bluuebunny/crossref_metadata_2025 huggingface dataset:
Evaluation of a latex agglutination test for the detection of Salmonella and Shigella spp. by using broth enrichment
Authors: Metzler J , Nachamkin I
Published: January 2020

We evaluated the Bactigen Salmonella-Shigella Latex Agglutination Slide Test (Wampole Laboratories, Div. Carter-Wallace, Inc., Cranbury, N.J.) for detection of Salmonella and Shigella spp. in enrichment broth cultures (gram-negative broth incubated for 24 h) as part of the routine testing of stool samples. A total of 1,128 stool samples were screened by using this test. Of 29 samples culture positive for Salmonella spp., 25 were positive with the Salmonella test (sensitivity, 86.2%; specificity, 96.2%). Of four stool samples culture positive for Shigella spp., two were detected with the Shigella latex reagent. Overall, the Shigella test had a specificity of 99.2%. Testing of enrichment broth cultures after 24 h of incubation was more sensitive than was testing after 6 h of incubation. When used for direct culture identification, both reagents had a specificity of greater than 98.0%. We conclude that the Salmonella test may be useful as an enrichment broth screening test to detect Salmonella spp.; however, we cannot make any conclusions about the Shigella test because of the low number of culture-positive specimens in this study.
Staphylococcus aureus capsular types and antibody response to lung infection in patients with cystic fibrosis
Authors: Albus A , Fournier J M , Wolz C , Boutonnier A , Ranke M , Høiby N , Hochkeppel H , Döring G
Published: January 2020

Chronic respiratory tract infections caused by Staphylococcus aureus are common in patients with cystic fibrosis (CF). Recently, it was shown in a few CF patients that S. aureus isolates produce capsular polysaccharides (CPs). However, it is not known whether this is a common feature and whether an immune response to CPs in CF is detectable. Therefore, we examined 170 S. aureus isolates from CF patients and healthy individuals for production of CP types 5 and 8 by using monoclonal antibodies. We found that CP-producing staphylococcal isolates were randomly distributed among CF patients and healthy carriers. Eighty-five percent of all isolates produced CPs, 77% of which were type 8. Examination of one sputum sample by an immunofluorescence technique suggested that production of CPs is not an in vitro phenomenon. S. aureus isolates from various sites of a single person often yielded more than one CP type. A random distribution of S. aureus strains with CP type 5 or 8 from the skin and respiratory tracts of patients and from the skin of healthy individuals was found. Antibody response to CP types 5 and 8, measured by enzyme-linked immunosorbent assay, was not elevated in CF patients with chronic S. aureus lung infection in comparison with healthy carriers. On the contrary, in CF patients the ratios of antibodies to CP 8 were significantly lower (P less than 0.005; alpha = 0.025). The ratios of antibodies to CP types did not change when monitored longitudinally over several months. This study suggests that the production of CPs is a universal property of S. aureus and that infected CF patients do not have elevated ratios of antibodies to these antigens.
Identification of a Yersinia pestis-specific DNA probe with potential for use in plague surveillance
Authors: McDonough K A , Schwan T G , Thomas R E , Falkow S
Published: January 2020

A 900-base-pair DNA fragment derived from a 9.5-kilobase plasmid in Yersinia pestis hybridized specifically with Y. pestis DNA. We demonstrated the feasibility of using this DNA fragment to detect plague bacilli directly in fleas, suggesting that this Y. pestis-specific DNA probe may be used for plague surveillance in the field. Additional applications for this DNA probe may include plague diagnosis and pathogenesis research.
Comparative evaluation of the oxoid signal and Roche Septi-Chek blood culture systems
Authors: Murray P R , Niles A C , Heeren R L , Curren M M , James L E , Hoppe-Bauer J E
Published: January 2020

The Oxoid Signal blood culture system (Oxoid USA, Inc., Columbia, Md.) was compared with the Roche Septi-Chek system (Roche Diagnostics, Div. Hoffmann-La Roche Inc., Nutley, N.J.), with the latter consisting of a tryptic soy broth (R-TSB) bottle with an attached agar slide unit and a Columbia broth bottle. A total of 5,034 cultures with equal volumes of blood in each bottle were processed. Overall, more organisms were recovered in the R-TSB bottle than in the Signal bottle, with significantly more aerobic organisms (Pseudomonas spp., Acinetobacter spp., and yeasts) recovered in the R-TSB bottles and anaerobes and viridans group streptococci recovered in Signal bottles. Approximately equivalent numbers of organisms were recovered in the Signal and Columbia broth bottles. The times of detection were essentially identical with the three blood culture broth systems. During the study, 30.6% of the Signal bottles had a positive indicator of growth, of which 1,103 (71.7%) were false-positive cultures. Additionally, nonviable organisms resembling streptococci were observed in 13.7% of the Signal bottles that were Gram stained and in unioculated blood culture bottles. With appropriate modifications of the preparation of the media, the latter problem can be eliminated.
Potential of in situ hybridization for early diagnosis of productive cytomegalovirus infection
Authors: Stöckl E , Popow-Kraupp T , Heinz F X , Mühlbacher F , Balcke P , Kunz C
Published: January 2020

