bluuebunny/crossref_metadata_2025 huggingface dataset:
Induction of histamine release from rat mast cells and human basophilic granulocytes by clinical Escherichia coli isolates and relation to hemolysin production and adhesin expression
Authors: Gross-Weege W , König W , Scheffer J , Nimmich W
Published: January 2020

We investigated the role of 27 disease-relevant Escherichia coli strains isolated from humans in the induction of histamine release from rat peritoneal mast cells and human basophilic granulocytes. Our data indicated that only the hemolysin-positive (HLY+) bacteria and the hemolysin-positive culture supernatants induced histamine release. For the latter, the hemolysin activity determined the degree of histamine secretion. Incubation of the target cells with washed HLY+ bacteria revealed a different secretory response. For the rat mast cells, histamine release paralleled expression of hemolysin activity, with the exception of strain S98 (O75:K5:H- HLY+), which induced less histamine, although its hemolysin activity was relatively high. No correlation between histamine secretion and hemolysin activity was observed when human basophils were stimulated with the HLY+ bacteria. Large amounts of histamine were still released, even when the hemolysin activity declined to zero. Our results support the potent role of the E. coli hemolysin as a pathogenicity factor in bacterial infection.
Simple method of inducing sporulation by Apophysomyces elegans and Saksenaea vasiformis
Authors: Padhye A A , Ajello L
Published: January 2020

Apophysomyces elegans and Saksenaea vasiformis are notorious for their failure to sporulate on routine media. Agar blocks, permeated with the mycelia of A. elegans and S. vasiformis, were cut aseptically from 7-day-old colonies grown on Sabouraud dextrose agar and transferred to plates containing 20 ml of sterile distilled water supplemented with 0.2 ml of 10% filter-sterilized yeast extract solution. When the plates were incubated at 37 degrees C, all 5 isolates of A. elegans and all 10 isolates of S. vasiformis produced abundant, characteristic sporangia within 7 to 10 days. The method is simple to use and yields consistent results.
Improved method for azole antifungal susceptibility testing
Authors: Gordon M A , Lapa E W , Passero P G
Published: January 2020

A reproducible method is described for the determination of the MICs of ketoconazole, miconazole, fluconazole, and itraconazole with sharp endpoints when employed with either yeasts or molds. A semisolid medium is used with controlled pH and standardized inoculum. The time of reading results is a critical factor in the conduct of this test. The medium is simple to prepare and has a relatively long refrigerator shelf life in a user-ready state, requiring only the addition of a freshly prepared inoculum after restoration to room temperature.
Disk diffusion interpretive criteria for extended-spectrum cephalosporins with Haemophilus influenzae
Authors: Jorgensen J H , Maher L A , Redding J S
Published: January 2020

Disk diffusion interpretive zone size criteria for use with Haemophilus test medium are proposed for five extended-spectrum cephalosporins. With cefuroxime, zones greater than or equal to 24 mm in diameter indicated susceptibility, while zones less than or equal to 20 mm implied resistance. With cefotaxime, ceftizoxime, and ceftriaxone, a single susceptible-zone size, greater than or equal to 26 mm, is proposed.
Comparison of antigen immunoassay and reverse transcriptase assay for monitoring human immunodeficiency virus infection in an antiviral trial
Authors: Burger H , Paul D , Siegal F P , Wendel I , Neff S , Eilbott D , Gehan K , Grimson R , Weiser B
Published: January 2020

We compared the Abbott enzyme immunoassay for human immunodeficiency virus (HIV) antigen with the reverse transcriptase assay (RTA) as a means of monitoring HIV infection during an antiviral trial. The Abbott enzyme immunoassay detected HIV earlier than RTA whether or not the patients were antigenemic and appears to be superior to RTA for detecting HIV in cultures used for monitoring clinical trials.
Odontogenic infection secondary to Leuconostoc species
Authors: Wenocur H S , Smith M A , Vellozzi E M , Shapiro J , Isenberg H D
Published: January 2020

Leuconostoc species are gaining importance as pathogenic organisms. We present the first case of odontogenic infection caused by Leuconostoc spp. Isolates initially identified as streptococci were found to be vancomycin resistant. Rigorous bacteriologic investigation subsequently classified these organisms as Leuconostoc mesenteroides.
Evaluation of 9-chloro-9-[4-(diethylamino)phenyl]- 9,10-dihydro-10-phenylacridine hydrochloride (C-390) in broth and agar media for identification of Pseudomonas aeruginosa
Authors: Fader R C , Latimer J , Bannister E , Lucia H
Published: January 2020

Brain heart infusion broth and Mueller-Hinton agar containing the compound 9-chloro-9-[4-(diethylamino)phenyl]- 9,10-dihydro-10-phenylacridine hydrochloride (C-390) were evaluated as selective media for identifying Pseudomonas aeruginosa. Of the 192 Pseudomonas spp. and 68 additional oxidase-positive or glucose-nonfermenting gram-negative organisms tested, only P. aeruginosa (120 of 121 isolates) and certain strains of Pseudomonas cepacia, Aeromonas hydrophila, and Achromobacter xylosoxidans were able to grow in brain heart infusion broth containing 15 micrograms of C-390 per ml. When production of green or yellow-green pigment after overnight growth on Mueller-Hinton agar containing 30 micrograms of C-390 per ml was used as the criterion for identification, only P. aeruginosa produced a positive test (96% sensitivity and 100% specificity). The agar medium, therefore, can provide a single differential medium for the overnight identification of P. aeruginosa.
Detection of bacteria in the presence of antibiotics by using specific monoclonal antibodies to neutralize the antibiotics
Authors: Sierra-Madero J G , Caulfield M J , Hall G S , Washington J A
Published: January 2020

