bluuebunny/crossref_metadata_2025 huggingface dataset:
Rapid detection of bacteria in cerebrospinal fluid by immunofluorescence staining on membrane filters
Authors: Lim L C , Pennell D R , Schell R F
Published: January 2020

A filter-fluorescent-antibody (FFA) staining procedure was developed for detection of bacteria in cerebrospinal fluid (CSF). The sensitivity of this procedure was determined and compared with that of the slide Gram stain of centrifuged samples, latex agglutination, and a previously developed filter Gram stain. Serial 10-fold dilutions of filter-sterilized CSF seeded with logarithmic-phase organisms were examined by each method and cultured to determine colony-forming bacteria. The bacteria included Haemophilus influenzae type b; Neisseria meningitidis group B, C, and W135; Streptococcus pneumoniae types 6A and 23F; and group B Streptococcus species. FFA was found to be equal in sensitivity to the filter Gram stain (P greater than 0.30). Both filter-staining procedures had greater sensitivity than the slide Gram stain of centrifuged sediment (P less than 0.02) and latex agglutination (P less than 0.0001). Addition of human leukocytes at a concentration of 5 to 10 cells per oil immersion field did not decrease sensitivity of the FFA procedure, although background fluorescence increased. FFA is a rapid and dependable procedure for detection of low numbers of bacteria in CSF. Evaluation of FFA with clinical specimens is needed.
Serological classification by monoclonal antibodies of Rickettsia tsutsugamushi isolated in Korea
Authors: Chang W H , Kang J S , Lee W K , Choi M S , Lee J H
Published: January 2020

Antigenic types of 113 strains of Rickettsia tsutsugamushi isolated from Korean patients were analyzed by using murine polyclonal and monoclonal antibodies. The isolates can be classified in six groups according to their reaction to a panel of monoclonal antibodies. Nine isolates of group I were identified as the Gilliam serotype, and 13 isolates of groups II and III were identified as the Karp serotype. There were two groups that were considered to be a mixture of groups I and II or groups I and III, respectively. The remaining 88 strains of group IV had a unique antigenic determinant that was not present in the prototype strains (Karp, Kato, Gilliam), in addition to sharing common antigens with the prototype strains. Therefore these strains, which are more prevalent in Korea, seem to belong to a new serotype closely related to the Karp serotype.
Implications of antibodies to pyruvylated glucose in healthy populations for mycobacterioses and other infectious diseases
Authors: Thayer W R , Bozic C M , Camphausen R T , McNeil M
Published: January 2020

Members of the Mycobacterium avium-Mycobacterium intracellulare (MAI) complex are typeable because each serovar is characterized by its own specific antigenic glycolipid. By means of an enzyme-linked immunosorbent assay, we studied serum specimens obtained from 148 healthy college students for antibodies to these glycopeptidolipids. Ninety-two (61.5%) of the serum specimens were positive to the specific glycolipid antigen from MAI serovar 8. In a study of a pediatric population, antibodies appeared to develop during adolescence. Individuals with overt mycobacterial disease had a significantly lower incidence (tuberculosis patients, 34.5%; leprosy patients, 25%). We found a lower incidence of positive results in a survey of 96 Japanese serum specimens (29.1%), but the results from a survey of sera obtained from Bombay, India, indicated a large degree of reactivity (55.5%). Antibodies to other MAI serovars (serovars 2, 4, and 11) were not found, except antibodies to MAI serovar 21 were seen in the same individuals with antibodies to serovar 8. The dominant epitope of the MAI serovar 8-specific glycopeptidolipid is a terminal pyruvylated 3-O-methylglucose residue [4,6-(1'-carboxyethylidene)-3-O-methyl-alpha-D-glucopyranosyl] unit, whereas that of the MAI serovar 21 has the same terminal pyruvylated glucose devoid of the 3-methoxy group. Thus the antibodies appear specific for the pyruvylated glucose. It is unclear whether the high prevalence of antibodies to these epitopes reflects a high incidence of subclinical colonization or infection with certain MAI serovars or whether they are acquired through contact with some other related antigen source.
Development of a highly specific and sensitive rubella immunoglobulin M antibody capture enzyme immunoassay that uses enzyme-labeled antigen
Authors: Seppänen H
Published: January 2020