In situ hybridization with a probe specific for immediate-early genes was used for detection of cytomegalovirus (CMV) transcripts in peripheral blood mononuclear cells, and the potential use of this technique as a diagnostic tool was assessed. The results were compared with those obtained with conventional assay systems. In 8 of 18 continually observed patients who developed a productive CMV infection, a high number of hybridization-positive cells were observed 1 to 2 weeks before the conventional tests yielded positive results. Thus, quantitative evaluation of hybridization results provided an early and specific marker for beginning CMV infection or reactivation. In three cases, quantitative in situ hybridization assays provided the only laboratory marker indicating CMV infection or reactivation. It was also found that a probe specific for immediate-early genes was superior to a probe specific for late genes for diagnosis of productive infections.
Selective medium for Branhamella catarrhalis with acetazolamide as a specific inhibitor of Neisseria spp
Authors: Vaneechoutte M , Verschraegen G , Claeys G , van den Abeele A M
Published: January 2020

Several semiselective media for Branhamella catarrhalis have been proposed. These media allow growth of all members of the family Neisseriaceae, and further differentiation is necessary. By addition of 10 micrograms of acetazolamide, a carbonic anhydrase inhibitor, per ml and incubation in air, a medium was created which reduced growth of Neisseria spp. When saliva samples from 178 healthy schoolchildren were screened for the presence of B. catarrhalis, the carrier rate for this organism was estimated to be 48.9% with the selective medium compared with 12.4% when a semiselective medium, which contains only 10 micrograms of vancomycin, 5 micrograms of trimethoprim, and 2 micrograms of amphotericin B per ml, was used and 6.2% when a nonselective blood agar plate was used. The number of Neisseria spp. isolated dropped from 297 on the semiselective agar to 55 on the selective agar.
Heat-stable-enterotoxin-producing Escherichia coli strains isolated from dogs
Authors: Wasteson Y , Olsvik O , Skancke E , Bopp C A , Fossum K
Published: January 2020

Five strains of hemolytic Escherichia coli isolated from dogs suffering from diarrhea were shown by radioactive and enzyme-labeled oligonucleotide probes to possess genes coding for heat-stable enterotoxin (STIa). Four of the strains were shown by immunoassay (enzyme-linked immunosorbent assay) and bioassay (infant mouse test) to produce STI in vitro. All five strains, however, were able to induce fluid accumulation in ligated dog intestinal loops. The four STI-producing strains all possessed the K99 fimbrial antigen (F5) and belonged to serotype O42:H37. In these strains, genes encoding STI were located on a 98-megadalton plasmid. In the fifth strain, which produced STI in vitro only after several subcultivations, the STI gene was located on an 80-megadalton plasmid. This strain was nontypable.
Rapid isolation of K88+ Escherichia coli by using immunomagnetic particles
Authors: Lund A , Hellemann A L , Vartdal F
Published: January 2020

Superparamagnetic polymer particles precoated with sheep anti-mouse immunoglobulin G were coated with immunoglobulin G2 monoclonal antibodies to the K88 antigen of Escherichia coli (MAb-K88). These immunomagnetic particles (IMP) were used for isolation and identification of K88 antigen-positive (K88+) E. coli. The bacteria presenting the K88 antigen were easily isolated in almost pure culture from a mixed culture of five different O serogroups of E. coli. Nonspecific binding of K88 antigen-negative (K88-) E. coli to the IMP was not observed. The sensitivity of the test to detect K88+ E. coli was found to be 4,000 CFU/ml with fluorescence microscopy. When bacteria attached to the MAb-K88 IMP were grown on blood agar, about 20% of the initial number of CFU was recovered. The test is promising as a rapid method for isolation and identification of K88+ E. coli from a mixed culture.
Time-resolved fluoroimmunoassay compared with virus isolation for rapid detection of respiratory syncytial virus in nasopharyngeal aspirates
Authors: Waris M , Halonen P , Ziegler T , Nikkari S , Obert G
Published: January 2020

Two monoclonal antibodies against two distinct conserved epitopes of the respiratory syncytial virus (RSV) nucleocapsid protein were used in a direct time-resolved fluoroimmunoassay (TR-FIA) for the detection of RSV antigens in nasopharyngeal aspirates. The capture antibody was adsorbed to the solid phase of microdilution strip wells, and the indicator antibody was labeled with a europium chelate. Specimens and label were incubated simultaneously for 1 h at 37 degrees C in the coated wells. After the test samples were washed, fluorescence enhancement solution was added, strips were shaken, and the time-resolved fluorescence was measured. The test procedure took only 75 min, and the total time for 20 specimens, with pretreatment by sonication, was 2 to 3 h. We prospectively evaluated the detection of RSV in nasopharyngeal aspirates of pediatric patients by TR-FIA and by virus isolation in human diploid fibroblasts. TR-FIA detected 40 of 42 isolation-positive specimens. Nine additional isolation-negative specimens were positive by TR-FIA; all proved to be true positives by a blocking-type confirmatory assay. The sensitivity, specificity, positive predictive value, and negative predictive value for TR-FIA were 95, 96, 82, and 99%, respectively, of the values obtained by virus isolation and 96, 100, 100, and 99%, respectively, of the values obtained by virus isolation and the confirmatory assay.
Occurrence of changes in human rotavirus serotypes with concurrent changes in genomic RNA electropherotypes
Authors: Nakagomi T , Akatani K , Ikegami N , Katsushima N , Nakagomi O
Published: January 2020