Inactivation of penicillin and gentamicin in cultures was achieved by using monoclonal antibodies against these antibiotics. A viridans group streptococcus (penicillin MIC, less than or equal to 0.06 micrograms/ml) and Escherichia coli ATCC 35218 (gentamicin MIC, less than or equal to 1 microgram/ml) were able to grow in broth containing 0.25 micrograms of penicillin per ml and 4 micrograms of gentamicin per ml, respectively, when the specific antibodies were added. This procedure may be useful to increase the yield of bacteria from body fluid specimens that contain antibiotics.
Solid-phase immunosorbent technique for rapid detection of Rift Valley fever virus immunoglobulin M by hemagglutination inhibition
Authors: Soliman A K , Botros B A , Morrill J C
Published: January 2020

A solid-phase immunosorbent technique (SPIT) was adapted to detect Rift Valley fever (RVF) virus-specific immunoglobulin M (IgM) in serum samples from humans vaccinated with Formalin-inactivated RVF vaccine. Microdilution plates coated with goat anti-human IgM were successively incubated with serum samples from human vaccinees, RVF virus hemagglutinating antigen, and goose erythrocytes. The RVF virus-specific IgM in the serum samples from vaccinees bound to the RVF virus antigen and inhibited hemagglutination of goose erythrocytes. SPIT was compared to the IgM capture enzyme linked immunosorbent assay (ELISA) and the indirect immunofluorescent-antibody (IFA) assay and was found to be sensitive in detecting RVF virus-specific IgM antibody, with high correlations between SPIT and the other two tests (Pearson's correlation coefficient [r] = 0.9 and 0.6, respectively). Results of SPIT were obtained within 5 h, offering speed over ELISA (8 h). In addition, SPIT does not require sophisticated equipment or expensive reagents. Serum rheumatoid factor did not produce false-positive reactions in SPIT as in the indirect immunofluorescent-antibody assay and IgM capture ELISA.
Evaluation of the Cobas-Bact system for direct and rapid identification and antimicrobial susceptibility testing of gram-negative rods from positive blood culture broths
Authors: Kamm W , Wenger A , Bille J
Published: January 2020

A direct antimicrobial susceptibility test and a direct identification of positive blood culture broths for gram-negative rods confirmed with Gram stain by using a new instrument, Cobas-Bact, were compared with the conventional Kirby-Bauer agar diffusion disk method and with the in-house set of identification or API 20E, respectively. The bacterial pellet of centrifuged positive blood culture broth was used to inoculate a Cobas-Bact susceptibility and identification rotor. Bacteria from 206 cases of monomicrobial septicemia due to members of the family Enterobacteriaceae were tested. In 198 episodes (96%), direct identification and antimicrobial susceptibility testing results were obtained for the same bacterial pathogen within 5 h of detection. Of 204 direct identifications obtained, 177 (86.6%) were "high-confidence" correct identifications (percentage of likelihood [P] greater than or equal to 80%) and 25 (12.5%) "low-confidence" correct identifications (P less than 80%), whereas only 2 misidentifications occurred (1 Escherichia coli and 1 Proteus mirabilis). Direct susceptibility testing was performed in 199 episodes (96%), providing 1,885 antibiotic-microorganism combinations. Full agreement reached 86.3%, and essential agreement reached 92.8%. Minor discrepancies were found in 120 (6.5%) of the tests, major discrepancies were found in 127 (6.8%) tests, and very major discrepancies were found in only 7 (0.4%) tests. Subsequent MIC determinations in cases of major or very major discrepancies reduced the number of major discrepancies involving cephalosporins from 60 to 16, whereas all those involving aminoglycosides remained. Overall, this direct and rapid Cobas-Bact identification and susceptibility testing procedure offered accurate information with 5 to 6 h after the laboratory detection of bacteremia and septicemia due to members of the Enterobacteriacease.
Monoclonal antibody solution hybridization assay for detection of human immunodeficiency virus nucleic acids
Authors: Viscidi R P , O'Meara C , Farzadegan H , Yolken R
Published: January 2020

In this report we describe a novel, nonisotopic hybridization assay for the measurement of viral RNA in biological samples. The assay involved a solution-phase reaction between a biotinylated DNA probe and RNA target sequences. Labeled hybrids were detected in an immunoreaction by using a solid-phase anti-biotin antibody and an enzyme-labeled monoclonal antibody specific for DNA-RNA hybrids. This monoclonal antibody solution hybridization assay was compared with an antigen-capture immunoassay for the detection of human immunodeficiency virus in 436 cell culture samples from 60 seropositive patients. The sensitivity and specificity of the hybridization assay were 93.5 and 94.6%, respectively. Detection of human immunodeficiency virus solely by hybridization in the initial sample but not subsequent samples from seven cultures may reflect detection of virus that was present in the patients' lymphocytes but did not replicate in vitro. Since the assay method is adapatable to the detection of either RNA or DNA, it could provide a means for the detection of a wide range of viral nucleic acids.
Molecular epidemiology and subgroup determination of bovine group A rotaviruses associated with diarrhea in dairy and beef calves
Authors: Theil K W , McCloskey C M
Published: January 2020

