bluuebunny/crossref_metadata_2025 huggingface dataset:
Impact on routine diagnosis of echovirus infections of intratypic differentiation and antigenic variation in echovirus type 25 studied by using monoclonal antibodies
Authors: Peigue-Lafeuille H , Fuchs F , Gharabaghi F , Chambon M , Aymard M
Published: January 2020

We studied the biological and antigenic properties of wild strains of echovirus type 25 isolated in France between 1982 and 1987 and compared them with the JV-4 prototype strains isolated in 1957. The wild strains differed from the prototype strain in their cellular tropism. The prototype strain grew readily in five cell lines (MRC5, MA 104, Vero, BGM, and HT 29-18), while for wild strains MRC5 and HT 29-18 cells were the most sensitive and supported growth to high titres (between 4.5 and 7.4 50% tissue culture infective doses per 0.05 ml). Plaques produced by wild strains were larger (6.05 +/- 0.94 mm in diameter [mean +/- standard deviation]) than those of the prototype strain (2.3 +/- 0.97 mm in diameter) and heterogeneous, even after cloning by three terminal dilution passages, which suggested heterogeneous virus populations. Virus neutralization with polyclonal monovalent sera showed that wild strains were significantly less neutralized by two reference immune sera than the prototype strain was. Monoclonal antibodies were raised against the echovirus type 25 JV-4 prototype strain. Nine clones with neutralizing activity were identified. Heterologous neutralizations of 14 clinical isolates revealed highly conserved, moderately conserved, and poorly conserved epitopes. The natural isolates differed from the prototype strain in two to four epitopes and can be classified into four different groups. We concluded that echovirus type 25, like coxsackie- and polioviruses, consists of heterogeneous viral populations with respect to biological and antigenic properties. In term of viral diagnosis, it may become increasingly difficult to identify recently isolated strains because of their antigenic variation.
Evaluation of the indoxyl acetate hydrolysis test for rapid differentiation of Campylobacter, Helicobacter, and Wolinella species
Authors: Popovic-Uroic T , Patton C M , Nicholson M A , Kiehlbauch J A
Published: January 2020

A total of 410 well-defined Campylobacter, Helicobacter, and Wolinella strains, comprising 26 named species, subspecies, and defined groups, were tested for indoxyl acetate hydrolysis by a disk method by using disks prepared at the Centers for Disease Control, Atlanta, Ga. All C. coli (43 strains), C. cryaerophila (34 strains), C. fennelliae (5 strains), C. fennelliae-Campylobacter-like organism 3 (2 strains), C. jejuni (66 strains), C. jejuni subsp. doylei (3 strains), hippurate-negative C. jejuni-C. coli (15 strains), "C. upsaliensis" (39 strains), H. mustelae (5 strains), W. curva (1 strain), and W. recta (1 strain) hydrolyzed indoxyl acetate. Four strains gave weak positive reactions, and the remaining 196 strains, which belonged to 15 species, subspecies, and defined groups, gave negative reactions. Of the 410 study strains, 246 and 125 strains were tested for indoxyl acetate hydrolysis by a disk method and a tube method, respectively, by using commercially produced disks. The disk method, regardless of source, required less time and interpretation than the tube method did. Better differentiation between Campylobacter spp. was obtained with the indoxyl acetate test than with the trimethylamine N-oxide test. The indoxyl acetate disk distinguished C. lari from C. jejuni and C. coli, C. cinaedi from C. fennelliae, and H. pylori from H. mustelae and suggested that W. succinogenes could be differentiated from W. recta and W. curva. The indoxyl acetate disk method could be performed in 5 to 30 min, was easy to read and interpret, and should be useful as a routine diagnostic test for identification of Campylobacter spp.
Identification of Giardia lamblia-specific antigens in infected human and gerbil feces by western immunoblotting
Authors: Stibbs H H , Samadpour M , Ongerth J E
Published: January 2020

Western immunoblot analysis of aqueous extracts of feces obtained from five giardiasis patients and from experimentally infected gerbils (Meriones unguiculatus) with rabbit antiserum to Giardia lamblia cysts has revealed antigens of three molecular weight groups. A stepladderlike, evenly-spaced set of strongly reactive antigens (darkest at a molecular weight [m.w.] of 55,000 to 70,000) appeared in the gerbil feces from day 4 (first experiment) or day 2 (second experiment) and lasted to about day 7 but disappeared completely by day 8 and did not reappear later. These antigenic bands were seen in gerbils infected with two isolates of G. lamblia. These bands were not revealed when antiserum to trophozoites was used as the probe, nor were they evident in specimens from the patients or in a preparation of sonicated cysts. A second group of antigens, represented by two to three low-m.w. bands of approximately 15,000 to 20,000, was evident in both the blots of gerbil feces after approximately day 8 and the specimens from the giardiasis patients. The third group of antigens revealed by blotting experiments was a high-m.w. band (approximately 110,000) which appeared on a number of days (beginning of day 8 of gerbil infection), but this band was not seen in the human specimens. A clear band corresponding to the previously reported GSA-65 antigen was not seen in either the gerbil or the human samples. Some low- and high-m.w. bands were also detected by antitrophozoite serum in the gerbil samples, but these were weak and unimpressive compared with those visualized using anticyst serum. A monoclonal antibody-based antigen capture enzyme-linked immunosorbent assay revealed that Giardia spp.-specific stool antigen rose suddenly at day 3 of gerbil infection, at the time when fecal cyst numbers began to rise rapidly.
DNA probe confirmatory test for Neisseria gonorrhoeae
Authors: Lewis J S , Kranig-Brown D , Trainor D A
Published: January 2020