To investigate the serotypic and genetic diversity of human rotavirus strains, we have tested 513 and 519 fecal rotavirus specimens, respectively, by an enzyme-linked immunosorbent assay with serotype-specific monoclonal antibodies and by polyacrylamide gel electrophoresis of the segmented RNA genome. Of the 513 specimens, 375 were typed as serotype 1 (47.3%), serotype 2 (2.9%), serotype 3 (2.9%), or serotype 4 (17.7%). In addition, a presumptive new human serotype, tentatively referred to as serotype X in this paper, was found in 1.6% of the specimens tested. The remaining 138 specimens (26.9%) were untypeable. Considerable variation in relative frequency of circulating serotypes was observed with respect to geographic locations and observation periods. Rotavirus RNAs were visualized in 481 of 519 specimens tested. Of these, 415 were typed as 33 electropherotypes, many of which were infrequently detected and were restricted to single epidemics. Analysis of the 291 specimens whose electropherotypes and serotypes were available indicated clearly that a given RNA pattern always corresponded to a particular serotype. Heterogeneity of electropherotypes within a serotype was similarly observed in strains belonging to the four previously established serotypes. The results obtained in this study indicated that antigenic changes on the major neutralization antigen occurred always with concurrent changes of genomic RNA electropherotypes. On the other hand, serotypic changes could not be predicted from the changes in RNA electropherotypes.
Application of immunoassay of encephalomyocarditis virus in cell culture with enzyme-labeled virus-specific monoclonal antibodies for rapid detection of virus, neutralizing antibodies, and interferon
Authors: Vlaspolder F , Harmsen T , van Veenendaal D , Kraaijeveld C A , Snippe H
Published: January 2020

Encephalomyocarditis virus (EMCV)-specific monoclonal antibody UM 21.1 labeled with horseradish peroxidase was used to detect EMCV in L-cell monolayers. This direct enzyme immunoassay of EMCV, performed in wells of 96-well plates, could be applied for various purposes, such as early detection of virus multiplication, determination of 50% tissue culture infective doses, and rapid titration of interferon and EMCV-neutralizing antibodies. Multiplication of EMCV is indicated by a rapid increase of the absorbance values measured against EMCV-infected L cells starting as early as 4.5 h after virus inoculation. The early rise of absorbance (i.e., virus multiplication) is inhibited by interferon, allowing its rapid titration. Preincubation of the virus inoculum with neutralizing antibodies also yielded decreased absorbance values. With the latter enzyme immunoassay for neutralizing antibodies, performed after an infection period of 8 h, antibody titers measured were comparable to those obtained with a conventional plaque reduction test. We assume that similar assays could be developed for other picornaviruses (e.g., polioviruses).
Evaluation of the Mini-ID Enterobacteriaceae screen system
Authors: O'Hara C M , Smith P B , Schable B A
Published: January 2020

A total of 932 typical and atypical enteric fermenters were used to evaluate the Mini-ID Enterobacteriaceae Screen/Identification System. At 4 h, final identifications were available for only 13.3%, but an additional 71% were screened into the correct group, according to the product's database. At 24 h, 58.8% were correctly identified to the species level, often with the use of an additional tube. When 118 of the cultures were arranged into a weighted assortment, as might be found in a clinical laboratory, 35 were definitively identified at 4 h, and another 72 were screened into the correct group. Of these 72, 31 were correctly identified to the species level at 24 h, for a total of 56.0%. False-negative ornithine and incorrect L-pyrrolidonyl-beta-naphthylamide and glucuronide xylopyranoside reactions accounted for 54% of the identification errors, while database problems accounted for 10.2% of the errors. Of the eight Salmonella paratyphi A cultures, seven were missed because of a false-positive lysine reaction. At best, the system serves only as a rough screen.
Clinical features of acute infantile gastroenteritis associated with human rotavirus subgroups I and II
Authors: Steele A D , Bos P , Alexander J J
Published: January 2020

During a prospective 3-year study, clinical and epidemiological features of rotavirus gastroenteritis in black infants were investigated. Fever and temperatures exceeding 39 degrees C were more frequent in children shedding subgroup II rotaviruses, whereas vomiting was more pronounced in children with subgroup I infection. Diarrhea, dehydration, duration of illness, and the need for admission to a hospital ward were similar in both groups.
Nucleic acid hybridization for detection of cell culture-amplified adenovirus
Authors: Huang C , Deibel R
Published: January 2020

A number of recombinant plasmids containing genomic segments of adenovirus were constructed. Seven cloned probes, as well as total adenovirus type 2 (Ad2) and Ad16 genomic DNA, were tested by a nucleic acid hybridization technique for sensitivity and specificity in detecting adenoviruses in infected cells. Adenovirus DNA was spotted onto a nitrocellulose filter and hybridized with 32P-labeled DNA probes. The probes, total Ad2 genomic DNA, and plasmid pAd2-H (containing the hexon gene from Ad2 DNA) all detected 10 reference serotypes of five genomic subgroups (A through E) with similar sensitivities. However, plasmid pAd2-H required less preparation time than did total Ad2 DNA. Probes pAd2-F (containing the fiber gene from Ad2) and pAd16-BD (containing the BamHI D fragment from Ad16) hybridized only with reference serotypes from the homologous subgroups (C and B, respectively). Of 101 patient isolates amplified in cells, pAd2-H detected 100% of all isolates from both the homologous and the heterologous subgroups. The detection rates for pAd2-F were 100% (subgroup C) and 3.6% (subgroups A, B, and D), and those for pAd16-BD were 100% (subgroup B) and 9.4% (subgroups A, C, and D). A commercial biotinylated product (Pathogene II) was also included in this study for comparison.
Comparison of modified Bordet-Gengou and modified Regan-Lowe media for the isolation of Bordetella pertussis and Bordetella parapertussis
Authors: Kurzynski T A , Boehm D M , Rott-Petri J A , Schell R F , Allison P E
Published: January 2020

