bluuebunny/crossref_metadata_2025 huggingface dataset:
Latex agglutination test for detection of antibodies to Toxoplasma gondii
Authors: Mazumder P , Chuang H Y , Wentz M W , Wiedbrauk D L
Published: January 2020

A resurgence of interest in Toxoplasma gondii has occurred because this coccidian parasite causes lethal infections in immunologically compromised hosts and is responsible for at least 3,000 congenitally infected infants in the United States annually. Thus, rapid, specific, and inexpensive serologic tests are required for routine screening of patients, especially pregnant women. We have developed a latex agglutination test for antibodies to T. gondii which utilizes covalently coupled T. gondii antigens. When compared with an indirect immunofluorescence assay, the latex test had a sensitivity of 94% and specificity of 100%. Compared with an enzyme-linked immunosorbent assay, the latex test had 86% sensitivity and 100% specificity. When testing samples which exhibited nonspecific polar staining by the immunofluorescence assay, the enzyme-linked immunosorbent assay had a 50% false-positive rate, whereas the latex agglutination test yielded no false-positive results. Thus, the latex agglutination test provided an efficacious method for routine serological screening for antibodies to T. gondii.
Evaluation of Abbott TESTPACK ROTAVIRUS with clinical specimens
Authors: Marchlewicz B , Spiewak M , Lampinen J
Published: January 2020

TESTPACK ROTAVIRUS was compared with the Pathfinder Rotavirus assay for the detection of rotaviral antigens in feces from 137 patients. The symptomatology and ages of the patients varied from no symptoms to watery diarrhea and from less than 3 months to greater than 60 years of age. The sensitivity and specificity, before resolution, were 89.2 and 90.0%, respectively. After resolution with a blocking reagent, the results were 100% sensitivity and 98.9% specificity. The performance was similar for all age groups and in both symptomatic and asymptomatic patients. Abbott TESTPACK ROTAVIRUS is a rapid (10-min), easy-to-perform assay for the detection of rotaviral antigens and has the sensitivity and specificity of enzyme-linked immunosorbent assays that require several hours to perform.
Development of serum antibodies of the immunoglobulin G class and subclasses against the capsular polysaccharide of Haemophilus influenzae type b in children and adults with invasive infections
Authors: Claesson B A , Lagergård T , Trollfors B
Published: January 2020

The development of total immunoglobulin G (IgG) antibodies and antibodies of the four IgG subclasses in serum against Haemophilus influenzae type b capsular polysaccharide (CPS) was studied in 24 children and 11 adults with invasive Haemophilus influenzae type b infections, by using an enzyme-linked immunosorbent assay. None of the 8 children aged 10 months or younger had increases in the IgG class or in any of the IgG subclasses. In contrast, 14 of 16 children between 10 months and 6 years of age and 10 of 11 adults had significant increases in total IgG, IgG1, or IgG2 antibodies in various combinations, but none of them had increases in IgG3 or IgG4 antibodies. The increases in IgG1 and IgG2 antibodies in the children were of similar magnitudes. Of 11 adult patients, 9 had significant increases in IgG2 antibodies, while only 4 had increases in IgG1 antibodies. In conclusion, this study shows that children younger than approximately 1 year have no IgG response to H. influenzae type b CPS, while individuals above this age have a mixed IgG1 and IgG2 response.
Genome variants of human adenovirus types 40 and 41 (subgroup F) in Japan
Authors: Shinozaki T , Araki K , Kobayashi M , Fujita Y , Abe T , Ushijima H
Published: January 2020

Nineteen strains of fecal adenovirus were typed as adenovirus type 40 (Ad40) by restriction enzyme analysis of the DNA, using SmaI. The DNAs of the strains were further compared by digestion with four other enzymes, namely, HindIII, EcoRI, BglI, and BamHI. At least two different restriction profiles were found to exist for BamHI. Of the 19 strains studied, 17 had the same combination of profiles for all of the enzymes. Thirteen strains of fecal adenovirus were typed as Ad41 by restriction enzyme analysis of the DNA, using SmaI. The DNAs of the strains were further compared by digestion with four other enzymes, as for Ad40. At least two different restriction profiles were found to exist for each enzyme. Of 13 strains, 9 had the same cleavage sites for all of the enzymes, indicating a close relationship. Two strains were genome variants which deviated from the common genome on digestion of the DNA with HindIII. One strain was a genome variant which deviated from the common genome on digestion of the DNA with HindIII and EcoRI. The last one had a combination of restriction sites which was not identical to those of other strains on digestion of the DNA with SmaI. All 13 Ad41 strains studied were easily distinguished from Ad40 strains by using any of the five restriction enzymes.
Monoclonal antibodies produced in BALB.B10 mice define new antigenic determinants in culture filtrate preparations of Mycobacterium tuberculosis
Authors: Worsaae A , Ljungqvist L , Heron I
Published: January 2020

A panel of monoclonal antibodies were derived from BALB.B10 mice immunized with a culture filtrate from Mycobacterium tuberculosis H37Rv. Of these antibodies, 10 were examined more closely for antigen specificity and interspecies reactivity. Six antibodies were used as immunosorbents for affinity purification of their corresponding antigens. Two monoclonal antibodies (HBT 2 and HBT 11) reacted with a 17-kilodalton antigen, and a competition assay showed that these antibodies are directed against the same epitope or against epitopes that are sterically very close to each other. Monoclonal antibody HBT 12 reacted with the same molecule with which a previously described 38-kilodalton reactive antibody reacted but was directed against a different epitope. Antibody HBT 10 reacted with a culture filtrate of M. tuberculosis but not of Mycobacterium bovis BCG. This latter finding was further studied by testing different preparations of M. tuberculosis H37Rv antigens and, additionally, culture filtrates of four M. tuberculosis and two BCG strains. Interspecies reactivity was assayed by immunoblotting and revealed that the majority of the monoclonal antibodies were specific to M. tuberculosis complex.
Stability of St. Louis encephalitis viral antigen detected by enzyme immunoassay in infected mosquitoes
Authors: Tsai T F , Happ C M , Bolin R A , Montoya M , Campos E , Francy D B , Hawkes R A , Roehrig J T
Published: January 2020