An enzyme immunoassay (EIA) for serum immunoglobulin M (IgM) antibodies to rubella virus based on enzyme labeling of viral antigen was developed. The sensitivity of the EIA for the detection of recent rubella virus infection was evaluated by using 115 rubella-IgM-antibody-positive serum specimens, which were confirmed as positive by Rubazyme M (Abbott Diagnostics). In addition, 12 individuals, 2 of whom were exposed to rubella through vaccination and 10 of whom were exposed through natural infection, were studied, and the results were compared with those obtained by indirect EIA (Rubelisa M; Electro-Nucleonics, Inc.) and immunoblotting. The sensitivity of the newly developed EIA with sera from these individuals was 100%. Serum specimens from two patients indicated that the IgM antibodies were detected by the newly developed EIA at the same time as IgM antibodies were detected by immunoblotting and before positive reactions were detected by an indirect EIA. The reference population consisted of 564 healthy blood donors and hospitalized patients (150 serum specimens). In addition, 145 serum specimens commonly giving false-positive reactions in conventional rubella IgM EIAs were studied. With these specimens, no false-positive reactions were observed. Positive IgM responses, which could not be confirmed by immunoblotting, were observed in two samples from the reference population. However, these two samples were rubella IgG positive. The overall specificity of the EIA was 99.8%.
Virulence-related genes in ColV plasmids of Escherichia coli isolated from human blood and intestines
Authors: Fernandez-Beros M E , Kissel V , Lior H , Cabello F C
Published: January 2020

DNA probes for the colicin V, traT, iss, and iu genes were used in this study of four representative ColV plasmids together with 200 Escherichia coli strains isolated from the stools of patients with diarrhea and 146 E. coli strains isolated from the blood of patients with bacteremia. The study indicated that the ColV plasmids are heterogeneous. Southern and colony hybridization analyses showed that in most of the colicin V-producing intestinal E. coli strains, the colicin V genes are located in the chromosome (14 of 16); in most of the colicin V-producing E. coli strains isolated from the blood, they are located in plasmids (18 of 22). In both intestinal and blood E. coli isolates, the traT, iss, and aerobactin receptor genes were present at similar frequencies, but the frequency of the aerobactin synthesis genes was significantly different. The aerobactin receptor gene was present in 25% of the intestinal E. coli strains that lack the aerobactin synthesis gene. In the blood isolates, the aerobactin synthesis and receptor genes were present at almost equal frequencies. Among the colicin V-producing isolates, the iss, traT, and iu genes were present in 95.5, 86.4, and 90.9% of the blood isolates and in only 68.8, 43.8, and 81.3% of the intestinal isolates, respectively. The ColV plasmids from blood isolates that were tested for the presence of traT, iss, and iu genes were homogeneous and had DNA sequences that hybridized with each of the probes. On the other hand, the two intestinal strains containing ColV genes in a plasmid were heterogeneous in regard to the carriage of these genes. The presence of ColV is not restricted to specific O types.
Quantitative culture of endocervical Chlamydia trachomatis
Authors: Barnes R C , Katz B P , Rolfs R T , Batteiger B , Caine V , Jones R B
Published: January 2020

We examined the number of Chlamydia trachomatis inclusions produced in the initial passage of cell cultures of endocervical specimens from 1,231 women with positive chlamydial cultures who attended a sexually transmitted diseases clinic. Youth, white race, oral contraceptive use, and concurrent infection by Neisseria gonorrhoeae were associated with high chlamydial inclusion counts. Youth, white race, and oral contraceptive use were independent determinants of a high chlamydial inclusion count in women without concurrent gonorrhea but not in women with gonorrhea. Results of our study suggest that the degree of chlamydial excretion from the infected cervix may be influenced by characteristics of the patient being tested and may affect the ability to detect C. trachomatis in different patient groups.
Effect of altered headspace atmosphere on yield and speed of detection of the Oxoid Signal blood culture system versus the BACTEC radiometric system
Authors: Weinstein M P , Mirrett S , Reimer L G , Reller L B
Published: January 2020

The one-bottle Oxoid Signal blood culture system altered to provide a more aerobic bottle headspace was reassessed in a comparative study versus the two-bottle BACTEC radiometric system in 5,426 blood cultures. The BACTEC system detected more microorganisms (P less than 0.02), particularly anaerobes (P less than 0.05) and fungi (P less than 0.05).
Simple assay of calcium dependency for virulent plasmid-bearing clones of Yersinia enterocolitica
Authors: Bhaduri S , Turner-Jones C , Taylor M M , Lachica R V
Published: January 2020

A simplified procedure to detect the calcium dependency of virulent plasmid-bearing strains of Yersinia enterocolitica was developed. A low-calcium, agarose-based medium of brain heat infusion with added magnesium effectively differentiated plasmid-bearing and plasmidless isolates. Further, the expression of calcium dependency in plasmid-bearing strains of Y. enterocolitica as measured by the average colony diameter was proportional to the calcium concentration of the assay.
Comparison of conventional and shell vial cultures for detecting cytomegalovirus infection
Authors: Rabella N , Drew W L
Published: January 2020