Culture and fluorescent-antibody methods for detection of Bordetella species were evaluated by two state public health laboratories. Field-inoculated plates of Regan-Lowe agar medium were most useful if incubation was initiated on the day of collection. Regan-Lowe and Bordet-Gengou media were comparable for subculturing nasopharyngeal specimens that were transported and enriched in half-strength Regan-Lowe agar. Maximum sensitivity was achieved when the media were used in parallel. Fluorescent-antibody-stained smears of nasopharyngeal specimens were more sensitive for detection of Bordetella pertussis than for detection of Bordetella parapertussis. The fluorescent-antibody method, however, was too insensitive for use without culture.
Early detection of influenza virus by using a fluorometric assay of infected tissue culture
Authors: Pachucki C T , Creticos C
Published: January 2020

A fluorometric substrate, 4-methylumbelliferyl-alpha-ketoside of N-acetylneuramide, was used directly on clinical specimens and infected tissue culture 24 h after inoculation for the detection of influenza viral neuraminidase. Viral neuraminidase was detected in infected tissue culture but not in clinical specimens. The sensitivity of the assay on tissue culture was 92%, and the specificity was 96%.
Isoelectric focusing of ureases from Campylobacter pylori and related organisms
Authors: Tompkins D S , Millar M R , West A P
Published: January 2020

Agarose gel isoelectric focusing was used to determine the isoelectric points of ureases from strains of gastric campylobacterlike organisms isolated from ferrets (pI 5.4), baboons (pI 5.7), and pigs (pI 5.9) and from isolates of Campylobacter pylori in humans (pI 5.9). This technique may help differentiate these closely related bacteria.
Rapid identification of Escherichia coli by Fluorocult media and positive indole reaction
Authors: Heizmann W , Döller P C , Gutbrod B , Werner H
Published: January 2020

To assess the specificity and sensitivity of Fluorocult media for the identification of Escherichia coli, the beta-glucuronidase activities of 1,258 bacterial strains, as well as 20 strains of Candida spp., were investigated. Fluorescence of colonies combined with positive indole reaction resulted in specificities of 99.6 to 99.8%. Sensitivities were 59.1% (MacConkey agar), 69.9% (brolacin agar), 85.5% (Columbia agar), and 85.8% (ECD agar).
Production and characterization of monoclonal antibodies directed against Bordetella pertussis lipopolysaccharide
Authors: Gustafsson B , Lindquist U , Andersson M
Published: January 2020

Hybrid cell lines producing monoclonal antibodies against Bordetella pertussis lipopolysaccharide (LPS) were established. The specificity of the antibodies was ascertained by enzyme-linked immunosorbent assay (ELISA) and ELISA-inhibition experiments with LPS and delipidated polysaccharide fragments (PS-1 and PS-2) prepared from B. pertussis LPS. Monoclonal antibody 9-1-H5 reacted with B. pertussis LPS only, whereas monoclonal antibodies 6-4-H6 and 9-2-A8 reacted with PS-1 and PS-2 as well as B. pertussis LPS. The antibodies did not react with LPS prepared from B. parapertussis and B. bronchiseptica in an LPS-specific ELISA. A monoclonal antibody-based sandwich ELISA was developed for detection of B. pertussis LPS. This assay had a detection limit of B. pertussis LPS in concentrations ranging from 0.16 to 0.32 microgram/ml. The assay was also shown to be specific for the detection of whole B. pertussis bacteria. No cross-reactions were observed with strains of Branhamella catarrhalis, Neisseria meningitidis, Streptococcus miteor, Haemophilus influenzae, or Legionella pneumophila. The monoclonal antibodies might be useful for the detection of soluble antigens and whole bacteria in clinical samples and for studies of the immunochemical structure of B. pertussis LPS.
Comparison of primary rabbit kidney and MRC-5 cells and two stain procedures for herpes simplex virus detection by a shell vial centrifugation method
Authors: Peterson E M , Hughes B L , Aarnaes S L , de la Maza L M
Published: January 2020

By using a conventional tissue culture method as a standard, four shell vial centrifugation culture (SVC) formats were compared for herpes simplex virus (HSV) detection in 300 clinical samples. Both MRC-5 and primary rabbit kidney (PRK) cells were used in the conventional and SVC systems. In addition, both a direct monoclonal fluorescent antibody to HSV (MAb-FA; Syva Corporation, Palo Alto, Calif.) and an indirect HSV polyclonal antibody-horseradish peroxidase stain (poly-HRP; Difco Laboratories, Detroit, Mich.) were used to stain shell vials of both cell types. Conventional tubes were incubated for up to 7 days with daily examination for cytopathic effect, which was confirmed as HSV by staining with an MAb-FA. Shell vials were inoculated, centrifuged, incubated for 16 to 24 h, and stained directly with MAb-FA or indirectly with a poly-HRP stain. Of the 300 specimens examined, 82 (27%) were HSV positive by conventional tissue culture. PRK cells detected 81 (99%) positive specimens, compared with 74 (90%) specimens detected with MRC-5 cells. Of the 82 positive specimens by conventional culture, the SVC formats detected 68 by MRC-5 and MAb-FA, 74 by MRC-5 and poly-HRP, 64 by PRK and MAb-FA, and 77 by PRK and poly-HRP. Therefore, PRK stained by an indirect method with poly-HRP was the most sensitive of the SVC formats tested, detecting 94% of the positive specimens.
Evaluation of methods for detecting human papillomavirus deoxyribonucleotide sequences in clinical specimens
Authors: Caussy D , Orr W , Daya A D , Roth P , Reeves W , Rawls W
Published: January 2020