The use of enzyme immunoassay to detect St. Louis encephalitis (SLE) viral antigen in vector mosquitoes enhances the effectiveness of surveillance because infected mosquitoes can be identified more rapidly than with conventional virus isolation systems and because it is a simple and accessible procedure. Infectivity among mosquitoes experimentally infected with SLE virus was lost within 24 h after the mosquitoes were stored at 27 degrees C and 80% relative humidity; however, viral antigen remained stable under these conditions and could be detected by enzyme immunoassay 2 weeks later. Desiccation further extended the period during which antigen could be detected to 6 weeks. Absorbances were higher in infected mosquitoes stored at 27 degrees C than in mosquitoes frozen continuously. Absorbances in infected mosquitoes also increased after repeated freezing and thawing and sonication. Both phenomena may be related to the release of antigen from decaying or disrupted cells. The relative stability of SLE viral antigen at ambient temperatures lends flexibility to schemes which use direct antigen detection to identify vectors. Surveillance systems can be designed without regard to collecting living mosquitoes, and a cold chain in unnecessary to preserve specimens, thus reducing the cost of surveillance and expanding the geographic areas to which it is accessible.
In vitro susceptibilities and biotypes of Candida albicans isolates from the oral cavities of patients infected with human immunodeficiency virus
Authors: Korting H C , Ollert M , Georgii A , Fröschl M
Published: January 2020

Candida albicans strains were isolated from the oral cavities of 62 human immunodeficiency virus (HIV)-infected patients at different stages of HIV infection. Only patients with persistent generalized lymphadenopathy-acquired immunodeficiency syndrome (AIDS)-related complex or full-blown AIDS showed typical clinical symptoms for oral candidiasis. In general, the microbiological recovery of Candida strains from the oral cavity increased with more advanced stages of HIV infection. The antifungal activity of ketoconazole, itraconazole, nystatin, amphotericin B, and flucytosine against all 62 strains was evaluated by means of a photometer-read broth microdilution method for determination of the 30% inhibitory concentrations of the drugs. The 95% ranges of 30% inhibitory concentrations were as follows: less than or equal to 0.063 to 32 micrograms/ml for ketoconazole, less than or equal to 0.063 to 8 micrograms/ml for itraconazole, 0.5 to 4 micrograms/ml for nystatin, less than or equal to 0.063 to 4 micrograms/ml for amphotericin B, and less than or equal to 0.063 to 8 micrograms/ml for flucytosine. Two strains were resistant to flucytosine, one was resistant to ketoconazole, and three were resistant to itraconazole. Isolates from patients with full-blown AIDS showed significantly less susceptibility to itraconazole, amphotericin B, and flucytosine. Strains were biotyped by using the API 20C carbohydrate assimilation system. The major biotype accounted for 63.9% of the isolates. At repeated evaluation, a change in biotype pattern was seen in 27.3%.
Borrelia burgdorferi infection surrounding La Crosse, Wis
Authors: Callister S M , Agger W A , Schell R F , Ellingson J L
Published: January 2020

This investigation defined the extent of Borrelia burgdorferi infection surrounding La Crosse, Wis. White-footed mice, Peromyscus leucopus or P. maniculatis, were captured from sites in Wisconsin, Minnesota, and Iowa and cultured for B. burgdorferi to define the local boundaries of the midwestern Lyme disease area. All foci of B. burgdorferi infection (N1, N2, N3, and N4) were located north of interstate highway 90 except focus S2, which was south of the highway near Fort McCoy, Wis. The interstate highway may have been a barrier to deer movement which slowed the southward dispersal of Ixodes dammini. B. burgdorferi was isolated from 12 (63%) of the mice captured from site N4, which was adjacent to the western border of Fort McCoy. Unexpectedly, no B. burgdorferi-infected mice were isolated at site N0, located north of interstate highway 90 and enclosed by areas in which B. burgdorferi infection is endemic. This site is surrounded by natural barriers which may have slowed the spread of I. dammini by deer. The Wisconsin area in which B. burgdorferi is endemic should now include the surrounding area north of interstate highway 90 west from Fort McCoy to the Mississippi River. Additional studies are needed to define the rapidity, limits, and means of I. dammini dispersal into southern Wisconsin.
Enzyme-linked immunosorbent assay with Brucella native hapten polysaccharide and smooth lipopolysaccharide
Authors: Alonso-Urmeneta B , Moriyón I , Díaz R , Blasco J M
Published: January 2020