We tested 606 specimens for cytomegalovirus infection. Fifty-two specimens were positive by conventional culture versus forty-three by pre-cytopathic effect detection. Twenty-nine specimens were positive by both methods. Because of toxicity, specimens should be set up on both conventional and shell vial culture systems to achieve maximum sensitivity.
Evaluation of enzyme-linked immunoassay systems for detection of human immunodeficiency virus type 1 antibody from filter paper disks impregnated with whole blood
Authors: Beebe J L , Briggs L C
Published: January 2020

Five commercial enzyme-linked immunoassay systems for the detection of human immunodeficiency virus type 1 antibody from filter paper disks impregnated with whole blood were evaluated for technical and operational performance. All five systems performed adequately in the technical challenges posed, with specificities in excess of 99% for 1,020 specimens. In a serial dilution sensitivity challenge, all of the kits were able to detect specific antibody within one dilution of a Western blot (immunoblot) standard, except for a Du Pont Co. kit, which detected antibody within two dilutions of the standard. The Du Pont assay showed the least variation in control values between test runs and between lots. All of the systems produced acceptable results, but their operational parameters differed significantly.
Biotype-specific probe for Vibrio cholerae serogroup O1
Authors: Alm R A , Manning P A
Published: January 2020

The O1 serogroup of Vibrio cholerae can be divided into two biotypes, El Tor and Classical. Current tests to distinguish between these biotypes are often difficult to interpret. On the basis of the difference in sequence of the hlyA gene in these biotypes, we have developed a simple probe that can easily and reliably differentiate between the two biotypes.
Rapid diagnosis of typhoid fever through identification of Salmonella typhi within 18 hours of specimen acquisition by culture of the mononuclear cell-platelet fraction of blood
Authors: Rubin F A , McWhirter P D , Burr D , Punjabi N H , Lane E , Kumala S , Sudarmono P , Pulungsih S P , Lesmana M , Tjaniadi P
Published: January 2020

Detection of Salmonella typhi in blood by culture of the mononuclear cell-platelet layer was compared with other methods currently used for the diagnosis of typhoid fever. Colonies of S. typhi were present in all mononuclear cell-platelet layer-positive cultures within 18 h of plating and were identified within an additional 10 min by a coagglutination technique. In contrast, identification of all positive cultures by conventional blood culture required 3 days.
Microbiology and potential virulence of Sporothrix cyanescens, a fungus rarely isolated from blood and skin
Authors: Sigler L , Harris J L , Dixon D M , Flis A L , Salkin I F , Kemna M , Duncan R A
Published: January 2020

Sporothrix cyanescens has been recovered from blood and a finger lesion at several medical centers in the United States. The morphology and physiology of these and three additional isolates were studied. S. cyanescens was distinguished from S. schenckii and S. fungorum by white to lavender colonial pigmentation and from S. schenckii also by the formation of secondary conidia. All isolates of S. cyanescens grew well at 37 degrees C, were cycloheximide susceptible, strongly urease positive, and benomyl resistant, failed to hydrolyze starch, and were inhibited by sodium chloride in vitro at a concentration of greater than or equal to 12%. Study of S. cyanescens in a murine model by using intravenous inoculation failed to demonstrate an invasive pathogenic potential. The validity of the transfer of S. cyanescens to the new genus Cerinosterus Moore is discussed.
Characterization and description of "Campylobacter upsaliensis" isolated from human feces
Authors: Goossens H , Pot B , Vlaes L , Van den Borre C , Van den Abbeele R , Van Naelten C , Levy J , Cogniau H , Marbehant P , Verhoef J
Published: January 2020

During a 3-year period, "Campylobacter upsaliensis" was isolated from 99 patients. Phenotypic characterization and numerical analysis of protein electrophoregrams showed evidence that "C. upsaliensis" is a distinct Campylobacter species with unique characteristics. The MBCs of 13 antibiotics were determined. In general, these organisms were highly susceptible to drugs that were present in the selective isolation media, making none of the available selective media suitable for the isolation of "C. upsaliensis." Ten strains were found to be resistant to erythromycin (MBCs, greater than or equal to 12.50 mg/liter). Plasmid DNA was detectable in 89 of the 99 strains; 16 plasmid profiles could be identified. Plasmid pattern 16, containing four plasmids of 52, 32, 5.5, and 2.6 megadaltons, represented 60.7% of the plasmid-containing strains. None of the "C. upsaliensis" strains could be agglutinated with antisera against heat-labile antigens from C. jejuni, C. coli, or C. laridis. "C. upsaliensis" was found to be susceptible to serum killing and was readily phagocytized by human polymorphonuclear cells.
Evaluation of a commercial microtiter system (MicroScan) using both frozen and freeze-dried panels for detection of high-level aminoglycoside resistance in Enterococcus spp
Authors: Fuller S A , Low D E , Simor A E
Published: January 2020