Specimens from 26 condylomatous lesions, 24 invasive cancer cells, and 33 cervices, without evidence of the diseases, were tested for the presence of human papillomavirus (HPV) types 6, 11, 16, and 18 by Southern blot hybridization, in situ filter hybridization, or in situ tissue hybridization methods. A total of 89% (23 of 26) of the condylomatous lesions contained HPV DNAs, as determined by one or more of the methods. The positive rates for the detection of HPV DNA in condylomas by the different methods were 82% for Southern blot hybridization, 62% for in situ filter hybridization, and 72% for in situ tissue hybridization. Among the specimens from patients with cancer, HPV DNA was found in 83% (19 of 23) by one or more of the methods. Positive rates of 89 and 70%, respectively, were obtained for cancer lesions tested by the filter in situ and Southern blot hybridization methods; however, only 30% of those lesions were positive by the in situ tissue hybridization method. Thirteen percent of the control cervices were positive for HPV DNA by one or more of the assays. With respect to all disease categories, the methods had comparable sensitivities and specificities, except for the in situ tissue hybridization method, which revealed a specificity of 72% for condylomatous lesions and 30% for invasive cancer cells.
Detection of staphylococcal exfoliative toxin by slide latex agglutination
Authors: Murono K , Fujita K , Yoshioka H
Published: January 2020

A simple and rapid method in which slide latex agglutination was used was developed to detect the exfoliative toxin (ET) elaborated by clinical isolates. ET types A and B (ET-A and ET-B) were purified by plate gel isoelectrofocusing, and anti-ET sera were obtained by immunizing rabbits. A specific immunoglobulin G antitoxin was then prepared from the immunized rabbit sera by fast protein liquid chromatography, and latex particles were coated with the antitoxin. Of 74 staphylococcal strains isolated from patients with staphylococcal scalded skin syndrome, 61 strains were found to produce ET by the newborn mouse bioassay. All 61 strains were shown to be positive for ET-A and ET-B production by the slide latex agglutination method. The lowest concentration of ETs detected by the latex agglutination method was 0.5 microgram/ml, which was much lower than that detected by the double immunodiffusion method, with a sensitivity of 50 micrograms/ml. It is crucial to prove ET production by clinical isolates for the diagnosis and surveillance of staphylococcal scalded skin syndrome. The latex agglutination method is a sensitive, simple, and rapid test which can be used as an alternative to the newborn mouse bioassay.
Comparative evaluation of three commercial tests for detection of heterophile antibody in patients with infectious mononucleosis
Authors: Tilton R C , Dias F , Ryan R W
Published: January 2020

Infectious mononucleosis (IM) is caused by the Epstein-Barr virus. Commonly used laboratory tests used for diagnosis of IM include a screening test based on the observation that horse erythrocytes are agglutinated by the Paul-Bunnell antibody found in the serum of patients with IM. This study evaluated two latex agglutination (LA) kits for IM, Monolatex (Wampole Laboratories) and Immunoscan-IM (American MicroScan) (formerly Monogen; Biokit, S.A.), and compared them with Monospot (Ortho Diagnostic Systems) results on 220 patient sera. Discrepancies in the three test results were resolved with complete Epstein-Barr virus antibody profiles. They indicated that any of the three kits tested can be successfully used as a screening test for IM. The advantage of the LA kits is that no differential absorption step is necessary. When discrepancies were resolved, sensitivity and specificity of both LA kits were greater than 93%.
Human coproantibody secretory immunoglobulin A response to Yersinia species
Authors: Fletcher K M , Morris C M , Noble M A
Published: January 2020

A semiquantitative indirect immunofluorescence assay to detect coproantibody secretory IgA (SIgA) was established to investigate the human intestinal immune response to Yersinia species. This assay was based on microagglutination of SIgA in fecal specimens with the patient's homologous organism. Two populations of patients were defined, those who produced an agglutinating (2+) SIgA response and those who did not. A comparison between SIgA production and standard in vitro virulence-related characteristics of infecting organisms, including autoagglutination, calcium dependence, plasmid carriage, and absorption of Congo red, mouse virulence, and clinical presentation, was performed. A positive (2+) SIgA result was associated with acute enteric illness (positive predictive value, 78.6%) and mouse virulence (positive predictive value, 85.7%). When patients with active inflammatory bowel disease were excluded, the positive predictive value of SIgA for mouse virulence and acute enteric disease became 100%. In addition to strains of Yersinia enterocolitica 4,O:3, strains generally characterized as nonpathogenic, including Yersinia frederiksenii, were found to be associated with acute disease, mouse virulence, and stimulation of SIgA. The indirect immunofluorescence assay for detection of SIgA response appears to be a useful indicator of pathogenic strains of yersiniae recovered from enteric specimens.
Fluorescent monoclonal antibody compared with carbohydrate utilization for rapid identification of Neisseria gonorrhoeae
Authors: Welch W D , Cartwright G
Published: January 2020