A DNA probe test for the culture confirmation of Neisseria gonorrhoeae in clinical isolates was evaluated with 156 isolates of N. gonorrhoeae and 120 isolates of nongonococcal Neisseria species, organisms representative of other genera within the family Neisseriaceae, and organisms isolated on media selective for N. gonorrhoeae. The 30-min test used a chemiluminescent DNA probe that was homologous to rRNA sequences of N. gonorrhoeae. We report here a specificity and a sensitivity of 100% with the 276 clinical isolates tested, including 43 gonococcal strains that had been misidentified by other methods.
Aerobic bacteria cultured from the mouth of the American opossum (Didelphis virginiana) with reference to bacteria associated with bite infections
Authors: Howell J M , Dalsey W C
Published: January 2020

The American opossum inflicts bite injuries both when hunted for food and when accidentally provoked when handled in captivity. This study involved aerobically culturing organisms from the mouths of seven wild opossums (Didelphis virginiana). Isolates included streptococci, coagulase-positive and -negative staphylococci, Aeromonas spp., Citrobacter freundii, Eikenella corrodens, and Escherichia coli.
Eubacterium lentum ATCC 43055, a new reference strain for quality control of anaerobic susceptibility tests
Authors: Barry A L , Zabransky R J
Published: January 2020

A strain of Eubacterium lentum (ATCC 43055) was selected for quality control of anaerobic susceptibility tests. Multilaboratory collaborative studies with 13 different antimicrobial agents were reviewed, and MIC control limits were proposed for agar dilution tests with the new control strain.
Irrigation-aspiration for culturing draining decubitus ulcers: correlation of bacteriological findings with a clinical inflammatory scoring index
Authors: Ehrenkranz N J , Alfonso B , Nerenberg D
Published: January 2020

Biopsy of infected decubitus ulcers for culture disrupts tissues and may disseminate infection. Antimicrobial prophylaxis to prevent dissemination of infection may adversely affect biopsy culture results. Irrigation-aspiration to obtain submarginal specimens from draining decubitus ulcers was studied as an atraumatic, noninvasive culturing technique to serve as an alternative to biopsy in research activities. Two aspirates were obtained serially from 32 subjects; in 12 subjects, biopsies were also performed immediately. A median of 4.5 bacterial species was recovered per ulcer by irrigation-aspiration. Recent antimicrobial treatment had no evident effect on the recovery of bacterial species in general or, specifically, on the recovery of Bacteroides species. Concordance of results for both aspirates was 97.6% for aerobes and 91.8% for anaerobes, indicating no interactive methodological effect of the first irrigation-aspiration on the second. Compared with biopsy isolates for one aspirate, the sensitivity was 93% and the specificity was 99.0%; for another aspirate, the sensitivity was 94.7% and the specificity was 99.5%. The positive predictive value for either aspirate was greater than or equal to 93.9%. A weighted clinical index to score inflammatory ulcer characteristics was devised (score range, 0 to 15). In the absence of anaerobes in 15 subjects, the mean score was 6.1 +/- 3.5; in the presence of anaerobes in 17 subjects, the mean score was 9.4 +/- 3.2 (P = 0.008). The presence of aerobic gram-positive or gram-negative species did not significantly affect scores. Irrigation-aspiration for culture and clinical scoring of inflammation should permit independent serial measures of bacteriological and clinical courses of draining decubitus ulcers without patient risk or discomfort.
Antigens in electron-dense granules from Entamoeba histolytica as possible markers for pathogenicity
Authors: Muñoz M L , Lamoyi E , León G , Tovar R , Pérez-García J , De La Torre M , Murueta E , Bernal R M
Published: January 2020

In vitro interaction of Entamoeba histolytica with collagen induces intracellular formation and release of electron-dense granules (EDG) and stimulation of collagenolytic activity. Purified EDG contain 1.66 U of collagenase per mg of protein. Thus, EDG may participate in tissue destruction during invasive amebiasis. Monoclonal antibodies (MAbs) L1.1 and L7.1 reacted specifically with EDG in enzyme-linked immunosorbent assay (ELISA) and immunofluorescence and immunoelectron microscopy. MAb L7.1 immunoprecipitated three polypeptides with molecular weights of 95,000, 68,000, and 28,000 from lysates of biosynthetically labeled E. histolytica. Both MAbs recognized the pathogenic E. histolytica axenic strains HM1:IMSS, HM38:IMSS, and HK-9 but failed to react in ELISA with Entamoeba moshkovskii, Entamoeba invadens, and E. histolytica-like Laredo. In addition, MAb L7.1 reacted with one E. histolytica isolate from a symptomatic patient but did not react with four of five isolates from asymptomatic patients. EDG antigens were detected by a MAb L7.1-based ELISA in E. histolytica-containing fecal samples from symptomatic, but not asymptomatic, individuals. These results suggest that the EDG antigen detected with MAb L7.1 may be differentially expressed in pathogenic and nonpathogenic E. histolytica.
Practical and economical method for using biotinylated DNA probes with bacterial colony blots to identify diarrhea-causing Escherichia coli
Authors: Gicquelais K G , Baldini M M , Martinez J , Maggi L , Martin W C , Prado V , Kaper J B , Levine M M
Published: January 2020