The MicroScan system was compared with agar dilution screen plates for the detection of high-level aminoglycoside resistance in 182 enterococcal isolates. Both the frozen Gram-Positive Combo Type 2 and the freeze-dried Type 5 panels were evaluated. The specificity of both panels for the detection of streptomycin and gentamicin resistance was 100%. However, the sensitivities for the detection of gentamicin and streptomycin resistance were 84 and 31%, respectively, for the Type 2 panels and 90 and 41%, respectively, for the Type 5 panels. The sensitivities of these panels for the detection of enterococcal high-level aminoglycoside resistance are inadequate for routine use.
Cellular fatty acid compositions of an unidentified organism and a bacterium associated with cat scratch disease
Authors: Moss C W , Holzer G , Wallace P L , Hollis D G
Published: January 2020

The cellular fatty acid composition of a gram-negative bacterium associated with cat scratch disease was determined by capillary gas chromatography-mass spectrometry (GC-MS). The fatty acid profile of this organism was distinct from those of other bacteria we have tested and was characterized by an unknown acid which was identified as 11-methyloctadec-12-enoic acid. The position of the branched methyl group in this acid was established by GC-MS of the reduced acid, and the location of the double bond was confirmed by GC-MS analysis of dimethyl disulfide derivatives. Another clinical isolate with no known relationship to cat scratch disease but with similar morphological and biochemical features had a similar fatty acid profile, including 11-methyloctadec-12-enoic acid.
Evaluation of direct wet mount parasitological examination of preserved fecal specimens
Authors: Neimeister R , Logan A L , Egleton J H , Kleger B
Published: January 2020

A retrospective study covering a 12-year period was performed to determine the contribution of the direct wet mount microscopic examination to the identification of intestinal parasites in preserved fecal material. It was determined that each of 898 specimens contained at least one parasite when processed by the direct wet mount, concentration, and trichrome staining procedures. Of these procedures, the direct wet mount examination was solely responsible for the identification of 45 (2.9%) of 1,581 parasites identified. This is in contrast to the 15.1 and 12.5% which were found exclusively by the concentration and trichrome methods, respectively. These percentages may vary, depending on the prevalence of parasites and the stages present in the stool specimen.
Microbiologic and clinical evidence supporting the role of Aeromonas caviae as a pediatric enteric pathogen
Authors: Namdari H , Bottone E J
Published: January 2020

Aeromonas caviae was recovered as the sole potential enteric pathogen from the stools of 14 of 17 symptomatic children (10 younger than 1 year of age) while Aeromonas hydrophila, Aeromonas sobria, and Plesiomonas shigelloides were isolated once each. The infants from whom A. caviae was isolated all presented with a watery diarrhea lasting 1 to 3 weeks. None of these infants was breast-fed, and all had a stool pH of greater than 7.5. All of the A. caviae isolates, including a reference strain (ATCC 15468), adhered to HEp-2 cells, and preliminary data showed that they produced a cytotoxin as well. Because A. caviae can survive at an elevated pH, as found in the gastrointestinal tract of formula-fed infants, and because of the adherence and cytotoxin production capabilities of the species, it should be regarded as an enteric pathogen in pediatric patients and most probably among adults as well.
Sensitive solid-phase immune electron microscopy double-antibody technique with gold-immunoglobulin G complexes for detecting rotavirus in cell culture and feces
Authors: Wu B , Mahony J B , Simon G , Chernesky M A
Published: January 2020

A new solid-phase immune electron microscopy double-antibody colloidal-gold technique (SPIEMDAGT) was developed and compared with direct electron microscopy, direct immune electron microscopy, and enzyme immunoassay for detecting rotavirus. Guinea pig and rabbit antirotavirus antisera were used as capture and detector antibodies, respectively, and goat anti-rabbit immunoglobulin G-gold complexes were employed as a label. Animal rotavirus in cell culture media and human virus in stool specimens were detected by this method. On average, SPIEMDAGT detected 800 times more virus particles than direct electron microscopy and 45 times more particles than direct immune electron microscopy and yielded 20% more positives than enzyme immunoassay. SPIEMDAGT could detect not only viral antigen associated with morphologically recognizable particles but also antigen present when whole virus particles were not visible.
Toxin A of Clostridium difficile is a potent cytotoxin
Authors: Tucker K D , Carrig P E , Wilkins T D
Published: January 2020