A commercially available fluorescein-conjugated monoclonal antibody (MAb) (Syva Co., Palo Alto, Calif.; Genetic Systems, Seattle, Wash.) against Neisseria gonorrhoeae was compared with a standard cystine Trypticase agar (CTA) sugar utilization method and with three rapid carbohydrate utilization tests, including the Minitek (BBL Microbiology Systems, Cockeysville, Md.), Neisseria-Stat (Richardson Scientific, Dallas, Tex.), and Neisseria-Kwik (Micro-Biologics, St. Cloud, Minn.) systems for the identification of Neisseria species. The MAb correctly identified all 86 clinical isolates of N. gonorrhoeae. Of these 86 isolates, 28 were found later (48 h after the initial inoculation) to be contaminated with non-Neisseria bacteria. In the other four test systems studied, the identification rates for pure and contaminated N. gonorrhoeae cultures were, respectively, as follows: CTA sugars, 88 and 32%; Minitek, 67 and 50%; Neisseria-Stat, 97 and 96%; and Neisseria-Kwik, 80 and 74%. The MAb did not identify any of the 50 nongonoccocal Neisseria isolates tested. The most expensive test system was the MAb, followed by the Neisseria-Kwik, Minitek, Neisseria-Stat, and CTA sugars systems. The MAb appears to be a rapid and accurate method to identify in vitro isolates of N. gonorrhoeae.
Measurement of antibodies to the Borrelia burgdorferi flagellum improves serodiagnosis in Lyme disease
Authors: Hansen K , Hindersson P , Pedersen N S
Published: January 2020

The isolation of Borrelia burgdorferi flagella and an enzyme-linked immunosorbent assay (ELISA) for detection of immunoglobulin G (IgG) and IgM to the B. burgdorferi flagellum are described. The diagnostic performance of the flagellum ELISA for serodiagnosis of Lyme disease was compared with the performance of a traditional whole cell B. burgdorferi sonic extract ELISA. We examined sera and cerebrospinal fluid (CSF) from 56 patients with lymphocytic meningoradiculitis (Bannwarth's syndrome), the most frequent secondary-stage manifestation of Lyme disease in Europe. Two hundred healthy individuals and patients with aseptic meningitis, encephalitis, Guillain-Barré syndrome, and syphilis served as controls. The flagellum ELISA was significantly more sensitive than the sonic extract ELISA. The diagnostic sensitivities were increased from 41.1 to 76.8% (P less than 0.01) for IgG and from 35.7 to 67.9% (P less than 0.05) for IgM detection in serum. The increase in sensitivity was most pronounced in patients with a short duration of disease (less than 20 days after onset). The diagnostic specificity increased for IgG detection but was almost unaltered for IgM. The flagellum ELISA did not improve the diagnostic sensitivity of measuring antibodies to borreliae in CSF, most likely owing to the low level of unspecific antibodies in CSF compared with serum. The cross-reactivity of sera and CSF from patients with syphilis decreased significantly. The flagellum antigen of B. burgdorferi shows no strain variation, is easy to purify in sufficient quantity, and is therefore a suitable reference antigen for routine serodiagnosis of Lyme disease.
Evaluation of enzyme-linked immunosorbent assay and reversed passive hemagglutination for detection of Crimean-Congo hemorrhagic fever virus antigen
Authors: Shepherd A J , Swanepoel R , Gill D E
Published: January 2020

Enzyme-linked immunosorbent assay (ELISA) and a reversed passive hemagglutination (RPHA) test were evaluated for rapid detection of Crimean-Congo hemorrhagic fever (CCHF) virus antigens. Both RPHA and ELISA detected CCHF antigen in the brains of infant mice 2 to 3 days after infection, several days before the animals sickened and died. Antigen was also detected after 1 to 2 days in infected cell culture extracts and after 2 to 4 days in culture supernatant fluids. Both tests detected CCHF antigen at threshold values of approximately 2.5 log10 tissue culture infective doses per ml and were more sensitive than complement fixation, immunodiffusion, or immunofluorescence. In a comparative study on specimens from CCHF patients, virus was isolated from 38 of 49 sera and 23 of 28 patients. Antigen was detected in 20 of 49 sera (15 of 28 patients) by RPHA and in 29 of 49 sera (18 of 28 patients) by ELISA. Antigenemia was detected more frequently in fatal cases (9 of 11) than in nonfatal cases (9 of 17). Although the antigen detection assays offered a more rapid approach than infectivity assays for diagnosing CCHF, the latter test was more sensitive. The results suggest that RPHA and ELISA may be of use in rapid diagnosis of CCHF infection, particularly in severe cases, in which the danger of nosocomial spread is greatest.
Panel of reference strains for evaluating serologic reagents used to identify gonococci
Authors: Evins G M , Pigott N E , Knapp J S , DeWitt W E
Published: January 2020

A panel of strains for evaluating Neisseria gonorrhoeae serologic reagents was developed. The strains selected for the panel were antigenically diverse and representative of strains isolated worldwide and had been isolated from a variety of anatomic sites. A few strains with characteristics that can cause problems in serologic tests were included. The panel of 52 gonococcal and 20 nongonococcal strains was used to evaluate two commercially produced kits with monoclonal antibody reagents, GonoGen and Phadebact, and one Phadebact kit with absorbed rabbit antiserum. The GonoGen reagent correctly identified all gonococcal strains and did not react with any of the nongonococcal strains. The Phadebact absorbed antiserum reagent correctly identified 47 of 48 gonococcal strains but reacted with 2 of the 20 nongonococcal strains. The Phadebact monoclonal antibody reagent correctly identified all the gonococcal strains; however, it gave positive reactions with 8 and trace reactions with 4 of the 20 nongonococcal strains.
Characterization of Neisseria gonorrhoeae reference strains used in development of serologic classification systems
Authors: Evins G M , Knapp J S
Published: January 2020

Certain strains of Neisseria gonorrhoeae have been used by numerous investigators to develop serologic classification systems. Some of these strains have been used by investigators to study gonococcal virulence. A reference consisting of strain classification by auxotype and serovar, a strain history, and a selected bibliography are provided cohesively.
Nosocomial urinary tract infection with a slowly growing, fastidious Escherichia coli
Authors: Isenberg H D , Vellozzi E M
Published: January 2020