Brucella melitensis native haptens (NH) are polysaccharides identical to the O-side chain of the smooth lipopolysaccharide (S-LPS) (E. Moreno, H. Mayer, and I. Moriyón, Infect. Immun. 55:2850-2853, 1987) which precipitate with sera from infected cattle but not from strain 19-vaccinated cattle. In the present work, NH was extracted by the hot-water method (R. Díaz, J. Toyos, M.D. Salvo, and M.L. Pardo, Ann. Rech. Vet. 12:35-39, 1981) and purified free of S-LPS and protein. Purified NH lacked the ability to coat polystyrene and sheep erythrocytes. In contrast, NH acylated with stearoyl chloride bound to both polystyrene and erythrocytes. By hemagglutination and enzyme-linked immunosorbent assay (ELISA), S-LPS and acylated NH gave similar results with blood sera from brucellosis-free, strain 19-vaccinated, and infected cattle. Moreover, a significant correlation between the results of NH ELISA and S-LPS ELISA was demonstrated with milk sera. However, in a competitive ELISA with milk sera, S-LPS in the liquid phase abrogated the binding of antibodies to acylated NH adsorbed to polystyrene, while NH in the liquid phase did not influence the binding of antibodies to polystyrene-adsorbed S-LPS. It is hypothesized that the different precipitations of NH and S-LPS with sera from infected or strain 19-vaccinated cattle are due to differences in the affinity of the antibodies produced upon vaccination or infection and in the physical state of aggregation of NH and S-LPS in aqueous solutions.
Improved Syva MicroTrak Chlamydia trachomatis direct test method
Authors: Judson B A , Lambert P P
Published: January 2020

Recent changes in the MicroTrak Chlamydia trachomatis Direct Specimen Test (Syva Company, Palo Alto, Calif.) have led to improved product performance. The use of the recommended cervical cytology brush can significantly increase the number of endocervical cells collected, and fixation with methanol increases the intensity of elementary-body staining in many specimens.
Microtiter method for MIC testing with spherule-endospore-phase Coccidioides immitis
Authors: Hector R F , Zimmer B L , Pappagianis D
Published: January 2020

A method was developed for susceptibility testing with spherule-endospore-phase Coccidioides immitis by using a microtiter format. Isolated endospores were used to inoculate wells containing modified Converse medium with various concentrations of azole or nikkomycin antifungal substances which then were sealed with an acetate film. The plate was incubated at 37 degrees C with shaking for 96 h, after which the control wells had visible turbidity and endpoints were discernible. Microscopic examination revealed that both control and treatment wells maintained cells predominantly in the spherule-endospore phase of growth.
Nonisotopic oligomeric probes for the human enteroviruses
Authors: Rotbart H A , Eastman P S , Ruth J L , Hirata K K , Levin M J
Published: January 2020

Oligonucleotide probes were prepared from highly conserved regions of human enteroviral genomes. These reagents were labeled with either 32P or alkaline phosphatase and were successfully used in blot assays to detect a wide variety of human enteroviruses, proving the potential utility of oligomeric sequences as pan-enteroviral probes. The availability of nonisotopic probes will ultimately make a hybridization assay for enteroviruses easier, shorter, and more adaptable for routine diagnostic laboratories.
Multicenter comparative evaluation of two rapid microscopic methods and culture for detection of Chlamydia trachomatis in patient specimens
Authors: Tilton R C , Judson F N , Barnes R C , Gruninger R P , Ryan R W , Steingrimsson O
Published: January 2020

Four hundred and seventy-three men and women at high risk for sexually transmitted disease were tested for the presence of Chlamydia trachomatis in the urethra or the endocervix. Four groups were involved in this multicenter study of two direct fluorescent-antibody microscopy tests, Kallestad Pathfinder and Syva Microtrak, compared with culture techniques. Results from the test sites indicated that there was no significant difference overall in the sensitivity and specificity of the two test kits. However, there was some interlaboratory variation seen in the sensitivity of the microscopy, but little difference in the specificity. Either kit could be an effective screening method for C. trachomatis in high-risk populations.
New porcine rotavirus serotype antigenically related to human rotavirus serotype 3
Authors: Nagesha H S , Holmes I H
Published: January 2020

Serotyping of porcine rotaviruses isolated in MA104 cells from Australian piglets with diarrhea showed that two strains belonged to serotype 3 and one strain was antigenically similar to the OSU strain of porcine rotavirus (serotype 5). In addition, neutralizing antibodies to human rotavirus serotype 4 (ST-3 strain) were detected in serum samples from sows in one area, and so it seems probable that porcine rotaviruses of at least three serotypes occur in Australia.
Simple method for rapid diagnosis of catheter-associated infection by direct acridine orange staining of catheter tips
Authors: Zufferey J , Rime B , Francioli P , Bille J
Published: January 2020

Direct acridine orange (AO) staining was used to detect bacteria adherent to intravascular catheters (IVC). Samples from 710 IVC tips were first cultured on blood agar plates by a semiquantitative technique and then independently colored with AO and screened dry at a magnification of x100 for 3 min. In the absence of fluorescence, they were considered negative. When fluorescence was present, they were further examined for the presence of microorganisms at x1,000 with immersion oil. Of 710 IVC tips, 37 (5.2%) were positive upon culture (greater than or equal to 15 colonies) and 673 were negative (640 were sterile and 33 [4.6%] had 1 to 14 colonies). The AO sensitivity was 84%, and the AO specificity was 99%. When restricted to the 212 long IVC, AO sensitivity rose to 94%. AO staining was positive in all cases of catheter-associated bacteremia. The negative predictive value of the preliminary screening at x100 was 99.5%. The direct examination of IVC tips stained by AO appears to be a simple and rapid method for diagnosing IVC-associated infections. In addition, AO staining is easier to perform than Gram staining.
Isolation of three distinct parasite populations expressing different circumsporozoite proteins from the Philippine strain of Plasmodium knowlesi
Authors: Cochrane A H , Gwadz R W , Kamboj K K
Published: January 2020