The genome electropherotyping technique was used to examine group A rotaviral infections of diarrheic calves ranging from 1 to 85 days of age in 2 beef and 27 dairy herds. Coelectrophoresis studies demonstrated 38 distinct bovine group A rotavirus genome electropherotypes; all were long genome electropherotypes, and none had extra segments or unusual segment rearrangements. Genome electropherotypes in fecal specimens from diarrheic calves previously inoculated orally with a commercial, modified-live group A rotavirus vaccine differed from the vaccine genome electropherotype. Generally, when fecal specimens for genome electropherotyping were collected from two or more different calves within the same herd over a relatively short time, only one genome electropherotype was detected within a given herd. Different genome electropherotypes were detected in the same herd, however, when fecal specimens were obtained from different diarrheic calves over longer intervals (6 months or more). Twenty-three group A rotavirus strains with distinct genome electropherotypes, from diarrheic calves in 22 herds, were isolated and plaque purified in cell culture, and all were subgroup 1 group A rotaviruses. Non-group A rotavirus genome electropherotypes were not detected in 131 fecal specimens, negative for group A rotavirus, collected from diarrheic calves in 17 dairy herds.
Detection by enzyme-linked immunosorbent assay of specific immunoglobulin G isotypes in primary and established cytomegalovirus infections
Authors: Joassin L , Somze A , Reginster M
Published: January 2020

An enzyme-linked immunosorbent assay using monoclonal antibodies was developed to study the subclass distribution of immunoglobulin G (IgG) to cytomegalovirus (CMV) in individuals from a number of clinical groups. Most CMV-seropositive individuals had IgG1 and IgG3. IgG2 and IgG4 were detected less frequently at very low levels of activity, mostly among mothers at delivery and renal patients. Most seroconversions were accompanied by an important increase of the IgG1 activity, whereas IgG3 appeared at lower levels; neither IgG2 nor IgG4 occurred. This suggests that these isotypes play a secondary role in the response to the CMV infection and that they may be considered markers of past infections. Anti-CMV IgG1 is the most efficiently transmitted through the placenta. Whether infected or not, newborns had the same subclass distribution and activity levels as their mothers. Isotype determination did not offer a decisive explanation of a number of discrepancies observed between CMV IgG enzyme-linked immunosorbent assay and complement fixation test results.
Immunoglobulin M, A, and G antibody response to lipopolysaccharide O antigen in symptomatic and asymptomatic Shigella infections
Authors: Cohen D , Block C , Green M S , Lowell G , Ofek I
Published: January 2020

The antilipopolysaccharide antibody response in sera obtained from subjects involved in 10 outbreaks of shigellosis occurring in Israeli military field units was determined by an enzyme-linked immunosorbent assay and a passive hemagglutination test. Both tests were found to be sensitive and specific for the diagnosis of shigellosis. A significant antibody response was detected in 73 to 82% of the symptomatic and 48 to 60% of the asymptomatic subjects during the Shigella sonnei and Shigella flexneri outbreaks. Fifty percent of the symptomatic and none of the asymptomatic subjects showed a significant antibody response in the Shigella boydii outbreaks. An examination of the kinetics of the antibody levels over a 10-week period after the onset of disease revealed that immunoglobulin A (IgA) levels were highest 2 weeks after infection and had declined to initial levels within 2.5 months. In contrast, IgG levels at the late convalescent stage were half those measured at early convalescence, still being about twice as high as the initial titers. Although the IgM levels showed a pattern similar to that of IgA, their elevation at the early convalescent stage was less pronounced. We conclude that the detection of an increase in the level of the IgA fraction appeared to be the best indicator for recent symptomatic, as well as symptomatic, infections due to Shigella organisms.
Bacteremia and urinary tract infection associated with CDC group Vd biovar 2
Authors: Van Horn K G , Gedris C A , Ahmed T , Wormser G P
Published: January 2020

A case of urinary tract infection and bacteremia caused by CDC group Vd biovar 2 in a 23-year-old woman with Hodgkin's disease is described. This is the first report of CDC group Vd biovar 2 isolated from a clinical specimen and considered as a pathogen. Detailed antimicrobial susceptibility data are presented.
Enzymatic extraction and spectral analysis of the chromophore from cell walls of nutritionally deficient streptococci
Authors: Stein D S , Libertin C R
Published: January 2020

We demonstrated by spectral analysis a method to enzymatically cleave the chromophore precursor from nutritionally deficient streptococcal cell walls by treatment with lysozyme. The peak absorbances without and with lysozyme pretreatment were 511.4 +/- 2.02 nm and 513.1 +/- 0.69 nm, respectively. Extraction yields varied among strains and were found to be growth phase dependent. A secondary peak of absorbance (mean of 477 nm) was found in only five of eight strains. The chromophore at a neutral pH undergoes a reaction with phenol consistent with that of a furan, indicating its carbohydrate composition.
Detection of cytomegalovirus from clinical specimens in centrifugation culture by in situ DNA hybridization and monoclonal antibody staining
Authors: Gleaves C A , Hursh D A , Rice D H , Meyers J D
Published: January 2020

An in situ DNA hybridization kit for cytomegalovirus (CMV) was evaluated for the detection of CMV in centrifugation culture. Of 61 clinical specimens, 17 (27.8%) were positive for CMV by monoclonal antibody staining following centrifugation. Of the 17 positive specimens, 15 were detected by DNA hybridization (24.5%). However, the earliest that CMV could be detected by DNA hybridization was 58 h as compared with 16 h with monoclonal antibodies following centrifugation. DNA hybridization remains of great interest for the study and detection of CMV infection. However, current DNA hybridization techniques are not sufficiently rapid to replace the use of monoclonal antibodies in centrifugation culture.
Flavimonas oryzihabitans (Pseudomonas oryzihabitans; CDC group Ve-2): an emerging pathogen in peritonitis related to continuous ambulatory peritoneal dialysis?
Authors: Bendig J W , Mayes P J , Eyers D E , Holmes B , Chin T T
Published: January 2020

