Sicilia M J
Published: January 2020

A total of 126 Candida albicans strains were enzymatically evaluated by the API ZYM system. Four enzymatically based groups of C. albicans are recognized.
Proposed zone standards for interpretation of fortimicin A disk diffusion tests
Authors: Barry A L , Thornsberry C , Jones R N
Published: January 2020

Fortimicin A microdilution tests and disk diffusion tests (10-, 30-, and 50-micrograms disks) were performed with 454 selected bacterial strains. Tests with 30-micrograms disks were preferred, with zone standards of less than or equal to 14 mm of resistant strains (minimal inhibitory concentration, greater than 16 micrograms/ml) and greater than or equal to 17 mm for susceptible strains (minimal inhibitory concentration, less than 16 micrograms/ml).
Comparison of direct immunofluorescence and direct immunoperoxidase procedures for detection of herpes simplex virus antigen in lesion specimens
Authors: Schmidt N J , Dennis J , Devlin V , Gallo D , Mills J
Published: January 2020

Direct immunofluorescence and direct immunoperoxidase staining were equally sensitive and specific for detection of herpes simplex virus antigen in lesion specimens, and each method showed 82% agreement with virus isolation results.
Comparison of two commercially available isolation systems for Chlamydia trachomatis
Authors: Baselski V S , Robison M K , Jennings B R
Published: January 2020

In a comparison of two commercially available chlamydial isolation systems in which cycloheximide-treated McCoy cell monolayers are used, the system from Bartels Immunodiagnostic Supplies, Inc., Bellevue, Wash., was found to be superior to that from M. A. Bioproducts, Walkersville, Md. for the detection of Chlamydia trachomatis by iodine staining. Of 288 clinical specimens run in parallel, 47 (16.3%) were positive, with 16 of 47 positive results detected in the Bartels system only and 1 of 47 positive results detected in the M. A. Bioproducts system only (P less than 0.001). A comparison of the number of inclusion-forming units per cover slip from clinical specimens and passaged isolates also showed that the Bartels cell system demonstrated higher inclusion counts than the M. A. Bioproducts system. In routine clinical use, overall isolation rates were higher (P less than 0.001) and contamination rates were lower (P less than 0.001) with the Bartels system as compared with results obtained in a previous time period in which the M. A. Bioproducts system was used.
Isolation of Clostridium pseudotetanicum from a patient with gas gangrene
Authors: Katoh N , Watanabe K , Miki Y , Muro T , Ueno K
Published: January 2020

Clostridium pseudotetanicum was isolated along with Serratia marcescens, Proteus mirabilis, and Staphylococcus epidermidis from a patient suffering from gas gangrene who had been injured in the right leg by a power cultivator. Experimental infection of the hind leg of mice with C. pseudotetanicum and the three kinds of aerobic bacteria did not produce any different macroscopic finding in the infection site, compared with aerobic bacterial injection, except for some enlargement of the involved tissue and some slightly altered histolytic findings. Ampicillin, rifampin, and tinidazole were the most active antimicrobial agents against C. pseudotetanicum.
An improved reagent for mycobacterial nitrate reductase tests
Authors: Warren N G , Body B A , Dalton H P
Published: January 2020

A new crystalline reagent for nitrate reductase tests was compared with standard liquid reagents on 437 strains of mycobacteria. The results for isolates of Mycobacterium avium complex, Mycobacterium kansasii, Mycobacterium gordonae, Mycobacterium scrofulaceum, Mycobacterium fortuitum, and Mycobacterium chelonei agreed 100% with the expected results. Of the 177 Mycobacterium tuberculosis isolates, 4 were negative by the conventional method. Two of these four isolates were positive with the new reagent. Of the positive nitrate tests carried out with liquid reagents, 42% flashed instantly or faded in color; none of the tests carried out with the new crystalline reagent flashed or faded. A stronger color reaction was seen for 28% of the positive tests with the new reagent.
Detection and characterization of mouse monoclonal antibodies to epidemic typhus rickettsiae
Authors: Black C M , Tzianabos T , Roumillat L F , Redus M A , McDade J E , Reimer C B
Published: January 2020

A solid-phase immunofluorometric assay was used to detect mouse monoclonal antibodies to epidemic typhus rickettsiae, Rickettsia prowazekii (the immunizing antigen), and to murine typhus rickettsiae, Rickettsia typhi, a related antigen. Of the 649 hybridoma cultures obtained, 628 contained antibodies either to R. prowazekii or to both R. prowazekii and R. typhi. A total of 72 cultures were cloned by limiting dilution and yielded 137 antibody-producing clones. Of these, 104 produced antibodies specific for R. prowazekii, 22 produced antibodies that reacted with R. prowazekii and R. typhi, and 11 produced antibodies that reacted with R. prowazekii, R. typhi, and R. canada. The immunoglobulin isotypes of the mouse monoclonal antibodies produced were identified by a related indirect immunofluorometric assay technique with fluorescein isothiocyanate-conjugated antisera specific for each isotype. Antibodies were also evaluated by indirect fluorescent antibody tests, and antibodies from selected clones were found to neutralize rickettsial toxic activity in mice.
Isolation and characterization of an equine rotavirus
Authors: Hoshino Y , Wyatt R G , Greenberg H B , Kalica A R , Flores J , Kapikian A Z
Published: January 2020

A rotavirus, designated as the H-1 strain, was isolated from a diarrheic foal in primary African green monkey kidney cells and MA104 cells. This cell culture-adapted strain hemagglutinated erythrocytes of human group O, rhesus monkeys, guinea pigs, and sheep. It was found to be similar, if not identical, to porcine rotaviruses (strains OSU, EE, and A-580) by plaque reduction neutralization and hemagglutination inhibition tests, and, in addition, it was found to belong to subgroup 1. This equine rotavirus has an RNA electrophoretic migration pattern which was distinct from those of the three strains of porcine rotavirus. The serological relationship established by plaque reduction neutralization and hemagglutination inhibition tests between the equine (H-1) and porcine (OSU, EE, and A-580) rotaviruses is an example of a rotavirus of the same serotype being isolated from different species. The H-1 strain was distinct from four human rotavirus serotypes (Wa, DS-1, P, and St. Thomas 4) as well as from bovine rotavirus NCDV, simian rotavirus MMU18006, and canine rotavirus CU-1 by plaque reduction neutralization tests. This equine isolate (H-1) was found to be related antigenically to canine CU-1 and bovine NCDV rotaviruses in a one-way fashion by hemagglutination inhibition tests.