Clostridium difficile is the cause of antibiotic-associated colitis in humans. The organism produces toxin A, which is generally known as the enterotoxin, and toxin B, which is known as the cytotoxin. Toxin A has been reported to have slight cytotoxic activity; in this study we show that cell lines (F9, OTF9-63, and P19) which express a carbohydrate to which toxin A binds are more sensitive to the toxin. These cell lines can be used as research tools for determining concentrations of biologically active toxin A and should also prove useful for studies of the mechanism of action of the toxin.
Serologic analyses of cottontail rabbits for antibodies to Borrelia burgdorferi
Authors: Magnarelli L A , Anderson J F , McAninch J B
Published: January 2020

An enzyme-linked immunosorbent assay was developed to detect antibodies to Borrelia burgdorferi in cottontail rabbits captured in Millbrook, N.Y., and New York, N.Y. Five antigenically variable strains of B. burgdorferi were analyzed to determine the variability of serologic test results. In analyses of 79 serum samples, seropositivity ranged from 56% for a strain cultured from kidney tissues of a cottontail rabbit to 68% for a strain isolated from a larva of Ixodes dentatus, a tick that parasitized a cottontail rabbit. There were false-positive results when reference rabbit antisera to B. hermsii and Treponema pallidum were screened against B. burgdorferi. Cross-reactivity with antisera to Leptospira interrogans serovars was less pronounced. Western blot (immunoblot) analyses revealed reactivities of test sera to two or more surface or subsurface proteins of B. burgdorferi with approximate molecular masses of 18, 25 to 27, 34, 36, 41, and 59 kilodaltons. Cottontail rabbits respond immunologically to B. burgdorferi, but the observed variations in serologic test results should not be a limitation in field and laboratory investigations of Lyme borreliosis.
Protein G-based enzyme-linked immunosorbent assay for anti-MPB70 antibodies in bovine tuberculosis
Authors: Harboe M , Wiker H G , Duncan J R , Garcia M M , Dukes T W , Brooks B W , Turcotte C , Nagai S
Published: January 2020

MPB70 is a highly species specific protein which is secreted from Mycobacterium bovis during culture. To investigate whether antibodies against MPB70 can be used as an indicator of infection with M. bovis, an enzyme-linked immunosorbent assay was developed, based on the use of biotinylated protein G, to provide a common indicator for antibody formation in different species. During experimental infection with M. bovis in cattle, a characteristic pattern of anti-MPB70 antibody production was observed with an initial flat plateau followed by a marked rise 18 to 20 weeks after infection. Skin testing with bovine tuberculin purified protein derivative (PPD), which was shown to contain antibody-reactive MPB70, was a potent stimulator of antibody production in infected animals. In experimentally infected cattle, we observed an inverse relationship between antibody activity and delayed-type hypersensitivity skin test reactions. In natural M. bovis infections, skin testing with PPD was also a potent stimulator of anti-MPB70 formation. Comparison between the enzyme-linked immunosorbent assay for antibodies to MPB70 and that for antibodies to the widely cross-reacting M. bovis BCG antigen 85B in animals with M. bovis, Mycobacterium avium, Mycobacterium paratuberculosis, and Corynebacterium pseudotuberculosis infections showed that formation of antibody to MPB70 was highly specific for infection with M. bovis. The use of an MPB70-containing PPD preparation for skin testing followed by this anti-MPB70 assay is a highly specific indicator of M. bovis infection. Adjustment of the test conditions is expected to provide an increased sensitivity of the procedure for the diagnosis of natural M. bovis infections.
Use of HL cells for improved isolation and passage of Chlamydia pneumoniae
Authors: Cles L D , Stamm W E
Published: January 2020

We compared growth of the recently discovered respiratory pathogen Chlamydia pneumoniae in McCoy, HeLa 229, BHK-21, and HL cells. When cells were not pretreated with DEAE-dextran, HL cells had significantly higher mean numbers of inclusion-forming units (IFUs) on initial inoculation than the other cell lines. When cells were pretreated with DEAE-dextran, HeLa 229 and HL cells had equivalent mean numbers of IFUs on initial inoculation. HL cells had strikingly higher mean numbers of IFUs in passage than HeLa 229, BHK-21, or McCoy cells. In addition, HL cells did not require pretreatment with DEAE-dextran and could be used from 2 to 4 days after seeding. We conclude that HL cells are an excellent cell culture system for laboratory propagation of C. pneumoniae and may be a more sensitive cell line for initial isolation.
Epidemiology of bacterial pathogens associated with infectious diarrhea in Djibouti
Authors: Mikhail I A , Fox E , Haberberger R L , Ahmed M H , Abbatte E A
Published: January 2020