A case of peritonitis caused by Flavimonas oryzihabitans (Pseudomonas oryzihabitans; CDC group VE-2) in a patient on continuous ambulatory peritoneal dialysis is reported. This is the seventh case of infection caused by this organism reported in the English literature and the third reported case of continuous ambulatory peritoneal dialysis-related peritonitis caused by this organism; it is the first case of infection of any kind caused by this organism in England.
Comparison of three methods for detection of Cryptosporidium oocysts in a low-prevalence population
Authors: Baron E J , Schenone C , Tanenbaum B
Published: January 2020

Four hundred fecal specimens which had been received for routine ova and parasite examination were concentrated by Formalin-ether sedimentation. Sediments were examined as saline and iodine-stained wet preparations and were stained with rhodamine-auramine O and a commercially available monoclonal fluorescent-antibody stain for oocysts of Cryptosporidium species. Examination with the fluorescent stains detected cryptosporidia in both positive specimens (0.5% prevalence), and routine direct wet-preparation examination detected cryptosporidia in one of them. Detection of only low numbers of positive specimens in our nonrisk population argues against routine use of specific and expensive stain reagents.
Evaluation of a method for rapid detection of penicillinase-producing Neisseria gonorrhoeae in urethral exudates
Authors: Herve V M , Georges A J , Massanga M , Martin P M
Published: January 2020

A substrate, pyridine-2-azo-p-dimethylaniline cephalosporin (PADAC; Diagnostic Pasteur, Marnes-La-Coquette, France), for detection of penicillinase-producing Neisseria gonorrhoeae (PPNG) on isolated colonies grown on agar was compared with the nitrocefin reference test (Cefinase; Biomerieux, Marcy l'Etoile, France). The PADAC test was also used to detect PPNG directly on urethral exudates. All 38 PPNG-containing and 29 of 30 PPNG-negative urethral pus isolates gave results in agreement with those obtained on cultured isolates. We conclude that direct detection of PPNG in urethral exudates from males is simple, rapid, reliable, and sensitive.
Monoclonal antibody-mediated, immunodiagnostic competitive enzyme-linked immunosorbent assay for equine monocytic ehrlichiosis
Authors: Shankarappa B , Dutta S K , Sanusi J , Mattingly B L
Published: January 2020

Competitive enzyme-linked immunosorbent assay (CELISA), mediated by a monoclonal antibody designated HybI, was developed for the diagnosis of equine monocytic ehrlichiosis. Inhibition of binding of HybI by the horse antibodies to Ehrlichia risticii was optimum at dilutions of 1:20 for serum and 1:10,000 for HybI. Mean optical densities (ODs) of positive and negative sera were 0.158 and 0.855, respectively. A comparison of ODs obtained by CELISA and indirect enzyme-linked immunosorbent assay (ELISA) indicated a marked tendency of positive and negative samples to cluster separately with respect to CELISA ODs, whereas the ELISA results displayed a continuum of ODs from negative to positive. Analysis of diagnosis by indirect fluorescent-antibody test (IFA), ELISA, and CELISA for 66 field-collected serum samples indicated that CELISA was superior to IFA and ELISA. Among 11 acute-phase serum samples negative by IFA which were obtained from horses that subsequently seroconverted, CELISA clearly demonstrated antibodies in 8 of these acute-phase sera, whereas 5 were borderline positive by ELISA. The presence of agent-specific humoral antibodies could be demonstrated conclusively by 14 days after infection. The results suggest that CELISA is more sensitive than IFA and ELISA and, owing to the marked differences between positive and negative samples, can be easily adapted for use in the field for detection of horse antibodies to E. risticii.
Detection of antigens in urine of mice and humans infected with Borrelia burgdorferi, etiologic agent of Lyme disease
Authors: Hyde F W , Johnson R C , White T J , Shelburne C E
Published: January 2020

Lyme disease is a seasonal tick-borne malady which has worldwide distribution. Early and accurate diagnosis of Lyme disease is essential for successful antibiotic therapy. Symptoms are too vague to make an early diagnosis based on conventional criteria. We report the detection of antigens of Borrelia burgdorferi, causative agent of Lyme disease, in the urine of infected mice and humans. This technique may eventually provide a rapid diagnostic test for the early and accurate detection of this illness.
Immunoglobulin G subclass antibodies to measles virus in patients with subacute sclerosing panencephalitis or multiple sclerosis
Authors: Mehta P D , Patrick B A , Sobczyk W , Kulczycki J , Woyciechowska-Camenga J , Camenga D , Thormar H
Published: January 2020

Mouse monoclonal antibodies specific for human immunoglobulin G (IgG) subclasses and a sensitive immunoassay were used to evaluate the IgG subclass antibody response to measles virus antigens in cerebrospinal fluid and serum samples from 20 patients with subacute sclerosing panencephalitis (SSPE), 12 patients with multiple sclerosis (MS), and 11 controls with high measles virus antibody titers in serum. In patients with SSPE, measles virus-specific antibodies were found mainly in the IgG1 subclass and the IgG subclass distribution remained unchanged, irrespective of the clinical stage or duration of the disease. In patients with MS and in controls, measles virus activity was also associated mainly with IgG1. However, the activity was significantly lower than that found in patients with SSPE. The results suggest that there is no primary abnormality in humoral immune response to measles virus in patients with MS. The disproportionately high levels of the measles virus-specific IgG1 subclass found in patients with SSPE may be due to persistent antigenic stimulation or reflect a defect in immunoregulatory mechanisms in response to viral infection.
Fecal isolation of Pseudomonas aeruginosa from patients with cystic fibrosis
Authors: Agnarsson U , Glass S , Govan J R
Published: January 2020

