bluuebunny/crossref_metadata_2025 huggingface dataset:
Toxoplasma antigens recognized by human immunoglobulin A antibodies
Authors: Huskinson J , Thulliez P , Remington J S
Published: January 2020

The immunoglobulin A (IgA) antibody response to antigens of Toxoplasma gondii during the acute and chronic stages of T. gondii infection was studied by immunoblotting with a nonreduced antigen (NRA) preparation. Serum samples were from individuals with acute or chronic T. gondii infection, and sequential samples were from women who seroconverted during gestation or congenitally infected infants. IgA antibodies to a variety of antigens were present in sera from each of the groups, irrespective of the titer in an IgA enzyme-linked immunosorbent assay. The predominant NRA recognized by IgA antibodies in sera from each of the four groups had an approximate molecular weight of 30,000. Other prominent NRAs had approximate molecular weights of 35,000 and 5,000; however, the latter NRA was not present in sera of congenitally infected infants.
Serodiagnosis of Helicobacter pylori infection in childhood
Authors: Thomas J E , Whatmore A M , Barer M R , Eastham E J , Kehoe M A
Published: January 2020

Sera from 100 children (ages, 6 to 16 years) presenting with upper gastrointestinal symptoms were examined for antibodies to Helicobacter pylori by enzyme-linked immunosorbent assay (ELISA) based on crude, loosely cell-associated antigens and a partially purified urease antigen preparation. All children underwent endoscopy, and 20 children were shown to have H. pylori infection by histology or direct culture. Serum anti-H. pylori immunoglobulin G (IgG) levels (crude antigen) were clearly raised in the infected group, particularly after preabsorption of sera against a Campylobacter jejuni antigen preparation, while IgM and IgA ELISA determinations did not discriminate between infected and H. pylori-negative patients. Only 14 children in the infected group had raised anti-urease IgG levels. Two patients in whom the organism was not demonstrated or cultured had raised specific IgG levels against both crude and urease antigens and pathological features consistent with H. pylori disease. Immunoblotting studies did not reveal any single protein antigen or simple combination of antigens that could be considered as a candidate for a more defined serodiagnostic reagent. Anti-H. pylori antibody determinations (crude antigen) performed on posttreatment samples from children in whom the organism could no longer be demonstrated suggested that sustained IgG levels may not be a reliable index of treatment failure. An IgG ELISA based on crude, loosely cell-associated antigens of H. pylori can be used for the serodiagnosis of H. pylori infection in childhood.
Polymerase chain reaction for detection of adenoviruses in stool samples
Authors: Allard A , Girones R , Juto P , Wadell G
Published: January 2020

The usefulness of the polymerase chain reaction (PCR) method for diagnosing adenovirus infections was investigated. Several primers, including primers specific for the hexon-coding region and for enteric adenovirus types 40 and 41, were evaluated. The PCR method was validated against cell culturing in routine diagnostic work and against restriction enzyme analysis of viral DNA. Sixty diagnostic specimens were selected for evaluation by the PCR method. Twenty of the 60 specimens were found positive on the basis of cytopathic effects and latex agglutination (Adenolex [Orion Diagnostica, Helsinki, Finland]), and 16 were identified and typed as adenoviruses by polyacrylamide gel electrophoresis. PCR was performed on all 60 specimens in parallel directly on diluted stool samples and on viral DNA extracted from cells inoculated with the same stool samples. When the general hexon primers were used 51 of the 60 specimens from infected cell cultures were found positive by PCR, whereas only 13 specimens were found positive when PCR was performed directly on stool samples. With the use of selective primers for enteric adenoviruses 16 of the 60 cell cultures were found to exhibit amplification products by PCR, whereas 4 were detected in stool samples. None of the 60 specimens were found positive by PCR when an adenovirus type 40-specific primer pair was used. PCR was found to be a fast, sensitive, and reliable method for the detection of adenoviruses in diarrheal disease, provided the amplifications were performed directly on diluted stool samples.
Respiratory tract carrier rates of Moraxella (Branhamella) catarrhalis in adults and children and interpretation of the isolation of M. catarrhalis from sputum
Authors: Vaneechoutte M , Verschraegen G , Claeys G , Weise B , Van den Abeele A M
Published: January 2020

Nonselective media and previously described selective media were used to study the occurrence of Moraxella (Branhamella) catarrhalis in sputum samples of good and poor quality and in samples taken from different sites of the upper respiratory tracts of healthy subjects. It was found that in healthy adults the carrier rate was 5.4%, as opposed to 50.8% in children and 26.5% in people older than 60 years. M. catarrhalis was recovered significantly more often from sputum samples of good quality (5%) than from poor quality samples (0.5%), and when present, it was found mostly in the presence of high inocula. From these data gathered from healthy and diseased subjects, it is concluded that the presence of M. catarrhalis in the sputum of adults is rarely due to oronasopharyngeal contamination of the sputum. Similar findings reported by others are discussed, and the origins of the currently held concept that M. catarrhalis is a common commensal organism of the human upper respiratory tract are traced.
Quantification of human cytomegalovirus viremia by using monoclonal antibodies to different viral proteins
Authors: Gerna G , Revello M G , Percivalle E , Zavattoni M , Parea M , Battaglia M
Published: January 2020