During a survey examining the causes of diarrhea in the East African country of Djibouti, 140 bacterial pathogens were recovered from 209 diarrheal and 100 control stools. The following pathogens were isolated at comparable frequencies from both diarrheal and control stools: enteroadherent Escherichia coli (EAEC) (10.6 versus 13%), enterotoxigenic E. coli (ETEC) (11 versus 10%), enteropathogenic E. coli (EPEC) (7.7 versus 12%), Salmonella spp. (2.9 versus 3%), and Campylobacter jejuni-C. coli (3.3 versus 5%). Surprisingly, the EAEC strains isolated did not correspond to well-recognized EPEC serogroups. No Yersinia spp., enteroinvasive E. coli, or enterohemorrhagic E. coli were isolated during the course of this study. Only the following two genera were recovered from diarrheal stools exclusively: Shigella spp. (7.7%) and Aeromonas hydrophila group organisms (3.3%). Shigella flexneri was the most common Shigella species isolated. Patients with Shigella species were of a higher average age than were controls (27 versus 13 years), while subjects with Campylobacter or Salmonella species belonged to younger age groups (2.6 and 1.6 years, respectively). Salmonella cases were more often in females. Shigella diarrhea was associated with fecal blood or mucus and leukocytes. ETEC was not associated with nausea or vomiting. Anorexia, weight loss, and fever were associated with the isolation of Salmonella and Aeromonas species. EAEC, ETEC, EPEC, and Shigella species were resistant to most drugs used for treating diarrhea in Africa, while the antibiotic most active against all bacteria tested was norfloxacin. We conclude that in Djibouti in 1989, Shigella and Aeromonas species must be considered as potential pathogens whenever they are isolated from diarrheal stools and that norfloxacin should be considered the drug of choice in adults for treating severe shigellosis and for diarrhea prophylaxis in travelers.
Evaluation of an indirect fluorescent-antibody stain for detection of Pneumocystis carinii in respiratory specimens
Authors: Ng V L , Yajko D M , McPhaul L W , Gartner I , Byford B , Goodman C D , Nassos P S , Sanders C A , Howes E L , Leoung G
Published: January 2020

Two prospective studies were undertaken to evaluate a commercial indirect fluorescent-antibody (IFA) stain for the detection of Pneumocystis carinii in respiratory specimens from individuals at risk for or with the acquired immunodeficiency syndrome. The first study compared IFA with Diff-Quik (DQ; a rapid Giemsa-like stain) for detecting P. carinii in 95 induced sputa obtained from 77 asymptomatic patients who had survived one previous episode of P. carinii pneumonia and who were being treated prophylactically with aerosolized pentamidine. Only one induced sputum specimen was found to contain P. carinii; organisms were detected by both stains. The second study compared the performance of the IFA stain versus DQ, modified toluidine blue O, and Gomori methenamine silver stains for detecting P. carinii in symptomatic individuals at risk for or with acquired immunodeficiency syndrome. Of 182 specimens examined, P. carinii was detected in 105 by one or more stains; the DQ stain detected 73 (70%), the modified toluidine blue O stain detected 75 (71%), the Gomori methenamine silver stain detected 76 (72%), and the IFA stain detected 95 (90%). The IFA stain was more sensitive (P less than 0.01) than the other traditional stains for detecting P. carinii; however, a subsequent clinical evaluation revealed that a subset of IFA-positive-only specimens were from patients whose clinical symptoms resolved without specific anti-P. carinii therapy.
Extended serogrouping scheme for motile, mesophilic Aeromonas species
Authors: Thomas L V , Gross R J , Cheasty T , Rowe B
Published: January 2020

A total of 1,255 strains of motile, mesophilic Aeromonas species isolated from clinical and environmental specimens in the United Kingdom and 258 strains isolated in Australia, Brazil, Peru, and the United States were examined by using antisera for serogroups O1 to O44 (R. Sakazaki and T. Shimada, Jpn. J. Med. Sci. 37:247-255, 1984) and for unpublished serogroup O45 (R. Sakazaki). The typeability rate for strains isolated in the United Kingdom was 35%; the strains isolated in other countries had typeability rates of between 14 and 43%. A total of 52 provisional new serogroups were identified, and the strains with unidentified O groups were examined by using antisera for these provisional new serogroups. The typeability rate for strains isolated in the United Kingdom was increased to 66% (70% of smooth strains). The typeability rates were 76% for A. hydrophila and 63% for both A. caviae and A. sobria. The 52 antisera for the provisional new serogroups increased the typeability rate for strains isolated outside the United Kingdom to between 43 and 68%. This extended serogrouping scheme would be of value in determining the importance of Aeromonas strains as human intestinal pathogens and in investigating the pathogenic mechanisms that may be involved in the production of diarrheal disease.
Effect of endocervical specimen quality on detection of Chlamydia trachomatis and on the incidence of false-positive results with the Chlamydiazyme method
Authors: Kellogg J A , Seiple J W , Murray C L , Levisky J S
Published: January 2020