Approximately 50% of the sporozoites of the Philippine (P) strain of Plasmodium knowlesi were found to react with monoclonal antibodies, produced against sporozoites of the H strain of P. knowlesi, by indirect immunofluorescence. Two of these anti-H strain monoclonal antibodies were used with the P-strain sporozoites in a neutralization assay. They selectively abolished the infectivity of part of the sporozoite population, and the remainder was designated the P2 strain. Monoclonal antibodies produced against sporozoites of the P2 strain identified two different sets of circumsporozoite proteins by Western blot analysis of an extract of these parasites, indicating the presence of two different populations of sporozoites. These two populations were separated by neutralization assays with monoclonal antibodies that recognized the two different sets of circumsporozoite proteins. The corresponding two strains were designated P3A and P3B. By Western blot analysis, two proteins with apparent molecular weights of 41,000 and 51,000 were identified in extracts of sporozoites of the P3A strain, and two proteins with molecular weights of 38,000 and 47,000 were identified in extracts of the P3B sporozoites.
Evaluation of number of shell vial cell cultures per clinical specimen for rapid diagnosis of cytomegalovirus infection
Authors: Paya C V , Wold A D , Ilstrup D M , Smith T F
Published: January 2020

Specimens submitted for the diagnosis of cytomegalovirus (CMV) infection were inoculated into three (blood) or two (urine, tissue, bronchoalveolar lavage [BAL]) shell vials seeded with MRC-5 cells for the diagnosis of CMV infection. We evaluated the detection of 993 specimens that were positive for CMV according to the number of shell vial cell cultures inoculated per specimen. For blood cultures, and considering one CMV-positive shell vial as 100%, inoculation of three shell vials versus one increased the detection rate of the virus by 51%. Inoculation of three shell vials compared with two yielded a 20% increase in the detection rate of CMV. For urine, tissue, and BAL specimens, inoculation of two shell vials compared with one resulted in increases of 7, 10, and 5%, respectively. For maximum detection of CMV in shell vial cell cultures, at least three vials should be inoculated with blood specimens, and two vials should be used for urine, tissue, and BAL samples.
Evaluation of antiviral drugs and neutralizing antibodies to human immunodeficiency virus by a rapid and sensitive microtiter infection assay
Authors: Montefiori D C , Robinson W E , Schuffman S S , Mitchell W M
Published: January 2020

A 96-well microtiter infection assay for the human immunodeficiency virus (HIV) is described. The assay utilizes human T-cell lymphotropic virus type I-immortalized MT-2 cells as targets for infection and requires only 4 to 5 days for completion. Cytolysis was quantitated by vital dye uptake of poly-L-lysine-adhered cells as an endpoint for infection. The assay's efficacy was proven by the sensitive and accurate assessment of several known anti-HIV agents including two inhibitors of reverse transcription (3'-azido-3'-deoxythymidine and 2',3'-dideoxycytidine), three biological response modifiers (recombinant interferons alpha and beta and mismatched double-stranded RNA), a direct inactivator of HIV virions (amphotericin B), and neutralizing antibodies from two HIV-positive human subjects. Evaluation of data was facilitated by computer-assisted analysis. This assay provides a means for rapid, sensitive, and inexpensive large-scale in vitro testing of potential anti-HIV therapeutic regimens and quantitation of HIV-neutralizing antibody titers.
Novel densitometric method for endonuclease analysis of Escherichia coli DNA samples containing multiple plasmids
Authors: Takahashi S , Nagano Y
Published: January 2020

We developed a novel method whereby the digestion pattern of each plasmid was distinguished in a sample of endonuclease-digested plasmid DNAs which contained multiple plasmids extracted from a bacterial strain. This method consisted of two procedures. (i) The concentration ratio of each undigested plasmid DNA and of each digested DNA fragment was calculated on the basis of densitometric scanning of an electrophoretogram, and the concentration ratio was then compared with the theoretical concentration ratio to determine from which plasmid each fragment was generated. (ii) The second procedure involved rapid visual identification with a scanning graph. We thus analyzed five strains of Escherichia coli that harbored several cryptic plasmids. The two procedures made it possible to analyze the restriction digestion pattern of each plasmid without the need to isolate the individual plasmids, except in the case of certain fragments. Even when the digested patterns of these exceptional fragments could not be distinguished completely, however, our method had the advantage that peak patterns of plasmids could be compared visually among different bacterial strains.
Effects of medium and inoculum variations on screening for high-level aminoglycoside resistance in Enterococcus faecalis
Authors: Sahm D F , Torres C
Published: January 2020