Human cytomegalovirus (HCMV) viremia in peripheral blood polymorphonuclear leukocytes (PMNLs) from 187 immunosuppressed patients (79 heart transplant recipients and 108 patients with acquired immunodeficiency syndrome [AIDS]) was investigated. Five mouse monoclonal antibodies (MAbs), one specifically reactive to HCMV immediate-early antigen (IEA) in PMNLs, two specifically reactive to IEA in infected cell cultures, and two specifically reactive to late antigens, were used in immunofluorescence and/or immunoperoxidase test systems for (i) detection of HCMV IEA in human embryonic lung fibroblast (HELF) cell cultures inoculated with PMNL samples, (ii) direct detection of HCMV IEA in PMNL nuclei of cytospin preparations, and (iii) identification of HCMV isolates from PMNL samples in HELF cells. Quantification of viremia was achieved by counting the number of infected PMNLs per 2 x 10(5) cells examined directly on cytospin preparations as well as by counting the number of IEA-positive HELF cells inoculated with 2 x 10(5) PMNLs. A significant correlation was found between the number of infected PMNLs and the number of infected HELF cells. When the number of infected PMNLs per 2 x 10(5) cells was greater than 10, both methods (PMNL IEA and HELF IEA) gave concordant results; when it was greater than 80/2 x 10(5) cells, clinical symptoms were consistently associated with HCMV viremia. Ten patients with heart transplants and three patients with AIDS who had high or increasing levels of HCMV viremia underwent antiviral treatment with ganciclovir. Treatment was discontinued only after disappearance of IEA-positive PMNLs from blood (the last marker of infection to become negative). On the other hand, in the presence of low levels of viremia (less than 10 infected PMNLs per 2 x 10(5) cells), different methods often provided discordant results and overt clinical symptoms were never observed. IEA-negative results with PMNL samples or HELF cells in the presence of positive virus isolation could never be attributed to the inability of IEA MAbs to recognize individual HCMV strains, since all of the relevant viral isolates were recognized by IEA MAbs. In addition, all five MAbs used in the study were capable of identifying all 135 conventional HCMV isolates obtained from the study population. It was concluded that (i) maximal sensitivity in diagnosing HCMV viremia may be achieved by combining different techniques; (ii) direct detection of HCMV IEA in PMNLs, yielding results on the day of blood collection, is a very rapid and sensitive technique, and thus one can rely on it for clinical management of HCMV infections; and (iii) low levels of viremia are clinically irrelevant, whereas high levels are associated with clinical symptoms.
Detection of Lassa virus RNA in specimens from patients with Lassa fever by using the polymerase chain reaction
Authors: Lunkenheimer K , Hufert F T , Schmitz H
Published: January 2020

Suitable oligonucleotide primers and probes were synthesized to amplify Lassa virus (Josiah strain)-specific nucleoprotein and glycoprotein gene fragments by using reverse transcription combined with the polymerase chain reaction (PCR). Our primers did not amplify the related lymphocytic choriomeningitis virus. By using PCR, about 50 50% tissue culture infective doses could be detected in the supernatant of infected cells. Furthermore, in all five serum specimens and four of five urine specimens of patients with acute Lassa fever, viral RNA could be demonstrated. Negative results were obtained with all serum and urine specimens of healthy subjects. Our data suggest that PCR may be applied as an alternative to virus isolation in the rapid diagnosis of Lassa fever.
Specific detection of Haemophilus influenzae type b lipooligosaccharide by immunoassay
Authors: Mertsola J , Munford R S , Ramilo O , Sáez-Llorens X , Mustafa M M , McCracken G H , Hansen E J
Published: January 2020

Three monoclonal antibody (MAb)-based immunoassays were developed for specific detection of Haemophilus influenzae type b (Hib) lipooligosaccharide (LOS). (i) Hib LOS was captured onto microtiter plates by polyclonal Hib-directed antibodies and detected with MAbs to the oligosaccharide component of Hib LOS in an enzyme-linked immunosorbent assay, (ii) The high affinity of polymyxin B for lipid A was used to bind Hib LOS to microtiter wells, and the oligosaccharide-specific MAbs were used as the detection system in the polymyxin B-MAb assay. (iii) Hib LOS solubilized in detergent was captured by MAbs, and the immobilized LOS was detected with a chromogenic Limulus amebocyte lysate method in the immunolimulus assay. Endotoxin concentrations were measured in in vitro samples and cerebrospinal fluid samples from rabbits with experimental Hib meningitis. The results were compared with those obtained with the standard chromogenic Limulus amebocyte lysate assay. There were significant correlations between the results of all four assays. These new immunoassays provide methods for specific detection of Hib LOS in laboratory fluids and in research involving quantification of Hib endotoxin in experimental animal models.
Longitudinal study of the biotypes of Gardnerella vaginalis
Authors: Briselden A M , Hillier S L
Published: January 2020