A simple and economical method was developed for using biotinylated DNA probes to hybridize with bacterial colonies belonging to the various categories of diarrhea-causing Escherichia coli. Simplification and cost containment were achieved by using Whatman no. 541 filter papers instead of nitrocellulose, by minimizing the concentration of proteinase K (an expensive but necessary reagent used to pretreat the colony blots prior to hybridization with biotin-labeled DNA probes) and by reusing hybridization solution containing labeled probe DNA. After exposing the colony blots to lysing solution and steam, followed by lysozyme (1.5 mg/ml), sucrose (25%), and proteinase K (10 micrograms/ml) treatments, biotinylated probes were used to detect enterotoxigenic, enteropathogenic, enterohemorrhagic, diffuse adherence, and enteroinvasive categories of diarrhea-causing E. coli with a high level of sensitivity and specificity. Three independent observers who were experienced in reading DNA blots recorded remarkably similar results, while less satisfactory results were obtained when the blots were read by an inexperienced observer. This technique will be useful in laboratories in which radioactive isotopes are unavailable or impractical and in which budgets are restricted.
Epitopes functional in neutralization of varicella-zoster virus
Authors: Forghani B , Dupuis K W , Schmidt N J
Published: January 2020

By competition neutralization assay using monoclonal antibodies (MAbs) to varicella-zoster virus (VZV) glycoproteins (gps), we attempted to determine the topographical relationship of epitopes which are functional in VZV neutralization. MAbs against gpI interfered moderately to strongly with neutralization of MAbs against gpIII, and one antigenic domain with two distinct epitopes was identified on gpIII. Competition neutralization assays performed with MAbs to gpI revealed at least three distinct antigenic domains: the first contained two complement-dependent neutralizing epitopes; the second contained five complement-dependent neutralizing, overlapping epitopes and one nonneutralizing, nonoverlapping epitope; and the third contained one complement-enhanced neutralizing epitope. Competition neutralization assays performed with MAbs to gpIV showed one antigenic domain with two distinct epitopes which competed with nonneutralizing gpI MAbs. gpII did not interfere with neutralization of gpI, gpIII, or gpIV. Our data suggest that neutralizing and nonneutralizing MAbs can interfere with the action of viral neutralization either by inhibition or by enhancement. This report describes the epitope mapping of VZV gps by a functional biological assay.
Characterization of Shigella dysenteriae serotypes 11, 12, and 13
Authors: Wathen-Grady H G , Britt L E , Strockbine N A , Wachsmuth I K
Published: January 2020

We conducted serologic and biochemical studies on strains of three provisional Shigella serotypes. Included were 19 strains of serotype 3873-50 from three countries (1 from Mexico, 1 from Netherlands Antilles, and 17 from the United States), 13 strains of serotype 3341-55 from three countries (1 from Canada, 1 from Bulgaria, and 11 from the United States), and 19 strains of serotype 19809-73 from two countries (16 from Israel and 3 from the United States). Reactions of these strains with homologous and heterologous Shigella antisera showed that serotypes 3873-50 and 19809-73 were unique. Strains of serotype 3341-55 shared an identical antigen with Shigella boydii 18; however, they were distinguished from S. boydii 18 biochemically and with absorbed sera. Strains of the three serotypes were biochemically most similar to Shigella dysenteriae. Like other shigellae, strains from each serotype were invasive in the Serény assay. Although biochemically similar to S. dysenteriae, none of the provisional serotypes produced Shiga toxin or the biologically related toxin, Shiga-like toxin II, produced by some strains of Escherichia coli. On the basis of these results and other published findings, we support earlier proposals to add the three provisional serotypes 3873-50, 3341-55, and 19809-73 to the Shigella schema and will include them as S. dysenteriae 11, 12, and 13, respectively.
Detection of circulating excretory secretory antigens in human fascioliasis by sandwich enzyme-linked immunosorbent assay
Authors: Espino A M , Marcet R , Finlay C M
Published: January 2020

We developed a sandwich enzyme-linked immunosorbent assay to detect circulating parasite antigen in humans with fascioliasis. The assay uses antibodies against Fasciola hepatica excretory secretory (ES) antigens. A monoclonal antibody was used to capture circulating ES antigens, and a human polyclonal antibody peroxidase conjugate was used to identify circulating ES antigens. Optimal dilutions of all reagents were determined by block titration. The antigen concentration in sera from patients was estimated by comparing the optical density at 492 nm of test sera with a standard curve. All of the serum samples from 25 patients with parasitological evidence of fascioliasis had a detectable antigen concentration (more than 10 ng/ml). None of the serum samples from 80 patients infected with parasites other than F. hepatica showed a positive reaction, suggesting the absence of cross-reactions in this assay.
Phosphatase activity of staphylococci is constitutive in some species and repressed by phosphates in others
Authors: Soro O , Grazi G , Varaldo P E , Satta G
Published: January 2020

The phosphatase activities of colonies of staphylococcal strains belonging to the 10 species most frequently isolated from specimens of human origin were evaluated on media with undetermined inorganic phosphate contents and on media supplemented with known amounts of phosphates. All strains of all species tested were phosphatase positive on plates that were not supplemented with inorganic phosphates when the pH of the medium was high. In media supplemented with 0.3% phosphates at low and high pH, all strains of Staphylococcus aureus, almost all strains of S. epidermidis and S. xylosus, and none of the other species tested were phosphatase positive. Thirty S. simulans strains were grown in broths at pH 7 with and without phosphates, and the phosphatase activities of the cells were assayed at pH 5.5 and 8 in the absence of phosphates. At pH 8, all strains gave a strong positive reaction when grown in the absence of phosphates and a negative reaction when grown in the presence of these salts at 0.3%. It was concluded that all the species of staphylococci tested possess phosphatase activity and that the staphylococcal phosphatases are constitutive in some species and repressed in others.
Detection and identification of Treponema hyodysenteriae by using oligodeoxynucleotide probes complementary to 16S rRNA
Authors: Jensen N S , Casey T A , Stanton T B
Published: January 2020