Enterococcus faecalis isolates that are refractory to aminoglycoside-penicillin synergy can be detected by their ability to grow in the presence of high concentrations of aminoglycoside (2,000 micrograms/ml). In past studies investigators have used a variety of media and inoculum sizes to perform high-level aminoglycoside resistance screens, but little is known about how these variations affect test accuracy. We screened 63 E. faecalis strains on different media by using various inoculum sizes and correlated the results with synergy test results obtained by time-kill studies. Screens were done with dextrose-phosphate agar, brain heart infusion agar, Trypticase soy agar with 5% sheep blood, Mueller-Hinton agar with 5% sheep blood, dextrose-phosphate broth, and Mueller-Hinton broth. Agar screens were inoculated with 10(2), 10(4), and 10(6) CFU; and broth screens contained a final inoculum of 10(5) CFU/ml. The E. faecalis isolates were tested for high-level resistance to streptomycin, kanamycin, amikacin, gentamicin, and tobramycin. Of the 63 isolates tested, 21 did not show high-level resistance to any of the aminoglycosides tested, and 42 demonstrated high-level resistance to one or more drugs. The sensitivity of most screens was greater than or equal to 90%. Regardless of the inoculum size or medium used, false-resistance results were seldom encountered. Screen specificity, which was used as the indicator of false susceptibility, was markedly influenced by both the inoculum size and the drug being tested. Specificity was low whenever a 10(2)-CFU inoculum was used, when amikacin was tested with any inoculum, and when tobramycin was tested in broth media. Data for kanamycin could be used to predict amikacin-penicillin synergy, and the highly accurate gentamicin screen obviated the need for the testing of tobramycin. We recommend a 10(6) -CFU inoculum for agar screens and a 10(5) -CFU/ml inoculum for broth screens. The type of medium used did not substantially influence screen accuracy. Among the aminoglycosides, only streptomycin, gentamicin, and occasionally, kanamycin need to be used to screen E. faecalis isolates for aminoglycoside-penicillin synergy.
High-content aminoglycoside disks for determining aminoglycoside-penicillin synergy against Enterococcus faecalis
Authors: Sahm D F , Torres C
Published: January 2020

We investigated the use of high-content aminoglycoside disks for determining Enterococcus faecalis susceptibility to aminoglycoside-penicillin synergy. The susceptibility of the organisms to synergy was established by 24-h time-kill studies performed with streptomycin, kanamycin, amikacin, gentamicin, and tobramycin, alone and in combination with penicillin. A total of 20 isolates that were susceptible to all drug combinations and 20 strains that were resistant to each aminoglycoside-penicillin combination were selected for testing against high-content disks. Disk-agar diffusion was performed on Mueller-Hinton agar, with and without 5% sheep blood, by using disks that contained either 300 or 2,000 micrograms of streptomycin and either 120 or 2,000 micrograms of kanamycin, amikacin, tobramycin, or gentamicin. Zone size results obtained for each aminoglycoside, except amikacin, could be used to differentiate between synergy-susceptible and -resistant isolates. No overlap occurred between the zone sizes of susceptible and resistant strains. Susceptibility to amikacin-penicillin synergy could reliably be tested with kanamycin, but not amikacin, disks. When the disks containing 120 micrograms were tested, a narrow zone size range of 6 to 7 mm could be used to identify all resistant strains. In contrast, when the disks containing 2,000 micrograms were used, the zone size ranges for resistant isolates varied widely with the aminoglycoside being tested. The presence of blood in the medium did not appreciably affect the disk test results. To detect resistance to every aminoglycoside-penicillin combination that may be considered for therapy, E. faecalis isolates need to be tested against a maximum of three different high-content disks (i.e., streptomycin, gentamicin, kanamycin). The disk-agar diffusion test performed with high-content aminoglycoside disks can provide laboratories with a convenient and reliable method for detecting E. faecalis isolates that are resistant to aminoglycoside-penicillin synergy.
Development and testing of invasion-associated DNA probes for detection of Shigella spp. and enteroinvasive Escherichia coli
Authors: Venkatesan M , Buysse J M , Vandendries E , Kopecko D J
Published: January 2020

Genetic determinants of the invasive phenotype of Shigella spp. and enteroinvasive Escherichia coli (EIEC), two common agents of bacillary dysentery, are encoded on large (180- to 210 kilobase), nonconjugative plasmids. Several plasmid-encoded antigens have been implicated as important bacterial ligands that mediate the attachment and invasion of colonic epithelial cells by the bacteria. Selected invasion plasmid antigen (ipa) genes have recently been cloned from Shigella flexneri serotype 5 into the lambda gt11 expression vector. Portions of three ipa genes (ipaB, ipaC, and ipaD) were tested as DNA probes for diagnostic detection of bacillary dysentery. Under stringent DNA hybridization conditions, all three DNA sequences hybridized to a single 4.6-kilobase HindIII fragment of the invasion plasmids of representative virulent Shigella spp. and EIEC strains. No hybridization was detected in isogenic, noninvasive Shigella mutants which had lost the invasion plasmid or had deleted the ipa gene region. Furthermore, these probes did not react with over 300 other enteric and nonenteric gram-negative bacteria tested, including Salmonella, Yersinia, Edwardsiella, Campylobacter, Vibrio, Klebsiella, Aeromonas, Enterobacter, Rickettsia, and Citrobacter spp. and various pathogenic E. coli strains. The use of unique invasion-essential gene segments as probes for the specific detection of invasive dysentery organisms should benefit both epidemiologic and diagnostic analyses of Shigella spp. and EIEC.
Morphologic appearance of inclusion bodies and their association with the antigenic composition of naturally occurring rabies viruses
Authors: Loretu K , Blenden D C , Torres-Anjel M J , Satalowich F T
Published: January 2020

A total of 112 rabies virus-infected skunk brain samples from naturally occurring cases (64 from Missouri, 48 from Kentucky) were code labeled and grouped into two morphologic categories according to the appearance and size of the discrete particles observed by immunofluorescent-antibody staining. The reactivity of the blind-labeled samples was then determined using a panel of 23 antinucleocapsid monoclonal antibodies to test whether morphologic appearance was associated with antigenicity. Two categories were defined and designated; they were morphologic type I (MO-I) and morphologic type II (MO-II). MO-I viruses produced granular-type fluorescing aggregates with no evidence of inclusions compatible in appearance with Negri bodies. MO-II viruses produced at least one and usually many large, round, or oval fluorescing inclusion bodies. No viruses categorized as MO-I reacted with antinucleocapsid monoclonal antibodies 103-7, 239-10, and 120-2; all MO-II viruses reacted positively to these three antibodies, with one exception involving antinucleocapsid monoclonal antibody 239-10. The results indicate a strong association between the reactivity of these antibodies and the morphologic appearance of the aggregates of antigen.
Monoclonal antibody enzyme-linked immunosorbent assay for specific identification and typing of subgroup F adenoviruses
Authors: Singh-Naz N , Rodriguez W J , Kidd A H , Brandt C D
Published: January 2020