Gardnerella vaginalis is the predominant vaginal microorganism in women with bacterial vaginosis. However, this organism is also frequently isolated from women without signs or symptoms of vaginitis. Earlier studies have not revealed whether certain biotypes of G. vaginalis are more often associated with bacterial vaginosis or are more common in women who acquire bacterial vaginosis. We used a typing scheme based on tests for beta-galactosidase, hippurate hydrolysis, and lipase, using oleate as a substrate. Of 261 strains tested, the distribution of biotypes observed was as follows: 1, 13%; 2, 9%; 3, 5%; 4, 7%; 5, 41%; 6, 15%; and 8, 10%. Biotype 7 was not observed. The distributions of biotypes from women with and without bacterial vaginosis were found to be significantly different, with the lipase-positive biotypes (biotypes 1, 2, 3, and 4) being more predominant in women with vaginosis (41 versus 23%, P = 0.003). Of 40 women with normal vaginal flora at the index visit who remained normal at follow-up, 23 (57%) acquired a new biotype of G. vaginalis. By comparison, 90% of the 30 women who developed bacterial vaginosis acquired a new biotype of G. vaginalis (P = 0.003). Women with bacterial vaginosis at the index visit who were not treated were no more likely than normal women to have a shift in G. vaginalis biotype. However, 86% of the 30 women with bacterial vaginosis who were treated with an antibiotic at the index visit acquired a different biotype (P = 0.04 compared with the value for untreated women) regardless of treatment success. A trend toward the acquisition of a new biotype was observed among women who had contact with a new sexual partner (81 versus 65%, P = 0.15). These data demonstrate that the lipase-positive isolates of G. vaginalis are associated with bacterial vaginosis. Women who acquire bacterial vaginosis are more likely to have a shift in biotype than women who had normal flora at he follow-up, suggesting that the G. vaginalis isolates recovered from women who develop bacterial vaginosis represent newly acquired strains rather than overgrowth of previously colonizing biotypes.
Detection of Cryptosporidium parvum oocysts in bovine feces by monoclonal antibody capture enzyme-linked immunosorbent assay
Authors: Anusz K Z , Mason P H , Riggs M W , Perryman L E
Published: January 2020

A monoclonal antibody enzyme-linked immunosorbent assay (ELISA) was developed to detect Cryptosporidium parvum oocysts in bovine feces. Fecal oocysts were concentrated by centrifugation through Formalin-ethyl acetate solution and captured with monoclonal antibody 18.280.2 reactive with C. parvum oocysts. Captured oocysts were detected with goat anti-oocyst serum, following the addition of a peroxidase conjugate of rabbit anti-goat immunoglobulin and O-phenylenediamine substrate. The assay was specific for Cryptosporidium sp. oocysts and did not detect oocysts of Eimeria auburnensis, Eimeria bovis, Eimeria ellipsoidalis, or Eimeria zuernii. Assay sensitivity allowed detection of 3 x 10(5) oocysts per ml of feces, compared with 1 x 10(6) oocysts per ml detected by examination of acid-fast-stained fecal smears and 1 x 10(3) oocysts per ml detected by indirect immunofluorescence. The capture ELISA was suitable for diagnostic analysis of bovine fecal samples and for assessment of oocyst shedding in experimentally infected calves. This assay may also prove useful for diagnostic assessment of humans in which cryptosporidiosis is suspected.
Diagnosis of Epstein-Barr virus infection in hairy leukoplakia by using nucleic acid hybridization and noninvasive techniques
Authors: De Souza Y G , Freese U K , Greenspan D , Greenspan J S
Published: January 2020

The presence of Epstein-Barr virus (EBV) DNA in the epithelial cells of oral hairy leukoplakia is the confirming criterion in the diagnosis of this lesion, which occurs mainly in persons infected by the human immunodeficiency virus. Because hairy leukoplakia often presages the development of the acquired immune deficiency syndrome, it is important that suspicious lesions be accurately diagnosed. Commonly, biopsy tissue is removed for detection of EBV DNA by in situ hybridization, but biopsy is contraindicated in some patients. This study evaluated filter and cytospin in situ hybridization, two noninvasive techniques that examine epithelial cells swabbed from the surfaces of the lesions, for their sensitivity in detecting EBV DNA. As compared with tissue in situ hybridization, the filter and cytospin techniques had sensitivities of 100 and 92%, respectively. We conclude that these two noninvasive techniques can provide the clinician with an accurate alternative to biopsy whenever this human immunodeficiency virus-associated lesion is suspected.
Quality control criteria for testing the susceptibility of anaerobic bacteria to meropenem
Authors: Zabransky R J , Bayola-Mueller L , Jenkins S G , Pitkin D R , Schreckenberger P C , Celig D M , Spiegel C A
Published: January 2020

Reference values for quality control of in vitro susceptibility tests with meropenem against anaerobic bacteria were determined in a multilaboratory study by the approved National Committee for Clinical Laboratory Standards agar dilution method for the four quality control strains. The study protocol also included the evaluation of microdilution testing, medium additives, and multiple lots of media. The recommended MIC control ranges for three of the control organisms are as follows: Bacteroides fragilis ATCC 25285, 0.06 to 0.125 micrograms/ml; Bacteroides thetaiotaomicron ATCC 29741, 0.125 to 0.5 micrograms/ml; and Eubacterium lentum ATCC 43055, 0.125 to 0.5 micrograms/ml. The modal MIC for Clostridium perfringens ATCC 13124 was at or below the lowest concentration of meropenem tested, and no values are recommended.
Rapid isolation of herpes simplex virus by using mink lung and rhabdomyosarcoma cell cultures
Authors: Johnston S L , Wellens K , Siegel C S
Published: January 2020

Highly sensitive and rapid results can be obtained by isolating herpes simplex virus from clinical specimens in simple cell culture with rhabdomyosarcoma (RD) cells. In this study, 3,186 clinical specimens were inoculated into locally produced, equivalent-age RD and mink lung (ML) cells. Of 727 positive isolates, all (100%) were isolated from RD cells and only 691 (95%) were isolated from ML cells. Furthermore, 162 of the positive isolates (22%) were isolated in RD cells earlier than in ML cells. RD cells are continuous and can be cultivated in house without decreasing sensitivity as the passage number increases. They produce a highly distinguishable cytopathic effect in response to herpes simplex virus and maintain intense confirmatory staining patterns.
Multicenter evaluation of Microring YT, a new method of yeast identification
Authors: Shankland G S , Hopwood V , Forster R A , Evans E G , Richardson M D , Warnock D W
Published: January 2020