Oligodeoxynucleotide probes (17 and 28 bases long) complementary to a unique region of Treponema hyodysenteriae 16S rRNA were developed. These probes bound specifically to partially purified rRNA and whole-cell rRNA of T. hyodysenteriae. No binding to partially purified rRNA or whole-cell rRNA of Treponema innocens, Treponema succinifaciens, Treponema bryantii, or Escherichia coli occurred under stringent conditions. The 28-base probe was 5 to 10 times more sensitive than the 17-base probe when hybridized with T. hyodysenteriae rRNA. The 28-base probe detected T. hyodysenteriae in the feces of experimentally inoculated pigs exhibiting clinical signs of swine dysentery.
Antigen capture assay for detection of a 43-kilodalton Mycobacterium tuberculosis antigen
Authors: Wadee A A , Boting L , Reddy S G
Published: January 2020

This study describes the development of an enzyme-linked immunosorbent assay (ELISA) to detect Mycobacterium tuberculosis antigens in body fluids. A double-antibody sandwich procedure that used human and rabbit anti-M. tuberculosis immunoglobulin G antibodies was followed. The ELISA was able to detect as little as 0.8 micrograms of protein of M. tuberculosis sonic extract. Of 253 cerebrospinal fluid specimens submitted for analysis, 11 (4.3%) false-positive results were recorded. Analysis of 317 pleural and ascitic fluid specimens resulted in 6 (1.9%) false-positive recordings. No false-negative results were recorded for any of the body fluids tested. This technique is rapid (5.5 h) and sensitive, may be developed and used in many laboratories with limited resources, and may prove useful in the diagnosis of extrapulmonary and pulmonary tuberculoses. Analysis of these body fluids by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western immunoblotting indicated that the antibody used in the ELISA detects a mycobacterial antigen of 43 kDa. Such antigens were not detected in body fluids of nontuberculous patients.
Detection of hippurate hydrolysis by Legionella spp. by using a rapid high-performance liquid chromatography method
Authors: Marmet D , Bornstein N , Fleurette J
Published: January 2020

A rapid high-performance liquid chromatography method was developed to determine hippurate hydrolysis by Legionella spp. Benzoic acid, an end product of enzymatic activity, was directly detected by high-performance liquid chromatography after 1 and 24 h of incubation in 1% sodium hippurate. Because of its sensitivity, this procedure offers more precise identification of some Legionella spp.
Antigenic and genetic analyses of human rotavirus with dual subgroup specificity
Authors: Urasawa T , Taniguchi K , Kobayashi N , Wakasugi F , Oishi I , Minekawa Y , Oseto M , Ahmed M U , Urasawa S
Published: January 2020

In our previous study (S. Urasawa, T. Urasawa, K. Taniguchi, F. Wakasugi, N. Kobayashi, S. Chiba, N. Sakurada, S. Morita, O. Morita, M. Tokieda, T. Kawamoto, K. Minekawa, and M. Oseto, J. Infect. Dis. 160:44-51, 1989) of antigenic characterization of about 300 human rotavirus (HRV) isolates collected at different localities in Japan, we found 4 HRV isolates having unique antigenic and genetic constructions. The four strains possessed both subgroup I and subgroup II antigens, serotype 3 antigen, and a long RNA electropherotype. The reactivity pattern of these four HRV isolates with three monoclonal antibodies (MAbs) directed to an outer capsid protein, VP4, and with one MAb directed to an inner capsid protein, VP2, was clearly different from those of usual subgroup II HRVs having serotype 1, serotype 3, or serotype 4 specificity and a long RNA pattern, whereas their reactivity pattern was similar to that of strain K8 (subgroup II, serotype 1), which possessed unique VP4 and VP2 proteins. RNA-RNA cross-hybridization analysis indicated that while the four isolates were genetically distinct from the two genetic groups of HRV reported previously, i.e., the Wa family (strains KU, S3, and YO) and the DS-1 family (strain S2), they were closely related to strain K8, a strain having unique antigenic and genetic properties (K. Taniguchi, K. Nishikawa, T. Urasawa, S. Urasawa, K. Midthun, A. Z. Kapikian, and M. Gorziglia, J. Virol. 63:4101-4106, 1989).
Kluyvera mediastinitis following open-heart surgery: a case report
Authors: Sierra-Madero J , Pratt K , Hall G S , Stewart R W , Scerbo J J , Longworth D L
Published: January 2020

Reports of serious infections caused by Kluyvera spp. have been rare. A case of Kluyvera sp. mediastinitis and bacteremia in a man after open-heart surgery is described. The clinical significance of Kluyvera sp. isolates was debated in early descriptions of the organism. More recent reports, as well as the present case, suggest that the pathogenic potential of this organism is being increasingly recognized by clinicians and microbiologists.
Serological typing of Branhamella catarrhalis strains on the basis of lipopolysaccharide antigens
Authors: Vaneechoutte M , Verschraegen G , Claeys G , Van Den Abeele A M
Published: January 2020