Monoclonal antibody specific for subgroup F enteric adenoviruses (EAds) was prepared by fusing P3-NS1/Ag4-1 mouse myeloma cells with lymphocytes from BALB/c mice immunized with G1105, an adenovirus type 41 (Ad41) strain. Monoclone 3F11/2H9, which specifically recognized Ad41, was successfully used as detector antibody in an enzyme-linked immunosorbent assay (ELISA). Additionally, previously prepared monoclones 5D8/2C2 and 2H6/1E11, recognizing Ad40 plus Ad41 and Ad40 alone, respectively, were used to study stool and/or tissue culture specimens from 106 patients with adenovirus-positive gastroenteritis. By ELISA, 91 had EAds (22 were Ad40 and 69 were Ad41) and 15 had non-EAds. ELISA results were in concordance with restriction endonuclease results for 38 of 39 specimens, with dot blot data for 19 of 20 specimens, and with neutralization test results for 74 of 78 specimens. ELISA was at least 10-fold more sensitive than direct electron microscopy was for the detection of EAds in stool specimens.
Evaluation of proteolytic activity to differentiate some dematiaceous fungi
Authors: Espinel-Ingroff A , Goldson P R , McGinnis M R , Kerkering T M
Published: January 2020

A total of 123 isolates of Cladosporium spp., Exophiala spp., Fonsecaea spp., Lecythophora hoffmannii, Phaeoannellomyces werneckii, Phialophora spp., Wangiella dermatitidis, and Xylohypha bantiana were tested for proteolytic activity by using 26 different formulations of gelatin, milk, casein, and Loeffler media. Other physiological properties examined included hydrolysis of tyrosine and xanthine, sodium nitrate utilization in Czapek Dox agar, and thermotolerance. Isolates of Exophiala jeanselmei, Fonsecaea compacta, Fonsecaea pedrosoi, W. dermatitidis, and X. bantiana lacked proteolytic activity. Proteolytic activity was variable among the remaining species, depending on the type of medium used. Thermotolerance had value in distinguishing some taxa.
Sensitivity and specificity of viral immunoglobulin M determination by indirect enzyme-linked immunosorbent assay
Authors: Champsaur H , Fattal-German M , Arranhado R
Published: January 2020

Four sources of error associated with virus-specific immunoglobulin M (IgM) determination by indirect enzyme-linked immunosorbent assay were recognized and analyzed. First, competitive inhibition due to specific IgG was demonstrated by experiments involving addition and subtraction of rubella-specific IgG. Second, the interference due to rheumatoid factors (RFs) of the IgM class (IgM-RFs) was studied thoroughly, and it appeared that the level of false positivity was more dependent on specific IgG titers than on IgM-RF titers. Third, it was found that some IgM-RFs, differing from conventional IgM-RFs in that they reacted only with isologous IgG, were responsible for further cases of false positivity. Fourth, the interference of an IgM reacting with some virus-unmasked cellular antigens was demonstrated for some uninfected individuals. All four interfering factors could be readily eliminated by simply premixing serum samples with a sheep anti-human gamma-chain serum. This single pretreatment was shown to eliminate false-negatives as well as false-positives in a further 2,004 sera tested for six viruses. These results also emphasize the frequency of RFs and their heterogeneity.
Comparison of antibodies for rapid detection of cytomegalovirus
Authors: Randazzo D N , Michalski F J
Published: January 2020

Antibodies were compared for use in the spin-amplified shell vial method for rapid cytomegalovirus detection. Commercial antibodies by Du Pont Co., Whittaker M.A. Bioproducts, Bartels Immunodiagnostic, Virostat, and Serono Diagnostics were compared at incubation times of 16 to 48 h on 22 patient specimens. Only the Du Pont antibody showed 100% sensitivity and specificity and no nonspecific reactions.
DNA probes for K-antigen (capsule) typing of Escherichia coli
Authors: Roberts M , Roberts I , Korhonen T K , Jann K , Bitter-Suermann D , Boulnois G J , Williams P H
Published: January 2020

DNA restriction fragments derived from the polysaccharide biosynthesis regions of cloned Escherichia coli K1, K5, and K12 capsular antigen genes hybridized only with DNA of strains determined by conventional methods to be of the same K serotype. A probe derived from the common transport region hybridized to all encapsulated E. coli strains.
Serotyping of Clostridium difficile
Authors: Toma S , Lesiak G , Magus M , Lo H L , Delmée M
Published: January 2020

A total of 246 live Clostridium difficile cultures were serotyped by a slide agglutination technique. Fifteen grouping antisera were produced which serotyped 98% of the cultures (241 of 246). Our results indicated that certain serogroups may have specific pathogenicity. Strains of serogroups A, G, H, K, S1, and S4 were cytotoxigenic and were isolated mainly from adult patients with pseudomembranous colitis or antibiotic-associated diarrhea. Nontoxigenic strains of serogroups D and Cd-5 were isolated mainly from asymptomatic neonates and small children. Some cross-reactions occurred among some strains of serogroups A, Cd-5, G, and K. These strains were further examined by analysis of protein profiles and restriction endonuclease patterns to elucidate their serology. Typing of C. difficile by using slide agglutination is a simple technique suitable for routine examination. Serogrouping may be a useful epidemiological marker and could help in elucidating the medical relevance of some C. difficile isolates.
Agglutinins and antibodies to Francisella tularensis outer membrane antigens in the early diagnosis of disease during an outbreak of tularemia
Authors: Bevanger L , Maeland J A , Naess A I
Published: January 2020