Microring YT was evaluated and compared with established methods for the identification of 142 clinical yeast isolates. Only 75 isolates (52.8%) were correctly identified by Microring YT. Results with this test were often difficult to read and subject to interlaboratory variations.
Bedside inoculation of blood culture bottles with ascitic fluid is superior to delayed inoculation in the detection of spontaneous bacterial peritonitis
Authors: Runyon B A , Antillon M R , Akriviadis E A , McHutchison J G
Published: January 2020

Ascitic fluids from patients suspected of having spontaneous bacterial peritonitis were inoculated into blood culture bottles (i) at the bedside and (ii) in the laboratory after a delay. In 29 episodes in which the bedside bottles were culture positive, only 22 (75.9%) of the laboratory-inoculated sets demonstrated growth; this difference was statistically significant (P less than 0.02).
Cross contamination of viral specimens related to shell vial caps
Authors: Salmon V C , Kenyon B R , Overall J C
Published: January 2020

A cluster of poliovirus type 3-positive specimens cultured in shell vials led to the discovery of significant cross contamination between vials related to the type of vial cap. Simulated testing with colored media in the shell vials indicated that the most contamination occurred with the plastic snap cap and that the least occurred with the screw cap. Measures to prevent cross contamination in clinical virology laboratories are discussed.
Rapid presumptive identification of gram-negative rods directly from blood cultures by simple enzymatic tests
Authors: Sepúlveda J L , Stager C E , Davis J R
Published: January 2020

Gram-negative rods were presumptively identified directly from blood cultures within 15 min as Escherichia coli, a member of the Klebsiella-Enterobacter group, or oxidase positive. Samples of artificially seeded blood cultures (193 cultures) and patient blood cultures (78 cultures) were filtered into a Dynadepth test card with the Bac-T-Screen instrument (Vitek, Inc., Hazelwood, Mo.). Triton X-100 was then filtered into the test card to lyse the blood cells but not the entrapped bacteria, and either methylumbelliferone-labeled substrates or oxidase reagent was applied to the filter surface. The oxidase test was read within 30 s, and the methylumbelliferone and indole tests were read after a 10-min incubation at room temperature. Positive beta-galactosidase, beta-glucuronidase, and indole test results predicted the identification of E. coli with a 96 to 100% sensitivity and a 99 to 100% specificity. Positive beta-xylosidase and beta-galactosidase test results and negative oxidase and beta-glucuronidase test results were 85 to 93% sensitive and 100% specific for a Klebsiella-Enterobacter organism. A positive oxidase test result and negative beta-glucuronidase, beta-xylosidase, and indole test results were highly predictive of Pseudomonas aeruginosa (sensitivity, 100%; specificity, 99%). The procedures described are rapid and simple and provide a direct presumptive identification of the gram-negative rods most commonly found in blood cultures.
Cytopathic astrovirus isolated from porcine acute gastroenteritis in an established cell line derived from porcine embryonic kidney
Authors: Shimizu M , Shirai J , Narita M , Yamane T
Published: January 2020

A cytopathic astrovirus was isolated from pigs with acute diarrhea in an established cell line that was derived from porcine embryonic kidneys with the aid of trypsin. The virus showed a distinct cytopathic effect characterized by an enlargement of cells and the appearance of fine granules in the cytoplasm. Porcine astrovirus was shown to have an RNA genome, as determined by the effect of 5-iodo-2'-deoxyuridine on its replication, and five polypeptides with molecular masses of 13,000, 30,000, 31,000, 36,000, and 39,000 daltons; and it was shown to be stable to lipid solvents and heating at 50 degrees C for 30 min but somewhat labile to acid (pH 3.0). The buoyant density of the isolate determined in CsCl was 1.35 g/ml. Seroconversion to the virus was evident in the paired serum specimens obtained from pigs with diarrhea that were housed at the farm where the disease occurred. The neutralization test on serum specimens collected randomly from 128 adult pigs of eight herds revealed that 50 of the serum specimens were positive for antibody to porcine astrovirus, although there was considerable variation in the prevalence among herds, ranging from 0 to 83%. Hysterectomy-produced, colostrum-deprived, 4-day-old pigs developed mild diarrhea after oral exposure to porcine astrovirus propagated in the cell culture; and the virus was isolated again from diarrheal stool specimens.
Hemolysin patterns of Actinobacillus pleuropneumoniae
Authors: Frey J , Nicolet J
Published: January 2020