Duplicate endocervical swabs were collected from 1,675 patients to assess the effects of variations in specimen quality on Chlamydiazyme (Abbott Laboratories) detection of Chlamydia trachomatis and the incidence of false-positive results. One swab (at random) from each patient was tested for C. trachomatis antigen by using the standard Chlamydiazyme procedure. A 200-microliter volume of 0.9% saline was added to the other swab from each patient. After vortexing, 20 microliters was smeared on a slide for Papanicolaou (Pap) staining and the remaining specimen was then tested with the Chlamydiazyme assay. The Chlamydiazyme result was positive for 170 (10.1%) and 165 (9.8%) of the stained and unstained duplicate specimens, respectively (no significant difference). Pap stains on smears from 1,536 (91.7%) of the patients were analyzed, and endocervical and/or metaplastic (E-M) cells were detected in 789 (51.4%) smears. Of these 1,536 stained and analyzed specimens, 150 (9.8%) were Chlamydiazyme positive but only 132 (88.0%) of the positive results were confirmed by repeating the test and using a monoclonal blocking antibody (Abbott). Confirmed Chlamydiazyme-positive results were obtained from only 34 (4.6%) of 747 specimens lacking E-M cells but from 98 (12.4%) of 789 specimens containing the cells (P less than 0.001). Of the 150 initially Chlamydiazyme-positive results obtained with Pap-stained, analyzed specimens, 12 (26.1%) of 46 were falsely positive from specimens lacking E-M cells but only 6 (5.8%) of 104 were falsely positive from specimens containing E-M cells (P less than 0.01). C. trachomatis antigen was detected significantly more frequently and false-positive results were significantly less common from specimens in which E-M cells were detected.
Comparison of restriction endonuclease analysis and phenotypic typing methods for differentiation of Yersinia enterocolitica isolates
Authors: Kapperud G , Nesbakken T , Aleksic S , Mollaret H H
Published: January 2020

Restriction endonuclease analysis of chromosomal DNA (REAC) was used to study polymorphism in restriction fragment patterns among Yersinia enterocolitica isolates belonging to serogroups O3, O5,27, O8, O9, O13, and O21. Using the enzyme HaeIII and electrophoresis on thin (0.75-mm) vertical 5% polyacrylamide gels, we were able to distinguish at least 22 DNA fragment patterns among the 72 strains examined. The method showed the greatest discriminatory power with regard to serogroup O8, within which as many as 10 different DNA fragment patterns were detected among the 16 strains examined. Compared with O8, serogroups O3 and O9 were relatively homogeneous with regard to REAC patterns. The discriminatory power of the method was compared with H-antigen typing, biotyping, phage typing, antimicrobial susceptibility typing, and restriction enzyme analysis of the virulence plasmid (REAP), by means of Simpson's index of diversity. The results showed that REAC and REAP constitute an effective supplement or alternative to conventional phenotypic methods for tracing epidemiologically related isolates of Y. enterocolitica. Our finding that human and porcine isolates exhibited the same REAC, REAP, and H-antigen patterns provides additional support for the hypothesis that pigs play an important role in the epidemiology of human Y. enterocolitica infection.
Specificity of bovine serum antibody to capsular carbohydrate antigens from Pasteurella haemolytica
Authors: McVey D S , Loan R W , Purdy C W , Shuman W J
Published: January 2020

A more complete understanding of the bovine immune response to antigens of Pasteurella haemolytica biotype A, serotype 1, will improve control of bovine respiratory disease (BRD). Sera were obtained from blood samples of calves as they transited the market system of eastern Tennessee and were transported to a feedlot in Texas. The clinical histories and performance data were recorded and compared with serologic findings. The calves underwent a natural challenge of BRD. Serologic and bacteriologic evaluation indicated that P. haemolytica A1 was a significant component of the challenge. Serum antibody titers against P. haemolytica A1 capsular antigens (in enzyme-linked immunosorbent assay and hemolysin-in-gel test) increased by day 15 and continued at high levels through day 56. The animals that remained free of BRD had higher initial serum antibody concentrations than those that succumbed to BRD. The specificity of the immunoglobulin G subclass 1 (IgG1) anticapsular antibody to P. haemolytica A1 increased from day 8 to day 29 as evidenced by a decrease in P. haemolytica A2 absorption inhibition from 60% (day 8) to 15% (day 29). However, IgA, IgG2, and IgM were more serotype specific on both days 8 and 29. There were no significant changes in anti-P. haemolytica A2 antibody titers. Both in vitro complement-dependent bacteriolysis and C3 deposition on the surface of the bacteria increased significantly (P less than 0.01) in a serotype-specific fashion from day 8 to day 29. These calves showed a humoral immune response to capsular polysaccharide antigens of P. haemolytica A1. Such a response may be an important component of immunity to BRD.
Characterization of the cellular antigens of Paracoccidioides brasiliensis yeast form
Authors: Casotto M
Published: January 2020