Tularemia was diagnosed in 57 patients during an outbreak in central Norway in 1984 and 1985. Clinical categories of the disease showed seasonal variations. A bacterial microagglutination test and an enzyme-linked immunosorbent assay (ELISA) with class-specific antibodies against Francisella tularensis outer membrane (OM) antigens were evaluated for the early diagnosis of tularemia. ELISA with immunoglobulin G (IgG), IgA, or IgM antibodies and the microagglutination test differed only marginally in diagnostic sensitivity. The OM preparation harbored F. tularensis agglutinogens and contained a variety of proteins, several of which functioned as immunogens in tularemia patients, as shown by Western blotting (immunoblotting). All 12 patients tested produced antibodies against a 43,000-molecular-weight OM protein. Individual variation was noted with regard to antibody response against other OM antigens. The OM is a suitable antigen preparation in ELISA for the diagnosis of tularemia and, presumably, contains antigens important in the immunobiology of tularemia.
Clinical laboratory challenges in the recognition of Leuconostoc spp
Authors: Isenberg H D , Vellozzi E M , Shapiro J , Rubin L G
Published: January 2020

Vancomycin-resistant Streptococcus salivarius-like bacteria were isolated from the blood and gastrostomy tube of a 6-month-old child during vancomycin therapy for Staphylococcus epidermidis central venous catheter-associated bacteremia. The isolates produced D-(-)-lactate and ethanol from glucose, hydrolyzed esculin, and produced no ammonia from arginine, thus fulfilling the major criteria as Leuconostoc spp. The enzymatic profile of the bacteria suggested that they were not lactobacilli or streptococci. Resistance to vancomycin and penicillin tolerance were noted. The formula fed through the gastrostomy tube was suspected as the vehicle that transmitted the organism to the patient. Penicillin and tobramycin therapy was used successfully.
Characterization of monoclonal antibodies to protein antigen of Salmonella typhi
Authors: Sarasombath S , Lertmemongkolchai G , Banchuin N
Published: January 2020

Two monoclonal antibodies were produced against protein antigens of Salmonella typhi. One of the antibodies (STP14) belongs to the immunoglobulin G1K subclass, and the other (STP13) was assigned to the immunoglobulin G2a(kappa) subclass. Both antibodies could recognize the 34.0-kilodalton protein antigen from S. typhi. The specificity of these antibodies was tested by immunoblotting with a panel of crude protein antigens from 12 bacteria causing enteric fever and enteric fever-like illness: S. typhi, S. paratyphi A, S. paratyphi B, S. paratyphi C, S. choleraesuis, S. enteritidis, S. krefeld, S. panama, S. typhimurium, Escherichia coli, Pseudomonas pseudomallei, and Yersinia enterocolitica. In a modified double-antibody sandwich enzyme-linked immunosorbent assay they could detect the protein antigen at ca. 0.6 microgram/ml. These monoclonal antibodies should be of great value in the diagnostic test for detecting S. typhi antigen in samples of bodily fluids isolated from patients with typhoid fever and in studies of the chemical structure and other immunological properties of this 34.0-kilodalton protein.
Characterization of freshly isolated Campylobacter coli strains and suitability of selective media for their growth
Authors: Ng L K , Taylor D E , Stiles M E
Published: January 2020

Typical and atypical Campylobacter strains were isolated from the colons of cattle and swine by techniques that enabled the selective pressures of antibiotics to be avoided. Some cephalothin-susceptible strains and a strain with an indeterminate hippurate reaction were classified as Campylobacter coli by DNA homology testing. Tetracycline-resistant isolates were obtained from animals with no recorded exposure to antibiotics. A selection of 12 C. coli and 6 C. jejuni strains was used to determine the ability of fresh isolates to grow on a range of selective media. C. coli isolates were inhibited more than C. jejuni on selective media containing antibiotics. The least inhibitory media were Skirrow medium (M. B. Skirrow, Br. Med. J. 2:9-11, 1977) and the charcoal-based media developed by Hutchinson and Bolton (D. N. Hutchinson and F. J. Bolton, J. Clin. Pathol. 37:956-957, 1984) and Karmali et al. (M. A. Karmali, A. E. Simon, M. Roscoe, P. C. Fleming, S. S. Smith, and J. Lane, J. Clin. Microbiol. 23:456-459, 1986). The plasmid contents of the isolates did not appear to be related to their sensitivity to growth on selective (antibiotic-containing) media. The study indicates that selective media used to detect Campylobacter spp. could select against the isolation of C. coli.
Production of gamma-hemolysin and lack of production of alpha-hemolysin by Staphylococcus aureus strains associated with toxic shock syndrome
Authors: Clyne M , De Azavedo J , Carlson E , Arbuthnott J
Published: January 2020