A total of 302 strains of Branhamella catarrhalis from different parts of the world were serologically typed according to their lipopolysaccharide (LPS) antigenicity. For this purpose, an inhibition enzyme-linked immunosorbent assay was developed using the following reagents: antisera raised against whole bacterial suspensions for a panel of 16 serotype strains and LPS prepared from these strains by phenol extraction. Antisera were absorbed with whole bacterial suspensions of the B. catarrhalis strains to be tested. The residual activity of the sera against the homologous LPS was determined by means of an enzyme-linked immunosorbent assay, using microdilution plates coated with LPS. Strains which gave greater than 90% reduction of activity were considered to carry the same LPS type as the serotype strain. It was shown that 93.4% of the strains tested carried one of three possible LPS types. LPS of B. catarrhalis are the rough type and have an apparent Mr of 5,500, as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
Experimental cryptosporidiosis in hamsters
Authors: Rossi P , Pozio E , Besse M G , Gomez Morales M A , La Rosa G
Published: January 2020

A new laboratory animal model for experimental cryptosporidiosis is described. Adult immunosuppressed hamsters were infected per os with 0.5 x 10(5) and 1 x 10(5) Cryptosporidium oocysts of calf origin. The mean numbers of oocysts shed per gram of feces per day and the patterns of infection are described. The susceptibility to Cryptosporidium infection, the total number of oocysts shed (a thousand times the infective dose), and the ease of handling in laboratory conditions make hamsters a good animal model for cryptosporidiosis.
Canine ehrlichiosis in Connecticut
Authors: Magnarelli L A , Litwin H J , Holland C J , Anderson J F , Ristic M
Published: January 2020

The first case of canine ehrlichiosis in Connecticut is reported. A female Brittany spaniel from Milford presented with lethargy, anorexia, fever, petechiae, splenomegaly, thrombocytopenia, anemia, elevated serum alkaline phosphatase, lymphopenia, and hypoalbuminemia. Serologic analysis revealed antibodies to Ehrlichia canis (titer, 1:2,560). This documents a more northern geographic distribution in the United States for this infectious agent than had previously been suspected.
Identification of an antigenic subtype of eastern equine encephalitis virus isolated from a human
Authors: Calisher C H , Karabatsos N , Foster J P , Pallansch M , Roehrig J T
Published: January 2020

Eastern equine encephalitis (EEE) virus was isolated from the cerebrospinal fluid of a 6-year-old male who had clinically diagnosed aseptic meningitis and subsequently died. Several standard serologic tests that use polyclonal antibody and indirect immunofluorescence and hemagglutination inhibition tests that use monoclonal antibody provided evidence that the isolate was an antigenic subtype of prototype North American EEE virus. We believe that this is the first evidence of an antigenic subtype of EEE virus.
Cultivation and characterization of spirochetes from cerebrospinal fluid of patients with Lyme borreliosis
Authors: Karlsson M , Hovind-Hougen K , Svenungsson B , Stiernstedt G
Published: January 2020

Attempts were made to culture spirochetes from cerebrospinal fluid samples of 105 patients suspected of having Lyme borreliosis with neurological complications. At the final evaluation, only 38 patients fulfilled the criteria of neuroborreliosis. Spirochetes were cultured from cerebrospinal fluid samples of four of these patients. All four patients had pleocytosis in their cerebrospinal fluid and a history of neurological symptoms of only 4 to 10 days in duration. Two of them had no detectable antibodies against any of the isolated spirochetes in their cerebrospinal fluid, both when tested with an enzyme-linked immunosorbent assay and when tested by immunoblotting. An antibody reaction against the homologous isolate that was distinctly stronger than that against the heterologous isolates was found in the serum and cerebrospinal fluid samples from one patient. The cells of the isolates were morphologically similar and showed a very similar protein pattern when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Cells of all isolates reacted with the monoclonal antibodies H5332 and H9724, which also react with Borrelia burgdorferi B31, the type strain. One isolate lost a major protein of 23 kilodaltons after subcultivation for 4 months. We conclude that isolation of spirochetes from cerebrospinal fluid might prove successful in clinically selected cases of Lyme borreliosis.
Comparison of directigen RSV with viral isolation and direct immunofluorescence for the identification of respiratory syncytial virus
Authors: Waner J L , Whitehurst N J , Todd S J , Shalaby H , Wall L V
Published: January 2020

An enzyme immunoassay membrane test (Directigen RSV) for the detection of respiratory syncytial virus in clinical specimens was compared prospectively with isolation in cell culture and direct immunofluorescence (IF). A total of 315 nasopharyngeal wash specimens from pediatric patients were examined. Directigen RSV was 86.1% sensitive and 91.3% specific for specimens positive by isolation in cell culture and/or IF, with 88.6% agreement. The false-positive rate was 16%; 2 of 20 specimens giving false-positive reactions by Directigen RSV were true-positives by blocking assay. Twenty-seven specimens (8.5%) whose results were initially uninterpretable by Directigen RSV due to filtration difficulties were diluted and upon retesting produced acceptable results. Sixty-three viral isolates and/or IF identifications of virus antigens representing seven virus groups other than respiratory syncytial virus were also found; cross-reactions between Directigen RSV and other viruses were not observed. Directigen RSV will be useful as an immediate procedure and in facilities lacking a comprehensive virology laboratory.
Detection of Borrelia burgdorferi infection in Ixodes dammini ticks with the polymerase chain reaction
Authors: Persing D H , Telford S R , Spielman A , Barthold S W
Published: January 2020