Escherichia coli recovered from a nosocomially acquired urinary tract infection required 48 h of incubation on blood agar and did not grow on other routine clinical laboratory media. This bacterium dissociated readily into three colony types, all of which were confirmed as E. coli by DNA hybridization studies. Preliminary studies indicate a prolonged lag phase that could not be corrected by the addition of a variety of peptones and yeast extracts. Better growth was achieved by the addition of 10% horse serum.
Bacteremia caused by Vibrio hollisae
Authors: Rank E L , Smith I B , Langer M
Published: January 2020

Vibrio hollisae was recovered from the bloodstream of a 36-year-old man with chronic active hepatitis who was admitted to the hospital with signs of gastrointestinal illness. V. hollisae is an infrequent human pathogen associated with seafood ingestion. No etiologic link to seafood was demonstrated in this case report.
First isolation of Legionella gormanii from human disease
Authors: Griffith M E , Lindquist D S , Benson R F , Thacker W L , Brenner D J , Wilkinson H W
Published: January 2020

Legionella gormanii, previously isolated only from the environment, was grown from the bronchial brush specimen of a patient with pneumonia. The organism was characterized by serologic, biochemical, and DNA hybridization studies.
Cross-reactions between Neisseria meningitidis group H and Escherichia coli K2 and K62 polysaccharides
Authors: van Alphen L , Arends A , Hopman C
Published: January 2020

Neisseria meningitidis group H polysaccharide was found to cross-react in immunoprecipitation with cultures, culture supernatants, and purified polysaccharides from Escherichia coli O86:K62 and O6:K2 but not with those fractions from E. coli O6:K-. Antibodies to N. meningitidis group H polysaccharide could be absorbed completely with E. coli K62 and partly with E. coli K2 bacteria but not with E. coli K- bacteria. N. meningitidis H and E. coli K2 and K62 polysaccharides share the structure -phosphate----galactose (Gal)p----glycerol (Gro)-.
Selective medium with DNase test agar and a modified toluidine blue O technique for primary isolation of Branhamella catarrhalis in sputum
Authors: Soto-Hernandez J L , Nunley D , Holtsclaw-Berk S , Berk S L
Published: January 2020

A selective medium with DNase test agar and incorporating vancomycin (10 micrograms/ml), trimethoprim (8 micrograms/ml), and amphotericin B (2 micrograms/ml) supported the growth of 305 Branhamella catarrhalis isolates. A modified toluidine blue O technique was used after 48 h of incubation in CO2 to overlay suspected B. catarrhalis colonies. A metachromatic color change was observed in 15 min, indicating DNase production. In 200 unselected sputum samples of hospitalized patients, this method was compared with routine microbiologic procedures; 31 B. catarrhalis isolates were recovered with the method, compared with 22 isolated from the clinical laboratory. This medium will be particularly useful for culture of sputum, which shows inflammatory cells and gram-negative diplococci on Gram-stained smears.
Comparison of antibody, antigen, and metabolite assays in rat models of systemic and gastrointestinal candidiasis
Authors: Greenfield R A , Troutt D L , Rickard R C , Altmiller D H
Published: January 2020

We compared serial measurements of antibodies to mannan and to a cytoplasmic antigen (enzyme-linked immunosorbent assays), detection of mannan and an unidentified candidal antigen (latex agglutination), and assays of mannose and arabinitol (gas chromatographic assay of per-O-acetylated aldonitrile derivatives). In a high-inoculum intravascular-infection model, antimannan assays were consistently positive beginning on day 2 postinoculation, anti-cytoplasmic antigen assays followed the same time course but were less sensitive, mannan was detected in all samples beginning on day 2 postinoculation, and serum mannose concentrations peaked on day 3 postinoculation and were less sensitive than mannan detection. Other assays were not useful. In a lower-inoculum intravascular-infection model, the antibody assays became positive after a similar interval and remained positive for 28 days, with antimannan again being the more sensitive. Mannan and mannose tests were positive in week 1 postinoculation only, with mannan detection being the more sensitive. In a gastrointestinal-colonization model, antimannan assays become positive after 2 weeks of colonization, whereas anti-cytoplasmic antigen and mannan tests remained negative. In a model of gastrointestinal colonization followed by invasive infection produced by induction of neutropenia, only mannan detection was diagnostically useful. These data, comparing this panel of modern serodiagnostic techniques in controlled models of clinically relevant syndromes of candidiasis, enhance understanding of previous efforts in serodiagnosis of candidiasis and provide a foundation for further prospective studies in patients.
Legionella cincinnatiensis sp. nov. isolated from a patient with pneumonia
Authors: Thacker W L , Benson R F , Staneck J L , Vincent S R , Mayberry W R , Brenner D J , Wilkinson H W
Published: January 2020

A Legionella-like organism (strain 72-OH-H [= ATCC 43753]) was isolated from an open-lung biopsy specimen from a hemodialysis patient with end-stage renal disease and bronchopneumonia. Growth characteristics and gas-liquid chromatographic profiles of the isolate were consistent with those for Legionella spp. The isolate was presumptively identified as a Legionella longbeachae serogroup 1 strain by direct immunofluorescence staining. However, the organism was serologically distinct in the slide agglutination test with absorbed antisera. DNA hybridization studies showed that strain 72-OH-H constitutes a new Legionella species, which is named Legionella cincinnatiensis (ATCC 43753).
Enzyme-linked immunosorbent assay detection of immunoglobulins G and M to Venezuelan equine encephalomyelitis virus in vaccinated and naturally infected humans
Authors: Rosato R R , Macasaet F F , Jahrling P B
Published: January 2020