Fecal isolation of Pseudomonas aeruginosa was observed in 8 of 10 patients with cystic fibrosis who at the time of sampling also exhibited colonization of the respiratory tract. In contrast, P. aeruginosa cells were isolated at low frequency (9.1%) from the stools of 44 patients with cystic fibrosis with no previous history of chronic colonization. The results of this study suggest that the gastrointestinal tract is not a significant chronic reservoir of P. aeruginosa prior to pulmonary colonization.
Sialic acid levels and lag time of growth in chemically defined medium containing 200 mM phosphate among strains of various serotypes of Streptococcus agalactiae
Authors: Nagano Y , Nagano N , Takahashi S , Suzuki A , Okuwaki Y
Published: January 2020

The type-specific capsular polysaccharide antigen of Streptococcus agalactiae has in previous experimental studies been considered a significant antiphagocytic factor, whereas the lipoteichoic acid moiety has been suggested to be a factor in adherence to human fetal cell lines. Since epidemiological data concerning these cell constituents in strains from the genital tract are lacking, we attempted serotyping and analysis of these constituents of 100 vaginal isolates. The capsular polysaccharide level was shown to be the amount of sialic acid that occupied the terminal side chains of the polysaccharide. We carried out a study to ascertain whether strains exhibited a lag time of growth in a chemically defined medium containing 200 mM phosphate, which has been suggested to be characteristic of strains with high lipoteichoic acid levels. Strains were classified, on the basis of the results of distribution of sialic acid levels, into three categories: (i) strains with a low sialic acid content of equal to or less than 9 micrograms/mg of cell dry weight; (ii) strains with a moderate sialic acid content of more than 9 but less than 12 micrograms/mg of cell dry weight; and (iii) strains with a high sialic acid content of equal to or more than 12 micrograms/mg of cell dry weight. Strains that belonged to the last category, which, as previous experimental data indicate, are potentially virulent strains, were significantly distributed among isolates of types Ia (P less than 0.001) and III (P less than 0.05). On the other hand, strains exhibiting a lag time of growth in the above-mentioned medium were detected to a significant extent in type III isolates (P <0.02). These results may be related to the epidemiological finding that isolates from neonates with late-onset infection were more frequently serotype Ia and III isolates.
Application of a numerical index of discriminatory power to a comparison of four physiochemical typing methods for Candida albicans
Authors: Hunter P R , Fraser C A
Published: January 2020

Four typing methods for the strain differentiation of Candida albicans (morphotyping, resistotyping, extracellular enzyme production, and carbon source assimilation reactions) were compared on a single population of 100 strains. An index of discriminatory power was used to facilitate this comparison. Extracellular enzyme production had poor discriminatory power, and carbon source assimilation reactions were poorly reproducible. Morphotyping and resistotyping had acceptable reproducibility and discrimination. The use of resistotyping and morphotyping in parallel enhanced discrimination without an unacceptable decrease in reproducibility, although other combinations did not enhance discrimination because reproductibility was impaired.
Serological response of patients with leprosy to a 28- to 30-kilodalton protein doublet from early cultures of Mycobacterium bovis BCG
Authors: Pessolani M C , Rumjanek F D , Marques M A , de Melo F S , Sarno E N
Published: January 2020

Analysis by Western immunoblotting of Mycobacterium bovis BCG short-term culture filtrates with a pool of serum samples from lepromatous leprosy patients revealed an immunodominant protein doublet with apparent molecular masses of 28 and 30 kilodaltons (kDa). The humoral response to these antigens was also investigated by using individual serum samples from patients representative of the whole spectrum of leprosy and from tuberculosis patients, as well as from contacts of leprosy patients and control groups. The protein doublet was recognized by 92% of the sera from patients with lepromatous leprosy (51 of 56), whereas essentially negative results were obtained with sera from the other groups. Similar immunodominant bands were also detected by Western blotting analysis of sonic extracts of seven other slow- and fast-growing mycobacterial species, suggesting a broad distribution of these antigens within the genus. Analysis of the purified doublet by Western blotting after two-dimensional gel electrophoresis fractionation showed that the 28- and 30-kDa doublet consisted of at least five different components with pls from 5.2 to 5.7 and molecular masses from 28 to 31 kDa. These results indicate that the protein doublet could be used as a potential marker in the diagnosis of lepromatous leprosy.
Identification of Clostridium difficile in stool specimens by culture-enhanced gas-liquid chromatography
Authors: Johnson L L , McFarland L V , Dearing P , Raisys V , Schoenknecht F D
Published: January 2020