The polymerase chain reaction (PCR) was used to amplify DNA sequences of the etiologic agent of Lyme disease, Borrelia burgdorferi, and was applied to the detection of the spirochete in its tick vector. The target for PCR amplification was the OSP-A gene of strain B31; analysis of isolates from different geographical areas indicated that this gene could be used to identify most North American isolates. These methods were extended to the analysis of colony-derived and field-collected Ixodes dammini. OSP-A-specific sequences were identified in 15 of 15 colony-derived nymphal ticks that had fed previously on an infected animal; no such amplification products were detected in 8 control ticks. Segregated midgut tissues of field-collected adult and nymphal ticks from Nantucket Island, Mass., and the Crane Reserve, Ipswich, Mass., were examined by both direct fluorescent-antibody (DFA) staining and PCR. The DFA technique identified 16 infected ticks of 30 paired specimens; 15 of these specimens were positive by PCR. One specimen was positive by PCR that was DFA negative. Both live whole ticks and desiccated dead specimens were suitable for this analysis. Because only five ticks are suitable for DFA analysis, the use of PCR may extend the range of specimens that can be analyzed for the presence of the Lyme spirochete.
Interpretive criteria and quality control limits for ceftibuten disk susceptibility tests. Collaborative Antimicrobial Susceptibility Testing Group
Authors: Barry A L , Jones R N
Published: January 2020

In vitro studies were undertaken to evaluate susceptibility tests with 30-micrograms ceftibuten disks. The following interpretive criteria were proposed: less than or equal to 17 mm for resistance (MIC, greater than or equal to 32 micrograms/ml) and greater than or equal to 21 mm for susceptibility (MIC, less than or equal to 8.0 micrograms/ml). A multilaboratory quality control study led to the conclusion that Escherichia coli ATCC 25922 should provide zones 29 to 35 mm in diameter.
Serological responses to papillomavirus group-specific antigens in women with neoplasia of the cervix uteri
Authors: Dillner L , Moreno-Lopez J , Dillner J
Published: January 2020

Certain types of human papillomaviruses have been linked to the development of carcinoma of the cervix uteri. We have analyzed 114 serum specimens from women with cervical intraepithelial neoplasia (CIN) or carcinoma of the cervix uteri for the presence of serum antibodies against purified, disrupted bovine papillomavirus (BPV). The titers of immunoglobulin A (IgA) antibodies against BPV were slightly elevated (P less than 0.025) in the sera from CIN or cervical carcinoma patients compared with the titers of 139 serum specimens from sex- and age-matched healthy controls. In contrast, both the IgG and IgM serum antibody titers against BPV were significantly decreased for CIN and cervical carcinoma patients compared with those of healthy controls (P less than 0.001 and P less than 0.005, respectively). These results suggest that the difference between IgA and IgG or IgM antibodies to papillomavirus group-specific antigens may represent interesting serological parameters that could possibly be used in the epidemiologic study of women at risk for CIN.
Whole-cell protein patterns of nonhemolytic group B, type Ib, streptococci isolated from humans, mice, cattle, frogs, and fish
Authors: Elliott J A , Facklam R R , Richter C B
Published: January 2020

Whole-cell protein and physiological patterns of nonhemolytic group B, type Ib, streptococci isolated from humans, cattle, frogs, fish, and mice were compared. Isolates from humans, fish, and mice were identical. Only minor differences were seen in the isolates from human, bovine, and frog sources. Nonhemolytic group B streptococci from humans, fish, and mice and, to a lesser extent, from cattle and frogs share several characteristics, including a high similarity in proteins (on the basis of molecular weight); this suggests that they may have a common ancestry.
Characterization of antibody reactivity to human T-cell lymphotropic virus types I and II using immunoblot and radioimmunoprecipitation assays
Authors: Hartley T M , Khabbaz R F , Cannon R O , Kaplan J E , Lairmore M D
Published: January 2020

We have characterized the immunoreactivity to human T-cell lymphotropic virus type I (HTLV-I) among 26,983 persons of various seroprevalence groups by using enzyme immunoassay, immunoblot (IB), and radioimmunoprecipitation assays (RIPA) in accordance with Public Health Service recommended guidelines for the interpretation of serologic test results for HTLV-I infection. IB-indeterminate serum specimens (n = 178) were reactive to HTLV-I gag proteins, and no serum contained only env reactivity. Overall, RIPA resolved 40% of IB-indeterminate serum samples; however, the probability that RIPA would confirm IB-indeterminate samples depended on the seroprevalence of the population tested. HTLV-I gag p19-only reactivity on IB was not a reliable marker of HTLV-I infection, while gag p24 reactivity on IB was clearly associated with positive seroreactive specimens. IB and RIPA tests did not clearly distinguish between HTLV-I and HTLV-II seroreactivities. These data emphasize that patterns of immunoreactivity to HTLV-I antigens are dependent upon the seroprevalence of the risk groups tested. In addition, RIPA detected antibodies to env proteins present in low titer in a substantial number of IB gag-only reactive sera and resolved the HTLV-I antibody status of these sera.
Purification and characterization of a pilin specific for Brazilian purpuric fever-associated Haemophilus influenzae biogroup aegyptius (H. aegyptius) strains
Authors: Weyant R S , Bibb W F , Stephens D S , Holloway B P , Moo-Penn W F , Birkness K A , Helsel L O , Mayer L W
Published: January 2020