The secreted hemolytic activities produced by the reference strains and field isolates of the 12 serotypes and 2 subtypes of Actinobacillus pleuropneumoniae were analyzed. Serotype 1 produced a Ca2(+)-inducible hemolysin, which was previously characterized as a 105-kilodalton protein and was named hemolysin I (HlyI). Serotypes 2, 4, 6, 7, and 8 produced a different hemolytic activity that was not inducible by Ca2+ but required this ion for its activity. The hemolytic activity produced by these serotypes was much weaker than that found in serotype 1 and was not neutralized by rabbit antibodies against HlyI. It was, however, neutralized by serum from pigs that were experimentally infected with a serotype 2 strain and was called hemolysin II (HlyII). Serotypes 5a, 5b, 9, 10, and 11 produced both HlyI and HlyII. In these strains, HlyI was the major contributor to the hemolytic activity. The remaining serotypes, 3 and 12, produced a very weak hemolytic activity, which was not further analyzed. Immunoblot analysis of the culture supernatants from all 12 serotypes with rabbit polyclonal antibodies directed against HlyI revealed reactions with a protein in the 105-kilodalton size range for all serotypes, indicating that HlyI and HlyII might be serologically related. Strains producing active HlyI seem to belong to serotypes that are generally considered to be virulent types and that are frequently isolated from pigs in severe pleuropneumonia outbreaks.
Evaluation of two single-plate incubation systems and the anaerobic chamber for the cultivation of anaerobic bacteria
Authors: Downes J , Mangels J I , Holden J , Ferraro M J , Baron E J
Published: January 2020

Three systems that are available for the incubation of anaerobic organisms were evaluated to assess their ability to support the growth of 25 anaerobic stock strains and to successfully recover anaerobic bacteria from clinical specimens. These were the anaerobic chamber, the Anaerobic Pouch System Catalyst-Free (Difco Laboratories, Detroit, Mich.), and the Bio-Bag Environmental Chamber Type A (Marion Scientific, Div. Marion Laboratories, Inc., Kansas City, Mo.). Three study centers were involved, the Wadsworth Anaerobe Laboratory (Los Angeles, Calif.), the Good Samaritan Hospital (San Jose, Calif.), and the Massachusetts General Hospital (Boston). A total of 171 anaerobic organisms were isolated from 49 clinical specimens that were cultured at the three test centers. Of these, 169 (99%) were recovered from media that were incubated in the anaerobic chamber, 163 (95%) were recovered from the Anaerobic Pouch, and 147 (86%) were recovered from the Bio-Bag. A similar trend was seen with the stock strains, in which the anaerobic chamber often supported better growth of the organisms than did either of the bag systems.
Serotypes of Escherichia coli that hybridized with DNA probes for genes encoding Shiga-like toxin I, Shiga-like toxin II, and serogroup O157 enterhemorrhagic E. coli fimbriae isolated from adults with diarrhea in Thailand
Authors: Bettelheim K A , Brown J E , Lolekha S , Echeverria P
Published: January 2020

Escherichia coli strains isolated from adults with diarrhea in Bangkok, Thailand, were examined for hybridization with DNA probes for genes that code for Shiga-like toxin (SLT)-I, SLT-II, and serogroup O157 enterhemorrhagic E. coli (EHEC) fimbriae. Seven isolates that hybridized with the SLT-I, SLT-II, and O157 EHEC fimbria probes and produced verocytotoxin (VT; group A) were isolated from two patients with diarrhea. Two strains that hybridized with only the SLT-II probe and the O157 EHEC fimbria probe and were VT+ (group B) were isolated from two patients with diarrhea, 7 strains that hybridized with only the SLT-II probe and were VT+ (group C) were isolated from two patients with diarrhea, and 26 strains that hybridized with only the O157 EHEC fimbria probe and were VT- (group D) were isolated from four patients with diarrhea. Seven strains in group A had serotypes O2:H1 (n = 1), O110:H19 (n = 1), and Ont:H8 (n = 5); 2 strains in group B were O112ab:H21 (n = 1) and O113:H21 (n = 1); 7 strains in group C were O6:H28 (n = 1), O22:H16 (n = 1), O52:H25 (n = 1), O112ab:H21 (n = 1), OR:H45 (n = 2), and OR:H11 (n = 1); and 26 strains in group D were O76:H7 (n = 18), O146:H3 (n = 2), O146:H10 (n = 1), O146:Hnt (n = 1), OR:H16 (n = 1), Ont:H2 (n = 1), Ont:H8 (n = 1) and Ont:H16 (n = 1). In Thailand, E. coli strains that hybridized with SLT-I, SLT-II, and O157 EHEC fimbria probes were of a variety of serotypes, none of which were O157:H7.
Use of synthetic oligonucleotide DNA probes for the identification of Bacteroides gingivalis
Authors: Moncla B J , Braham P , Dix K , Watanabe S , Schwartz D
Published: January 2020

Six different oligonucleotide probes complementary to the hypervariable regions of 16S rRNA of Bacteroides gingivalis were tested for specificity and sensitivity against 77 field strains of B. gingivalis and 105 strains of 12 other Bacteroides species. The data demonstrated that these probes were very specific (range, 0.85 to 1.00) and sensitive (1.00). Some limited cross-reactions with other Bacteroides species were observed. Four of these probes should be useful for rapid detection and identification of B. gingivalis.
Use of partially purified 54-kilodalton antigen for diagnosis of nocardiosis by Western blot (immunoblot) assay
Authors: Boiron P , Provost F
Published: January 2020