We have developed a sensitive and specific method for the identification of Clostridium difficile in stool specimens based on the detection of metabolic breakdown products of the organism by gas-liquid chromatography after incubation of stool samples in a selective broth medium containing cefoxitin. Use of this approach to test samples from two different populations of patients at separate medical centers showed this method to be superior to plate cultures or cytotoxin testing alone for both populations. The combined results from the two patient populations showed that 225 of 226 confirmed isolates were identified correctly, resulting in a sensitivity of 99.6% and a specificity of 99.0%. This method eliminates the delay caused by subculturing for tests requiring a pure isolate. The culture phase amplifies even low numbers of C. difficile in fecal samples (due to low in vivo concentrations or delayed transport) and thus increases sensitivity. Other advantages include the ability to detect C. difficile in the mixed flora of the stool and the ability of most clinical laboratories to use this procedure. Given the complexities of the detection of C. difficile toxins and the increasing importance of this organism as a nosocomial agent, culture-based methods remain the preferred approach to screening and routine workup for cases of diarrhea.
Cerebral phaeohyphomycosis caused by Chaetomium globosum in a renal transplant recipient
Authors: Anandi V , John T J , Walter A , Shastry J C , Lalitha M K , Padhye A A , Ajello L , Chandler F W
Published: January 2020

A 32-year-old male patient developed headaches, vomiting, blurring of vision, and focal seizures of the left side of the face 2 months after a renal transplant. He developed a brain abscess and died. Direct KOH examination of the brain tissue demonstrated hyaline as well as dematiaceous, septate hyphae. Histologic examination of brain sections revealed polymorphous fungal elements consisting of septate, dark-pigmented hyphae, intercalary and terminal swollen fungal cells, and budding yeastlike cells characteristic of phaeohyphomycosis. Chaetomium globosum was isolated from the brain tissue on all of the fungal media used. This case represents the first histologically and culturally documented phaeohyphomycotic brain infection caused by C. globosum.
Evaluation of a rapid method of extracting DNA from stool samples for use in hybridization assays
Authors: Coll P , Phillips K , Tenover F C
Published: January 2020

The ability of the Extractor system (Molecular Biosystems, Inc., San Diego, Calif.) to isolate nucleic acid (NA) from stool samples for use in hybridization assays was investigated. Crude NA was recovered from 45 of 50 stool samples by using this system. The amount of NA recovered varied considerably depending on the microbial flora present in the sample (mean +/- standard deviation, 50.2 +/- 46.7 micrograms; range, 2 to 228 micrograms) but did not correlate with the consistency of the sample. Samples containing primarily gram-positive organisms or yeast cells gave lower yields of NA (less than 10 micrograms) than those containing gram-negative bacilli. The five samples which did not yield NA were sterile when cultured aerobically on blood agar plates. Samples of the 45 stools yielding NA were inoculated into broth and grown overnight, and a 10-microliters sample of broth was spotted onto nitrocellulose filters. The NA samples recovered from the Extractor column were applied to nylon membranes by using the Centri-dot system. The NA on the broth blots and the NA on the Centri-dot filters were hybridized with a 310-base-pair probe specific for the 2"-O-aminoglycoside adenylyltransferase [ANT(2")] resistance gene. The Extractor-Centri-dot system demonstrated 61.9% sensitivity and 95.8% specificity in detecting the ANT(2") gene in stool samples containing colonies demonstrating the ANT(2") phenotype. The positive and negative predictive values of the NA blot were 92.8 and 74.2%, respectively.
Nonculture detection of Haemophilus influenzae in sputum with monoclonal antibodies specific for outer membrane lipoprotein P6
Authors: Groeneveld K , van Alphen L , van Ketel R J , Geelen-van den Broek L , Eijk P P , Zanen H C
Published: January 2020

Isolation of Haemophilus influenzae from sputum is hampered by overgrowth by other microorganisms or by antibiotic treatment of the patient. To overcome this problem in the detection of H. influenzae, an in situ immunoperoxidase staining technique was developed with monoclonal antibody (MAb) 8BD9, immunoglobulin subclass G2a. MAb 8BD9 appeared to be directed to an epitope on the outer membrane lipoprotein P6 of H. influenzae. The species specificity of MAb 8BD9 was analyzed by staining isolates from different bacterial species. MAb 8BD9 reacted with all 300 H. influenzae strains tested and with H. aegyptius and H. haemolyticus. Twenty-six of 30 H. parainfluenzae strains, other Haemophilus species, and other bacterial species often isolated from sputum were not stained. The staining technique was compared with culture of 845 routinely obtained sputum samples from patients with respiratory tract infections. In 829 sputa (98.1%), the results of both techniques were in agreement; 173 were positive for H. influenzae, and 656 were negative. With 14 sputum samples, the staining method gave a positive result for H. influenzae, but the bacterium was not cultured. This could be ascribed to antibiotic treatment of the patient (n = 7), the presence of other MAb 8BD9-positive Haemophilus species in the sputum (n = 5), and overgrowth by swarming Proteus mirabilis or by Branhamella catarrhalis. In the immunoperoxidase- and Gram-stained smears of two sputum samples, no bacteria were seen, although some H. influenzae was cultured. On the basis of these results, we conclude that immunoperoxidase staining with MAb 8BD9 is a fast and reliable extension of the available detection techniques for H. influenzae.
Lack of protection against bacterial infections in patients with advanced cancer treated by biologic response modifiers
Authors: Maoleekoonpairoj S , Mittelman A , Savona S , Ahmed T , Puccio C , Gafney E , Skelos A , Arnold P , Coombe N , Baskind P
Published: January 2020