The hemolytic activity of toxic shock syndrome isolates of Staphylococcus aureus is enhanced when agarose is substituted for agar in blood plates or when strains are grown in liquid culture in the presence of 20% (vol/vol) CO2 in air. Hemolytic activity of a representative panel of toxic shock syndrome isolates was rigorously assessed both on blood agar and in liquid culture to unequivocally identify the predominant hemolysins produced. As determined by isoelectric focusing and Western immunoblotting, 15 of 15 TSS isolates produced gamma-lysin and 10 of 15 produced delta-lysin. None produced beta-lysin, and only 2 of 15 produced alpha-lysin. The low rate of alpha-lysin production was a most striking characteristic, since all strains were found to have the alpha-lysin gene by Southern blot hybridization.
Comparison of four hemolysin-producing organisms (Escherichia coli, Serratia marcescens, Aeromonas hydrophila, and Listeria monocytogenes) for release of inflammatory mediators from various cells
Authors: Scheffer J , König W , Braun V , Goebel W
Published: January 2020

We investigated the role of various hemolysin-producing strains (Escherichia coli, Serratia marcescens, Aeromonas hydrophila, and Listeria monocytogenes) in induction of inflammatory mediators, e.g., histamine release from rat mast cells as well as the chemiluminescence response and the release of lipoxygenase transformation products from human polymorphonuclear neutrophils. Our data show that the hemolysin-positive bacteria as well as the hemolysin-positive culture supernatants were active in inducing the chemiluminescence response, leukotriene (LTB4 and LTC4) release from human granulocytes, and histamine release from rat mast cells. The degree of leukotriene release was dependent on the hemolysin type and on the expression of hemolysin activity. The E. coli alpha-hemolysin and the aerolysin-producing A. hydrophila were the most potent stimuli whether washed bacteria or bacterial supernatant was used. Bacteria expressing the S. marcescens hemolysin and the listeriolysin were only poor inducers of leukotriene generation. In contrast to leukotriene generation, all hemolysin-positive strains induced nearly the same histamine release in a dose-dependent manner. Our data suggest a potent role for various hemolysins as virulence factors in inducing the release of inflammatory mediators.
Immunologic and clinicopathologic evaluation of adult dogs inoculated with Encephalitozoon cuniculi
Authors: Szabo J R , Shadduck J A
Published: January 2020

Beagle dogs inoculated intravenously with 1.75 x 10(9) viable Encephalitozoon cuniculi spores at 12.5 months of age were monitored for 18 months to assess infection in the adult dog. Parameters monitored included packed cell volume, total and differential leukocyte counts, and humoral and cellular responses to infection. Immunoglobulin M and G antibodies directed against the parasite tegument were present throughout the 18 months. Peripheral blood monocytes treated with lymphokines, either antigen specific (E. cuniculi) or nonspecific (concanavalin A), killed E. cuniculi spores in vitro. Pretreatment of E. cuniculi spores with normal dog serum or infected dog serum enhanced the killing of the parasite (normal dog serum less than infected dog serum) by canine monocytes. Histologic examination of selected tissues revealed microfocal plasma cell and lymphocyte aggregates at the renal corticomedullary junction and in the medullary interstitium. The results of these experiments suggest that the adult dog is able to mount an effective defense to infection, to minimize host tissue damage, and to eliminate the parasite through complex interactions between monocytes-macrophages and lymphocytes.
Radiolabeled total parasite DNA probe specifically detects Trypanosoma cruzi in mammalian blood
Authors: Ashall F , Yip-Chuck D A , Luquetti A A , Miles M A
Published: January 2020

A DNA-DNA hybridization procedure is described in which radiolabeled total parasite DNA is used to detect Trypanosoma cruzi spotted directly onto nylon membrane. Under conditions that favor hybridization of repetitive DNA sequences, the radiolabeled total DNA was able to detect as few as five parasites spotted onto nylon membrane. Trypanosomes were detectable in blood from mice with T. cruzi parasitemias spotted directly onto membranes. All 10 strains of T. cruzi examined, which came from different areas of the United States and South America, were readily detected with a total DNA probe from any 1 strain of the parasite. No signals were detected with up to 20,000 Trypanosoma rangeli, T. brucei, Crithidia fasciculata, or Leishmania parasites or with normal mouse blood or mammalian DNA. The hybridization method is sensitive, specific, rapid, and inexpensive and is potentially applicable to the detection of other parasites.
Thirteenth serogroup of Legionella pneumophila isolated from patients with pneumonia
Authors: Lindquist D S , Nygaard G , Thacker W L , Benson R F , Brenner D J , Wilkinson H W
Published: January 2020

A Legionella-like organism (strain 82A3105; ATCC 43736) was isolated from a lung aspirate taken from a patient with pneumonia. Results of physiologic, gas-liquid chromatographic, genetic, and serologic tests showed that strain 82A3105 and four additional clinical isolates belong to a new Legionella pneumophila serogroup, serogroup 13.
Use of cryopreserved normal peripheral blood lymphocytes for isolation of human immunodeficiency virus from seropositive men
Authors: Balachandran R , Thampatty P , Rinaldo C , Gupta P
Published: January 2020

The possibility was investigated of using frozen stocks of phytohemagglutinin (PHA)-stimulated normal human peripheral blood lymphocytes (PBL) in cocultivation with human immunodeficiency virus (HIV)-infected lymphocytes for the isolation of HIV. Fresh and cryopreserved PBL from eight healthy volunteers were compared for their susceptibility to HIV infection in vitro. Fresh lymphocytes, as well as lymphocytes that were stimulated with PHA before or after cryopreservation, displayed comparable susceptibilities to HIV infection in vitro. In addition, HIV was recovered in all cases when lymphocytes stimulated with PHA before or after cryopreservation were cocultured in parallel with PBL from 15 patients with acquired immune deficiency syndrome. However, the cryopreserved PBL were less efficient in isolating HIV from asymptomatic men.