A Western blot (immunoblot) assay is presented for the diagnosis of nocardiosis with a specific immunodominant 54-kilodalton (kDa) antigen purified from a culture filtrate of Nocardia asteroides by immunoaffinity chromatography. The chromatography column was prepared with immunoglobulin G obtained from sera from patients with lepromatous leprosy. Unbound solutes consisted of specific, partially purified N. asteroides antigens, primarily a 54-kDa band, accompanied by two others of 31 and 62 kDa. The Western blot technique was applied to detecting the immunologic response to nocardiae. Immunodetection was performed according to the biotin-avidin system, which greatly improved the detection of antibodies even in immunosuppressed hosts. Each of 16 serum samples from immunosuppressed or immunocompetent patients infected with N. asteroides reacted with the 54-kDa band, and two reacted with the 31- and 62-kDa bands. Each of the serum specimens obtained from patients with mycetoma caused by Nocardia brasiliensis or Rhodococcus rhodochrous reacted with the 54-kDa band. There was no reaction to either the 54- or the 31-kDa antigen with all serum samples obtained from patients with tuberculosis, except one, with all serum samples obtained from patients with leprosy, or with all sera obtained from healthy controls. The 54-kDa protein is a candidate to be used as a probe to study the humoral immunologic response to nocardiae.
Detection of Entamoeba histolytica immunoglobulins G and M to plasma membrane antigen by enzyme-linked immunosorbent assay
Authors: Sathar M A , Bredenkamp B L , Gathiram V , Simjee A E , Jackson T F
Published: January 2020

Sixty-one serum specimens from 22 patients with clinically diagnosed amoebic liver abscess (ALA), 10 hospitalized patients with a variety of diseases other than amoebiasis, 12 normal healthy controls, and 17 subjects from an amoebiasis-endemic area were assayed by enzyme-linked immunosorbent assay (ELISA). The plasma membrane fraction of axenic cultures of Entamoeba histolytica HK9 separated from other subcellular fractions by differential centrifugation was used as the antigen to detect specific immunoglobulin G (IgG) and IgM antibodies. Using a single serum dilution of 1/100 and optical densities at 492 nm of 0.200 and 0.250 as the cutoff values for the IgM and IgG ELISAs, their respective sensitivities in 22 ALA patients were 91% (20 of 22) and 95% (21 of 22). In 22 patients (10 hospitalized and 12 normal healthy controls), the specificities of the IgM and IgG ELISAs were 95% (21 of 22) and 91% (20 of 22), respectively. All five asymptomatic carriers of pathogenic E. histolytica were seropositive by the IgG ELISA and the amoebic gel diffusion test (AGDT). The AGDT was positive for three of six culture-negative controls, while the IgG ELISA was positive for all six. For six asymptomatic carriers of nonpathogenic zymodemes, the AGDT was positive for two, and the IgG ELISA was positive for three. There was an excellent correlation (r = 0.96) between the IgG ELISA and the AGDT. Only one of six culture-negative controls, none of the asymptomatic carriers of pathogenic E. histolytica, and one of six carriers of nonpathogenic E. histolytica were seropositive by the IgM ELISA, thus highlighting the specificity of the IgM ELISA in the diagnosis of ALA. It is believed that the use of plasma membrane fractions has improved the diagnostic potential of the IgM ELISA.
Reactivity of coagulase-negative staphylococci isolated from cow and goat milk with monoclonal antibodies to Staphylococcus aureus capsular polysaccharide types 5 and 8
Authors: Poutrel B , Mendolia C , Sutra L , Fournier J M
Published: January 2020

Monoclonal antibodies to Staphylococcus aureus capsular polysaccharide types 5 and 8 were used in an enzyme-linked immunosorbent assay to serotype 74 and 42 coagulase-negative isolates from cow and goat milk, respectively. Eighteen (15.5%) isolates were typable: 13 Staphylococcus haemolyticus, 1 S. hyicus, 1 S. simulans, and 1 S. warneri from bovine origin and 2 S. lentus from caprine origin. Type 5 was predominant, accounting for about 89% of typable isolates. Reactivity with monoclonal antibodies varied considerably according to isolates. The significance and the potential importance of these findings are discussed.
Discordant results between the broth disk elution and broth microdilution susceptibility tests with Bacteroides fragilis group isolates
Authors: Aldridge K E , Henderberg A , Schiro D D , Sanders C V
Published: January 2020

Susceptibility testing of 161 clinical isolates of the Bacteroides fragilis group was performed to compare interpretive results generated by the broth disk elution and broth microdilution methods recommended by the National Committee for Clinical Laboratory Standards. Among the cephalosporin-cephamycin compounds tested, correlation was poorest for ceftizoxime (71%), ceftriaxone (57%), and cefotaxime (47%); when the tests did not correlate, false resistance was seen 92, 95, and 93% of the time, respectively. Cefotetan and cefoperazone showed lack of correlation in 19 and 20% of the tests, respectively. For cefotetan, false resistance was more frequent, while with cefoperazone, false susceptibility occurred more often. Cefoxitin produced the fewest discrepancies; 10% of the disk elution tests produced either false-resistance or false-susceptibility results. Mezlocillin and piperacillin showed lack of correlation in 8 and 14% of the tests, respectively, and discrepancies were due primarily to false-resistance results. Overall with the beta-lactams, 84% of the discordant interpretive results were false resistance by the broth disk elution test. Clindamycin had a discrepancy rate of 10%, with the majority of discrepancies being false susceptibility disk elution results. Because of the high number of discrepancies noted with ceftizoxime, ceftriaxone, and cefotaxime, we recommend that these drugs not be tested by the disk elution method and that they be tested by a quantitative MIC method such as the broth microdilution test. Furthermore, caution should be exercised when interpreting broth disk elution results with all the beta-lactams included in this study except imipenem. These data indicate the lack of correlation of results between these two tests for many beta-lactams and suggest the need for a reexamination of the disk elution method to provide a more accurately standardized test.
Evaluation of the rapid Patho Dx Latex Strep Grouping Kit
Authors: Beck R , Etzion R , Sandler S , Kohn L
Published: January 2020