A survey of patients with advanced cancer treated by biologic response modifiers (BRMs), including (i) recombinant interleukin-2 and lymphokine-activated killer cells, (ii) recombinant interleukin-2 and alpha interferon, and (iii) tumor necrosis factor, was done. A total of 52 patients were reviewed. A total of 73 courses of BRMs were administered. Prior to the initiation of therapy, all patients were infection free and not receiving antibiotics. Twelve patients developed bacteremia during treatment with these BRMs. Five of these 12 patients had catheter-related bacteremia. Six patients had bacteremic infections without an obvious source, and one patient had a urinary tract infection with bacteremia. Staphylococcus epidermidis accounted for six of the isolates. Other organisms were Staphylococcus aureus, group B streptococci, viridans group streptococci, and gram-negative bacilli. This was an unexpectedly high incidence of bacterial infections in patients treated with BRMs. These BRMs have been previously shown to be efficacious against infections (by bacteria and other intracellular organisms) in experimental animals. In this study BRMs did not influence host defense mechanisms or offer protection against bacterial infections.
Enumeration of viral antigen-reactive helper T lymphocytes in human peripheral blood by limiting dilution for analysis of viral antigen-reactive T-cell pools in virus-seropositive and virus-seronegative individuals
Authors: Clouse K A , Adams P W , Orosz C G
Published: January 2020

A limiting-dilution analysis technique was developed which enumerates human T cells with the capacity to secrete T-cell growth factors such as interleukin 2 after contact with herpes simplex virus type 1 (HSV-1) or cytomegalovirus (CMV) antigens (operationally defined as virus-reactive helper T cells [HTL]). By using this limiting-dilution analysis technique, the peripheral blood of HSV-seropositive individuals was analyzed for the frequency of HSV antigen-reactive HTL and for the ability either to proliferate or to secrete detectable T-cell growth factors in conventional HSV antigen-stimulated lymphocyte cultures. We found that the magnitudes of the latter two responses did not correlate directly with the frequency estimates of HSV antigen-reactive HTL. The study was expanded to analyze both HSV and CMV reactivities within individuals. Those who were seropositive for HSV or CMV were found to have relatively high HTL frequencies for the viral antigens to which they were sensitized. However, those who were seronegative for one of the viruses often had HTL reactive with that virus in their peripheral blood. These latter HTL frequencies were highly variable and ranged from undetectable to quite prominent, even within the same individual at different times. In general, it was found that viral antigen-reactive serologic activity does not necessarily reflect the status of viral antigen-reactive cell-mediated immunity in humans and that viral antigen-induced T-cell responses may be unexpectedly complex, rather than absent, in individuals who are seronegative for a particular virus.
Analysis of Brucella lipopolysaccharide with specific and cross-reacting monoclonal antibodies
Authors: Palmer D A , Douglas J T
Published: January 2020

Monoclonal antibodies which bind Brucella A lipopolysaccharide (LPS)-specific, M LPS-specific, or cross-reactive epitopes were used as reagents in quantitative dot blot, Western blot (immunoblot), and immunoprecipitation analysis of Brucella whole cells, whole-cell extracts, and purified LPS preparations. This set of monoclonal antibodies detected four unique epitopes on Brucella LPS. The specificity of monoclonal antibodies reactive with Brucella unique (A and M) and common (C and C/Y) LPS epitopes was demonstrated by blot analysis. The serotype specificity of monoclonal antibodies for A LPS of B. abortus 1119.3 or M LPS of Brucella melitensis 16M was confirmed. Type C monoclonal antibodies recognized epitopes on Brucella A and M LPS and did not cross-react with Yersinia enterocolitica O:9. In Western blots, type C monoclonal antibodies were bound by epitopes on Brucella A and M LPSs ranging in Mrs from 30,000 to 70,000, relative to marker proteins. Type C/Y monoclonal antibodies were cross-reactive with Y. enterocolitica O:9 and recognized Brucella A LPS epitopes with a restricted Mr ranging only from 40,000 to 50,000, relative to marker proteins. Type C/Y monoclonal antibodies also displayed a more restricted pattern of binding to Brucella M LPS. The monoclonal antibodies were able to detect 5 to 50 pg of a purified A LPS preparation in dot blots. The limits of detection by the monoclonal antibodies of a purified M LPS preparation ranged from 0.05 to 50 pg. Monoclonal antibody analysis of whole-cell preparations also demonstrated quantitative differences in the presence of the respective epitopes. The binding profiles of the monoclonal antibodies to Brucella whole cells varied between acetone- and chloroform-killed organisms as well as between species and strains. The lower limit of detection of any whole-cell preparation by the dot blot technique was 10(5) CFU. Binding profiles in Western blots and endotoxin activity of immunoprecipitates obtained with these monoclonal antibodies further defined the Brucella LPS antigens. These monoclonal antibodies and the techniques described may be useful in monitoring the antigenic content of Brucella vaccines and diagnostics.
DNA probe for the identification of Histoplasma capsulatum
Authors: Keath E J , Spitzer E D , Painter A A , Travis S J , Kobayashi G S , Medoff G
Published: January 2020

A 1.85-kilobase HindIII nuclear DNA probe from Histoplasma capsulatum G217B detected polymorphic restriction fragments within whole-cell DNA from different clinical isolates of H. capsulatum, consistent with the previous system of classification. The probe failed to hybridize to DNA from Blastomyces dermatitidis, Candida spp., Saccharomyces cerevisiae, Sepedonium chrysospermum, and Chrysosporium keratinophilum under low-stringency conditions and therefore may have value as a diagnostic reagent to identify H. capsulatum.
Human herpesvirus 6 infection and Kawasaki disease
Authors: Okano M , Luka J , Thiele G M , Sakiyama Y , Matsumoto S , Purtilo D T
Published: January 2020