Enzyme-linked immunosorbent assays (ELISAs) were developed for detection of immunoglobulin G (IgG) and IgM antibodies to Venezuelan equine encephalomyelitis (VEE) virus in vaccinated and naturally infected humans. A total of 441 sera found negative for VEE antibodies by plaque reduction neutralization were examined by IgG ELISA and gave a 1.0% false-positive rate; no false-positives were found in the IgM ELISA. Sera with neutralizing antibody to western or eastern equine encephalomyelitis virus did not react with VEE antigen in the IgG ELISA. Sensitivity of the IgG ELISA was determined by testing 100 coded pre- and postvaccination human sera. Sixty-two were positive by ELISA; 58 of these 62 were also positive by neutralization tests, and 38 were negative by both tests. No neutralization-positive, ELISA-negative sera were found. Comparison of titers obtained by ELISA and neutralization tests indicated that 88% varied randomly by a fourfold dilution factor or less, while 61% were identical or varied only twofold. In sera obtained sequentially from 10 vaccinees and 5 naturally infected patients, both IgG and IgM antibodies appeared between 2 and 3 weeks after vaccination or onset of symptoms. The IgG and IgM antibody ELISAs described are rapid, specific, and sensitive indicators of VEE antibody status in vaccinated and naturally infected individuals.
Quality of commercially produced Shigella serogrouping and serotyping antisera
Authors: Evins G M , Gheesling L L , Tauxe R V
Published: January 2020

Shigella grouping antisera from five manufacturers and typing antisera from two were purchased and evaluated with homologous and heterologous Shigella strains in the slide agglutination test. Only 31 of 73 (42%) antisera were satisfactory. In many instances, the antisera gave negative, as opposed to weak, reactions when they should have given strong positive reactions. Four reagents cross-reacted with Shigella strains. Of the 19 polyvalent grouping antisera to subgroups Shigella dysenteriae serotypes 1 through 7, S. flexneri serotypes 1 through 6, S. boydii serotypes 1 through 7, and S. sonnei forms I, II, only one S. sonnei reagent and five S. flexneri reagents were satisfactory with greater than or equal to 90% of the homologous strains. The reagent of poorest quality was satisfactory with only 18% of the homologous strains. There were three polyvalent antisera to the higher types of S. dysenteriae and S. boydii, which were available from only one company, that adequately identified 80, 63, and 65% of the homologous strains. Typing antisera were available from only two companies, and 30 of 51 (59%) were satisfactory. Commercially available Shigella antisera are inadequate for the laboratory testing required for planning the development of and evaluating Shigella vaccines.
Investigation of human immune response to Micropolyspora faeni antigens by enzyme-linked immunoelectrodiffusion assay and immunoblotting
Authors: Aznar C , Andre P M , Deunff J , Robert R
Published: January 2020

Immune response to Micropolyspora faeni was analyzed in 10 patients suffering from farmer's lung by two techniques: enzyme-linked immunoelectrodiffusion assay (ELIEDA) and immunoblotting. ELIEDA revealed the presence in all patients of various specific immunoglobulin G (IgG), IgM, IgA, and IgE antibodies, with the number of arcs ranging from 4 to 19. M. faeni proteins were isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred to nitrocellulose, and immunoblotted with human sera and specific immunoglobulin-peroxidase conjugates. In immunoblotting, the predominant immunoglobulin class was IgG for all patients. At least 20 bands ranging from 15,000 to 60,000 in molecular weight were observed in a highly positive serum, whereas IgM- and IgA-specific reactivity was directed mainly to the 28,000- and 49,000-molecular-weight bands; M. faeni-specific IgE antibodies appeared less often. The rheumatoid factor (IgM-RF), which also had high titers in these patients (greater than 1/512), interfered with ELIEDA, while only slightly interfering with the immunoblotting detection of specific IgM. This latter technique provided a better characterization of immune response in patients with farmer's lung than ELIEDA did and should also permit discrimination of recently exposed individuals from chronic patients. Moreover, this technique should make it possible to determine whether the response of one particular immunoglobulin class to an antigen fraction can be associated with a specific state of the disease.
Detection of Bacteroides fragilis, Bacteroides thetaiotaomicron, and Bacteroides ovatus in clinical specimens by immunofluorescence with a monoclonal antibody to B. fragilis lipopolysaccharide
Authors: Viljanen M K , Linko L , Lehtonen O P
Published: January 2020

A total of 1,897 clinical specimens (1,019 aspirates and 876 swabs) were studied by indirect immunofluorescence (IF) with a mouse monoclonal antibody (MAb) against a D-galactose oligomer of Bacteroides fragilis lipopolysaccharide. The MAb has been shown to react with 96% of clinical B. fragilis isolates and with about 50% of Bacteroides ovatus and Bacteroides thetaiotaomicron isolates but not with other aerobic or anaerobic organisms tested. The sensitivity of IF in comparison with culturing was 78.9% for all three species. Of the 32 strains originating from culture-positive, IF-negative specimens, 13 lacked the target determinant for the MAb. Sensitivity was highest with specimens taken from the perineal area (87.1%) and lowest with those taken from undefined sites (56.6%). Sensitivity was better with aspirates (86.8%) than with swabs (72.6%). The specificity of IF was 95.6% for all of the material. Positive and negative predictive values were 51.1 and 98.0%, respectively. Neither long transportation times of specimens nor antimicrobial therapy seemed to correlate with the occurrence of IF-positive, culture-negative specimens. This study shows that a single MAb can be used to establish an IF assay that can complement isolation in the detection of these three members of the B. fragilis group.