A variety of clinical specimens from throat, nose, ear, eye, wounds, urine, and vagina were collected, cultured, and screened for beta-hemolytic streptococci. The Patho Dx Latex Strep Grouping Kit (Diagnostic Product Corp., Los Angeles, Calif.) technique was applied to colonies taken right from the primary cultures. Isolated strains were sent to the reference laboratory where they were grouped by standard techniques. The kappa coefficient of agreement between the Patho Dx Kit and the standard method was 0.958. We believe that, although better agreement was achieved by others with isolated colonies, a very good agreement is also achieved with primary cultures. The fact that the laboratory is able to report an accurate answer after only 24 h seems most advantageous.
Protein banding patterns of the outer membrane-enriched fraction of Bacteroides bivius
Authors: Dinsmoor M J , Ebersole J E , Gibbs R S
Published: January 2020

Bacteroides bivius is a common anaerobe in female genital infections. Although protein banding patterns of outer membrane (OM) preparations of Bacteroides fragilis are well described and are homologous within the species, similar work has not been done with B. bivius. Our aims were to (i) characterize the OM banding patterns of B. bivius and compare them with those of other Bacteroides species and (ii) test clinical isolates of B. bivius against anti-B. bivius and anti-B. fragilis sera to identify different serogroups that might also exhibit different OM banding patterns. OM-enriched fractions of 27 clinical strains of B. bivius, 6 Bacteroides disiens strains, 10 B. fragilis strains, and 12 other Bacteroides strains were prepared, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis was performed. Antisera to B. bivius ATCC 29303 and B. fragilis ATCC 25285 were raised in rabbits and tested against Bacteroides strains in an indirect enzyme-linked immunosorbent assay. All 27 B. bivius strains contained protein bands at 32, 27, 25, and 23 kilodaltons. This pattern was present in only 2 of 28 other strains; both of these were B. disiens. All B. bivius strains were reactive with the anti-B. bivius serum, while only 6 of 39 other strains (2 of 6 B. disiens) were reactive. Non-B. fragilis Bacteroides strains did not react with the anti-B. fragilis serum. Although there was marked homogeneity in the OM banding patterns of B. bivius, some B. disiens strains exhibited similar OM banding patterns. There appears to be some antigenic cross-reactivity between strains of B. bivius and B. disiens and very little with other Bacteroides species. These results may ultimately allow the development of rapid diagnostic tests for the B. bivius-B. disiens group.t
Evaluation of a new latex agglutination test for detection of streptolysin O antibodies
Authors: Gerber M A , Caparas L S , Randolph M F
Published: January 2020

Acute- and convalescent-phase serum specimens were collected from 50 patients with group A streptococcal pharyngitis. The anti-streptolysin O (ASO) titer for each serum specimen was determined by using both the standard neutralization assay and the latex agglutination (LA) test (Rheumagen ASO; Biokit Inc., New Britain, Conn.). When the ASO titers derived by the two methods were compared, the correlation coefficient was 0.93. When the ability of the LA test to demonstrate a significant ASO titer rise (greater than or equal to 2 dilutions) was compared with that of the standard neutralization assay, the LA test had a sensitivity of 91%, a specificity of 86%, a positive predictive value of 83%, and a negative predictive value of 92%. Triplicate LA test determinations were performed on a subset of 31 serum specimens, and for 29 (94%), the repeated ASO titers were all within 1 dilution of each other; the width of the 95% confidence interval for the triplicate measurements of each serum specimen was +/- 32.8 IU. We found the Rheumagen ASO to be a simple, rapid LA procedure for measuring ASO titers that produces results that are highly reproducible, show little lot-to-lot variability, and are comparable to the ASO titers obtained with the standard neutralization assay.
Phenotypic characterization, cellular fatty acid composition, and DNA relatedness of aerococci and comparison to related genera
Authors: Bosley G S , Wallace P L , Moss C W , Steigerwalt A G , Brenner D J , Swenson J M , Hebert G A , Facklam R R
Published: January 2020

Aerococci can be misidentified as streptococci, enterococci, pediococci, lactococci, or leuconostocs. To distinguish the genus and determine if another species is needed in the present taxon, we analyzed 37 aerococci for cellular fatty acids and compared them with 377 strains of gram-positive cocci, including the species type strains from each of the related genera. The cellular fatty acid profile of aerococci was distinguishable from other genera. Two relatively novel fatty acids found in the aerococci were identified as C16:1 omega 9c and C16:1 omega 9t. Eleven strains of aerococci (including a strain originally identified as "Gaffkya" species) were chosen for DNA-DNA reassociation studies with the type strain Aerococcus viridans ATCC 11563; DNAs from eight of these strains were more than 75% related to the type strain and had 1 to 4% divergence in related sequences. The remaining three strains were 60 to 70% related to the type strain, had 7 to 11.5% divergence, and may represent a second species, Aerococcus genospecies 2. beta-Glucuronidase, alpha-galactosidase, and beta-galactosidase were useful in characterizing the aerococci.
Use of Haemophilus test medium for broth microdilution antimicrobial susceptibility testing of Streptococcus pneumoniae
Authors: Jorgensen J H , Maher L A , Howell A W
Published: January 2020