Eighteen of a total of 22 serum specimens (81.8%) from patients with Kawasaki disease were positive for immunoglobulin G or M antibodies to human herpesvirus 6, whereas 10 of 16 age- and sex-matched healthy controls (62.5%) were seropositive. Additionally, increased geometric mean antibody titers of immunoglobulin G were shown in these patients. These results suggest that the status of human herpesvirus 6 infection may be a reflection of the immunologic alterations that are associated with Kawasaki disease.
Evaluation of four newer antimicrobial agents in the Avantage susceptibility test system
Authors: Wright D N , Matsen J M , DiPersio J R , Kirk M , Saxon B , Ficorilli S M , Spencer H J
Published: January 2020

Antimicrobial elution disks containing amoxicillin-clavulanic acid (Augmentin), cefotetan, ciprofloxacin, or norfloxacin were tested in the Avantage automated susceptibility test system. Performance was compared against an agar diffusion procedure in a three-site collaborative study. Results of 1,500 comparison with amoxicillin-clavulanic acid showed a full accord (agreement of both systems) of 93.6% and an essential accord (agreement excluding minor discrepancies) of 97.6%. Results for cefotetan showed a full accord of 95.1% and an essential accord of 98.3% by the two methods. Results for both ciprofloxacin and norfloxacin were in full accord for more than 98% of tests with gram-negative bacilli and staphylococci, but tests with enterococci gave 38 and 26.1% minor discrepancies (the result of one method was resistant or susceptible and the result of the other method was intermediate), respectively. The results indicated that the Avantage test system is accurate and reliable and provides appropriate determination of bacterial susceptibility with the four antibiotics tested.
Rapid screening test for the diagnosis of rotavirus infection
Authors: Chudzio T , Kasatiya S , Irvine N , Sankar-Mistry P
Published: January 2020

The early diagnosis of human rotavirus infection is essential for effective patient management and infection control. We report here a rapid, easy-to-perform, and inexpensive test for rotavirus detection. The viral RNA is extracted directly from the stools and electrophoresed on 1% agarose gels. Currently available immunoassays for routine diagnostic purposes are directed at the common group A-specific antigen. As reports become available on human gastroenteritis caused by the atypical or novel rotaviruses, this technique presents an added advantage in that it can detect both group A and non-group A rotaviruses.
Clinical significance of Blastocystis hominis
Authors: Qadri S M , al-Okaili G A , al-Dayel F
Published: January 2020

A total of 19,252 stool specimens from 12,136 patients were examined by direct microscopy and the ethyl acetate-Formalin concentration method during the last 2 years. All liquid specimens and those in which parasite identification was difficult or equivocal were also examined in trichrome-stained preparations. A total of 3,070 intestinal parasites were seen in 2,889 patients. Blastocystis hominis was found in fecal material from 647 patients (17.5%). A total of 132 cases (25.6%) were observed to be in association with other enteric pathogens. B. hominis in large numbers was present as the only parasite or with other commensals in 515 specimens from patients (79.6%). Of these patients, 239 (46.4%) had symptoms, the most common being abdominal pain (87.9%), constipation (32.2%), diarrhea (23.4%), alternating diarrhea and constipation (14.5%), vomiting (12.5%), and fatigue (10.5%). Forty-three (18%) of the patients were treated with metronidazole (0.5 to 1.0 g/day) because of recurrent symptoms and the presence of large numbers of B. hominis cells in repeated stool specimens. After 7 to 10 days of treatment, all patients became asymptomatic with negative stools on follow-up examinations for B. hominis.
Evaluation of microparticle enzyme immunoassays for immunoglobulins G and M to rubella virus and Toxoplasma gondii on the Abbott IMx automated analyzer
Authors: Schaefer L E , Dyke J W , Meglio F D , Murray P R , Crafts W , Niles A C
Published: January 2020

The ability of the Abbott IMx automated analyzer to detect immunoglobulin G (IgG) and IgM antibodies to rubella virus and to Toxoplasma gondii was compared with the abilities of RUBAZYME, RUBAZYME-M, ABBOTT TOXO-G enzyme immunoassay, and ABBOTT TOXO-M enzyme immunoassay, respectively. Specimens that produced discordant results were evaluated by RUBACELL II, Behring Enzygnost-Rubella enzyme-linked immunosorbent assay, Behring Enzygnost Toxoplasmosis/IgG, and bioMerieux Toxo-ISAGA (immunosorbent agglutination assay), respectively. After resolution of discordant results, IMx Rubella IgG, IMx Rubella IgM, IMx Toxo IgG, and IMx Toxo IgM antibody assays had sensitivities of 99.9, 100, 98.0, and 100%; specificities of 98.9, 99.0, 97.5, and 98.7%; and accuracies of 99.8, 99.3, 97.8, and 98.8%, respectively.
Detection of infectious bursal disease viruses by using cloned cDNA probes
Authors: Jackwood D J , Kibenge F S , Mercado C C
Published: January 2020

A molecular clone representing 445 base pairs at the 3' end of infectious bursal disease virus (IBDV) genome segment B was used in a dot blot hybridization assay to detect viral RNA from cell culture and from chicken bursa and spleen tissue specimens. The cloned nucleotide sequence represents approximately 14% of the virus-encoded polymerase (VP-1) gene. The lower detection limit of radiolabeled probes prepared from this clone was 0.1 ng of IBDV double-stranded RNA. The probe had broad specificity and was used to detect four serotype 1 IBDV strains and one serotype 2 IBDV strain. This probe, however, did not cross-react with nucleic acid extracted from nine unrelated poultry viruses. A rapid procedure for isolation of IBDV genomic RNA from bursa and spleen tissue specimens was developed and used with the dot blot hybridization assay to detect IBDV strains in tissue samples from experimentally infected and commercially reared chickens.