Brazilian purpuric fever (BPF) is a recently described fatal pediatric disease caused by systemic infection with Haemophilus influenzae biogroup aegyptius. Previous studies have shown that all H. influenzae biogroup aegyptius strains isolated from BPF cases and case contacts share several unique phenotypic and genotypic characteristics that differentiate them from other H. influenzae biogroup aegyptius strains isolated from conjunctivitis cases in Brazil. One key characteristic of this BPF clone is reactivity in a BPF-specific monoclonal antibody enzyme-linked immunosorbent assay. We have purified and partially characterized a pilin, referred to as the 25-kilodalton (kDa) protein. Aggregates of this protein contain a heat-labile epitope which is recognized by a monoclonal antibody used in the BPF-specific enzyme-linked immunosorbent assay. The protein has a molecular weight of approximately 25,000, is insoluble in most detergents, and fractionates with outer membrane vesicles after LiCl extraction. Biochemical analysis of the 25-kDa protein shows it to have an amino acid composition similar but not identical to that of the H. influenzae type b pilin. The sequence of 20 N-terminal amino acids of the 25-kDa protein shows almost complete homology with the N terminus of the H. influenzae type b pilin and the types 1 and P pilins of Escherichia coli. Transmission electron microscopic analysis of the purified protein shows the presence of filamentous structures similar in morphology to those of H. influenzae pili. Reactivity between the 25-kDa protein and the BPF-specific monoclonal antibody is demonstrated by Western blotting (immunoblotting) and colloidal gold-enhanced immunoelectron microscopy. Hemadsorption analysis shows that expression of this protein is associated with increases in piliated cells and enhanced binding of these cells to human erythrocytes. These studies indicate that expression of the 25-kDa protein is a characteristic unique to the BPF clone and suggest that this protein plays a role in the pathogenesis of BPF.
Effect of iron on production of a possible virulence factor by Plesiomonas shigelloides
Authors: Gardner S E , Fowlston S E , George W L
Published: January 2020

Plesiomonas shigelloides, when grown in an iron-poor medium (syncase), produces a substances that causes elongation of Chinese hamster ovary (CHO) cells similar to that produced by cholera toxin. When syncase is supplemented with iron, the ability of P. shigelloides (but not of Vibrio cholerae) to produce this elongation of CHO cells is lost. Iron depletion of the growth medium appears to be essential for the CHO cell elongation produced by P. shigelloides but is not essential for the production of toxin by V. cholerae. The possible role of iron regulation of this potential virulence factor warrants further study.
Rapid screening of urine for bacteria and cells by using a catalase reagent
Authors: Berger S A , Bogokowsky B , Block C
Published: January 2020

Five hundred urine samples were tested for cells and bacteria by using a commercial dipstick, a catalase screening device, standard culturing, and chamber counts. The sensitivities of the catalase test were 83% for all samples and 97.6% for specimens containing significant numbers of leukocytes and bacteria. The catalase screening test is simple to perform and should prove useful for the detection of urinary tract infections.
Major subtypes of invasive Haemophilus influenzae from 1983 to 1985 in Atlanta, Ga
Authors: Elliott J A , Pigott N , Cochi S L , Facklam R R
Published: January 2020

We compared outer membrane protein (OMP) patterns of Haemophilus influenzae isolated in metropolitan Atlanta, Ga., from July 1983 to June 1985. Of 74 randomly selected H. influenzae serotype b, biotype I, isolates (24% of the total number of H. influenzae, and 32% of the total number of H. influenzae serotype b, biotype I, isolates), 66 (89.2%) had the same OMP pattern. Of the remaining eight, five (6.7%) had an identical OMP pattern. The other three isolates had separate and distinct patterns. A greater diversity of OMP patterns was found with H. influenzae serotype b, biotype II, and nonserotypeable H. influenzae. Of the 18 H. influenzae serotype b, biotype II, isolates (5.8% of the total number of H. influenzae isolates), 1 had an OMP pattern similar to that of the predominate biotype I OMP type, 6 (33% of the biotype II) had the same pattern, and 11 had heterogeneous patterns. Of the 19 recoverable, nonserotypeable biotype II isolates (6.8% of the total number of H. influenzae), 18 had different OMP patterns, and no pattern was similar to those observed with serotype b. These findings indicate that most H. influenzae strains isolated during this 2-year period were indistinguishable by serotype, biotype, or OMP patterns.
Administration of bacterial lipopolysaccharide elicits circulating tumor necrosis factor-alpha in neonatal calves
Authors: Adams J L , Semrad S D , Czuprynski C J
Published: January 2020

The presence of tumor necrosis factor-alpha (TNF-alpha) during endotoxemia in ruminants has not been reported previously. In this study, we detected the in vivo release of bovine TNF-alpha by using WEHI-164 murine fibrosarcoma cells as targets in an 18-h cytotoxicity assay. Treatment of the WEHI-164 cells with 1 microgram of actinomycin D (dactinomycin) enhanced approximately twofold the susceptibility of the cells to TNF-alpha activity. TNF-alpha activity in sera from neonatal calves injected intravenously with 2.7 micrograms of Escherichia coli lipopolysaccharide (LPS) increased rapidly within the first 2 h postinjection and then declined until it was undetectable by 4 h postinjection. Sera taken before LPS administration had no TNF-alpha activity. LPS (10 micrograms/ml) and fetal, newborn, and pooled adult bovine sera alone and in combination had no direct cytotoxic effects on WEHI-164 cells. TNF-alpha cytotoxic activity is probably not due to the presence of interleukin-1 (IL-1), alpha interferon, or gamma interferon in the sera since recombinant human IL-1, natural bovine IL-1, and recombinant bovine alpha and gamma interferons had no direct cytotoxic effects on WEHI-164 cells. A monoclonal antibody that neutralizes recombinant human TNF-alpha significantly reduced the cytotoxic activity of sera from LPS-injected calves.
Human malignant melanoma cell line (HMV-II) for isolation of influenza C and parainfluenza viruses
Authors: Moriuchi H , Oshima T , Nishimura H , Nakamura K , Katsushima N , Numazaki Y
Published: January 2020