A recently described medium (Haemophilus test medium [HTM]) for antimicrobial susceptibility testing of Haemophilus influenzae was evaluated in this study for broth microdilution testing of Streptococcus pneumoniae. A total of 137 clinical isolates was tested against 11 antimicrobial agents, using Mueller-Hinton broth supplemented with 3% lysed horse blood in parallel with HTM. Inocula of 5 X 10(5) CFU/ml and incubation for 20 to 24 h were used with both media. All isolates of S. pneumoniae produced acceptable growth in both media, and MICs determined in HTM agreed closely with those determined in lysed horse blood. Drugs which provided a MIC within 1 log2 concentration difference in both media included penicillin (100%), ampicillin (98.0%), amoxicillin-clavulanate (100%), ampicillin-sulbactam (100%), cephalexin (98.9%), cefaclor (96.8%), cefuroxime (99.0%), chloramphenicol (96.2%), tetracycline (96.2%), and erythromycin (100%). HTM MICs with trimethoprim-sulfamethoxazole were 1 to 2 log2 concentration increments higher in 92.0% of isolates than MICs determined in lysed horse blood. Based on the results of this study, HTM appears to represent a promising alternative medium for broth microdilution susceptibility testing of S. pneumoniae.
Enzymatic RNA amplification of the enteroviruses
Authors: Rotbart H A
Published: January 2020

Enteroviruses are among the most common causes of childhood infection. Current diagnostic techniques are often too slow and too insensitive to benefit the patient optimally. This report describes a modified polymerase chain reaction technique by which enteroviral RNA can be amplified, over a few hours, to a level detectable by agarose mini-gel electrophoresis or nucleic acid hybridization or both. Three oligomeric regions of great homology among the enteroviruses were identified and designated as a potential primer pair and probe. With this combination, all 11 of the enterovirus serotypes tested, representing the major subgroups of these pathogens, were successfully amplified and detected. The sensitivity and rapidity of this new assay speak to its potential clinical applicability in the diagnosis of enterovirus infections.
Encapsulation of Staphylococcus aureus isolates from mastitic milk: relationship between capsular polysaccharide types 5 and 8 and colony morphology in serum-soft agar, clumping factor, teichoic acid, and protein A
Authors: Sutra L , Mendolia C , Rainard P , Poutrel B
Published: January 2020

A total of 193 Staphylococcus aureus isolates from bovine, caprine, and ovine mastitis producing type 5 or 8 capsular polysaccharides were investigated for colony morphology in serum-soft agar and agglutinability by an anti-teichoic acid serum, after cultivation in modified staphylococcus medium no. 110. Also, 40 of these strains were cultivated in brain heart infusion and submitted to clumping factor and protein A detection tests. Considering capsular serotyping as a reference method, diffuse growth in serum-soft agar and inagglutinability by anti-teichoic acid serum identified, respectively, 57.5 and 45% of encapsulated strains cultivated in brain heart infusion and 85.5 and 77.2% of those cultivated in modified staphylococcus medium 110. Consequently, these indirect techniques underestimated encapsulation and were greatly influenced by culture conditions. Whatever the medium used, diffuse colony morphology in serum-soft agar was generally characterized by a masking of teichoic acid and protein A. By contrast, these surface antigens were detected in association with compact morphology; the presence of a thin or discontinuous capsular material could explain this result. Moreover, the masking of teichoic acid and the removal of capsular polysaccharide by washing in saline suggest that type 8 capsular polysaccharide is more abundant and labile than type 5.
Agreement between visual and automated UniScept API readings
Authors: O'Hara C M , Rhoden D L , Smith P B
Published: January 2020

The UniScept API system was evaluated for agreement of visual versus automated readings of both its identification panels and its antimicrobial susceptibility panels. The biochemical responses of 340 oxidase-negative and oxidase-positive fermentative bacterial cultures were read both visually and automatically in the UniScept API 20E system. Automated and visual readings agreed with 99.3% of the biochemicals. Of the 45 tests that disagreed, the tests for indole and citrate were most often in disagreement. A total of 470 fermentative and nonfermentative cultures were used in the UniScept MIC system to compare visual and automated readings of susceptibility results with 17 antimicrobial agents. Agreement within +/- 1 dilution occurred with 94.1% of the enteric fermenters and with 91.7% of the other cultures. Comparison of visual and automated readings resulted in very major discrepancies in 0.95% of the readings, with the largest percentage of discrepancies associated with glucose nonfermenters (1.8%). It was felt that an automated reading is an acceptable alternative to a visual reading of the biochemicals but that 0.95% was just within the acceptable range of the 1% allowable very major discrepancies in the automated reading of susceptibilities.
Comparison of different medium bases for the semiquantitative isolation of anaerobes from vaginal secretions
Authors: Sheppard A , Cammarata C , Martin D H
Published: January 2020

Two studies were performed to determine the best medium for the isolation of anaerobes from vaginal secretions. In the first, three different medium bases (brucella, Centers for Disease Control [CDC], and Schaedler) were compared semiquantitatively for ability to support the growth of gram-negative anaerobes from vaginal fluid. Media were supplemented with laked sheep blood, kanamycin, and vancomycin. The brucella base agar formulation supported the growth of anaerobic gram-negative bacilli better than either the CDC or Schaedler base agar formulation. In a second study, nonselective brucella and CDC base sheep blood agar were compared for ability to support the growth of anaerobic gram-positive cocci. Anaerobic gram-positive cocci grew in higher concentrations on CDC base agar than on brucella base agar. On the basis of these observations, we recommend that a CDC base sheep blood agar be used for the nonselective plate and a brucella base plate supplemented with laked sheep blood, kanamycin, and vancomycin be used for isolation of gram-negative bacilli in studies of the anaerobic flora of the female genital tract.