HMV-II, a human malignant melanoma cell line, was compared with other cell lines (MDCK, Vero, and LLC-MK2) and primary cultures of monkey kidney (PMK) cells for the isolation and quantification of influenza and parainfluenza viruses. HMV-II cells were superior to MDCK and LLC-MK2 cells in quantification of the influenza C virus and were used successfully in the isolation of the virus from clinical specimens. The HMV-II cell line was also more sensitive for isolating parainfluenza viruses from clinical specimens than were Vero and PMK cells; there was, however, no significant difference in the quantification of the viruses among these cultures. As far as influenza A and B viruses were concerned, the HMV-II cell line was significantly less sensitive than MDCK cells, and no virus was isolated from clinical specimens with HMV-II cells. Thus, HMV-II cells are useful for the isolation of influenza C and parainfluenza viruses as an alternative to embryonated hen's eggs and PMK cells.
Evaluation of a human immunodeficiency virus test algorithm utilizing a recombinant protein enzyme immunoassay
Authors: Lepine D G , Neumann P W , Frenette S L , O'Shaughnessy M V
Published: January 2020

Many truly human immunodeficiency virus (HIV) antibody-negative serum samples may be unnecessarily subjected to costly and time-consuming Western blots (immunoblots). An investigation was undertaken to evaluate the efficiency of using a recombinant protein-based enzyme immunosorbent assay (EIA; Cambridge BioScience [CBC] Recombigen HIV EIA) as an adjunct to whole viral lysate EIA. A total of 2,212 serum samples which had been screened by viral lysate EIA were tested by CBC EIA in parallel with the Western blot. The sensitivity and specificity of the CBC kit were 99.9 and 99.7%, respectively. Positive and negative predictive values were 99.7 and 99.9%, respectively. The high sensitivity of this kit and its high negative predictive value make it an attractive addition to an HIV testing algorithm by reducing the number of Western blot tests on truly antibody negative serum samples.
Use of polymerase chain reaction for detection of Chlamydia trachomatis
Authors: Ostergaard L , Birkelund S , Christiansen G
Published: January 2020

A polymerase chain reaction (PCR) assay was developed for detection of Chlamydia trachomatis DNA. From the published sequence of the common C. trachomatis plasmid, two primer sets were selected. Detection of amplified sequences was done by agarose gel electrophoresis of cleaved or uncleaved amplified sequences, Southern hybridization, or dot blot analysis. The PCR assay was optimized and, after 40 cycles of amplification with primer set II, demonstrated a sensitivity of 10(-17) g of DNA, which corresponds to the detection of one copy of the plasmid. Because of the high sensitivity, we developed a closed system in which airborne contamination was minimized. Analysis of 228 clinical samples tested by cell culture, IDEIA enzyme immunosorbent assay (Medico-Nobel, Boots-Celltech Ltd., Berkshire, United Kingdom), and PCR showed a sensitivity of 100%, a specificity of 93% when PCR was compared with cell culture, and a corrected specificity of 99% when PCR was compared with cell culture or IDEIA.
Stability of viable-bacterium counts in liquid media used for preparation of inocula and subsequent impact on antimicrobial susceptibility test results
Authors: Hindler J A , Gonzalez A H , Drake T A
Published: January 2020

We evaluated the consequences of prolonging the time between initial bacterial inoculum suspension preparation and susceptibility test inoculation. Extending the current National Committee for Clinical Laboratory Standards-recommended time of 15 min between suspension preparation and test inoculation should allow laboratories more flexibility to optimize efficiency from the standpoint of workflow. We assessed the length of time for which viable-bacterium counts remain stable in three liquid media at room temperature. Fifty isolates were examined in water, saline, and cation-supplemented Mueller-Hinton broth (CSMHB). Disk diffusion and microdilution MIC tests were performed on nine of these. Our results suggest that directly prepared inoculum suspensions of members of the family Enterobacteriaceae, Pseudomonas aeruginosa, staphylococci, and enterococci can be held for up to 6 h in water or saline prior to inoculation of disk diffusion and MIC tests without compromising test accuracy. The same organisms can be held for at least 1 h in CSMHB. Viridans group streptococci can be held for up to 6 h in saline and CSMHB and for up to 3 h in water. Similarly, Haemophilus influenzae and Streptococcus pneumoniae isolates may be held in CSMHB for up to 3 h. Because of an early decrease in viable-bacterium counts in water and saline with some H. influenzae and S. pneumoniae isolates, we recommend that National Committee for Clinical Laboratory Standards recommendations be followed for these species.
Cross-reactivity of nonspecific treponemal antibody in serologic tests for Lyme disease
Authors: Magnarelli L A , Miller J N , Anderson J F , Riviere G R
Published: January 2020

Serum samples obtained from 59 persons who had acute necrotizing ulcerative gingivitis, periodontitis, syphilis, or Lyme disease were tested against Treponema phagedenis biotype Reiter, Treponema denticola, Treponema vincentii, and Treponema scoliodontum by indirect fluorescent-antibody staining methods. Although there were positive reactions for sera representing each of these study groups and for 20 (13%) of 156 samples collected from the general population (premarital screening for syphilis), titration endpoints were relatively low (less than or equal to 1:256). Serum samples from 18 persons who had gingivitis or periodontitis but no history of Lyme borreliosis were tested by enzyme-linked immunosorbent assay for antibodies to Borrelia burgdorferi. Of these, five (28%) had immunoglobulin M antibody and four (22%) contained immunoglobulin G antibodies to this spirochete. Adsorption with either sorbent commercially prepared from T. phagedenis biotype Reiter or with washed, whole cells of T. phagedenis biotype Reiter reduced cross-reactivity in the enzyme-linked immunosorbent assay for Lyme borreliosis.