Antigenic components of the yeast extract of Paracoccidioides brasiliensis Linder 2511 cultured for 3, 8, 20, 30, and 60 days were examined by the Western blot (immunoblot) technique. The 3-day extract was chosen for characterization of the antigenic components because its stability did not vary with time and it contained all antigens identified by patient sera. Antibodies to cross-reacting antigens of P. brasiliensis extracts were detected in sera from patients with histoplasmosis, candidiasis, and aspergillosis. The 58-, 57-, 21-, and 16-kilodalton (kDa) antigens were specific for P. brasiliensis, while the 48- and 45-kDa antigens were specific for paracoccidioidomycosis. The Western blot technique is a useful tool for the diagnosis of disease and revealed heterogeneity in the responses of patient sera. The combination of the 58-, 57-, and 45-kDa proteins confirmed a diagnosis of paracoccidioidomycosis (87% of the cases).
Major Trypanosoma cruzi antigenic determinant in Chagas' heart disease shares homology with the systemic lupus erythematosus ribosomal P protein epitope
Authors: Mesri E A , Levitus G , Hontebeyrie-Joskowicz M , Dighiero G , Van Regenmortel M H , Levin M J
Published: January 2020

A Trypanosoma cruzi lambda gt11 cDNA clone, JL5, expressed a recombinant protein which was found to react predominantly with chronic Chagas' heart disease sera. The cloned 35-residue-long peptide was identified as the carboxyl-terminal portion of a T. cruzi ribosomal P protein. The JL5 13 carboxyl-terminal residues shared a high degree of homology with the systemic lupus erythematosus (SLE) ribosomal P protein epitope. Synthetic peptides comprising the 13 (R-13), 10 (R-10), and 7 (R-7) carboxyl-terminal residues of the JL5 protein were used to study, by enzyme-linked immunosorbent assay, the specificity of the Chagas' disease anti-JL5 and SLE anti-P antibodies. The R-13 peptide defined a linear antigenic determinant of the JL5 recombinant protein. As was proved for JL5, R-13 defined antibody specificities which were significantly increased in chronic Chagas' heart disease patients. Only SLE anti-P positive sera were found to react with JL5 and R-13. Fine epitope mapping showed that Chagas' disease anti-JL5 and SLE anti-P antibodies define similar epitopes within the R-13 peptide. The binding of the SLE sera to JL5 was completely blocked by the R-13 peptide, indicating that the shared specificity between anti-JL5 and anti-P autoantibodies was exclusively limited to the conserved linear epitope(s) within the R-13 peptide. The prevalence of high anti-R-13 antibody titers in Chagas' heart disease patients supports the hypothesis that postulates the existence of autoimmune disorders in Chagas' heart disease.
Procaryotic expression of phosphorylated tegument protein pp65 of human cytomegalovirus and application of recombinant peptides for immunoblot analyses
Authors: Plachter B , Klages S , Hagelmann S , Britt W , Landini M P , Jahn G
Published: January 2020

The tegument of human cytomegalovirus (HCMV) contains a phosphorylated protein of 65 kilodaltons, termed pp65, which was reported to carry significant epitopes for the stimulation of the humoral immune response during natural infection. A monoclonal antibody directed against this protein was used to screen a lambda gt11 cDNA library for recombinant polypeptides. Two DNA fragments from purified lambda clones and one fragment from genomic DNA were used for cloning in a bacterial high-level expression vector. The resulting fusion proteins were tested for their reactivity with a panel of monoclonal antibodies directed against pp65 and with polyspecific anti-HCMV rabbit antisera. The binding site for all the monoclonal antibodies tested was found to be contained in one of the recombinant proteins with a viral portion of 26 amino acids. Immunoblot analyses with HCMV-positive human sera revealed that pp65 alone is not a reliable antigen for serodiagnosis but may be very useful in combination with other HCMV proteins.
Serological response to Chlamydia pneumoniae infection
Authors: Campbell L A , Kuo C C , Wang S P , Grayston J T
Published: January 2020

The human serological response was analyzed by using sera from patients who were serologically positive but isolation negative for Chlamydia pneumoniae and from patients with proven C. pneumoniae infection based on serology and isolation. To assess whether seroreactivity to C. pneumoniae proteins had potential diagnostic value, the cross-reactivities of these sera to other Chlamydia species and of sera from patients infected with C. trachomatis and C. psittaci to C. pneumoniae proteins were determined. In all serum samples from patients with proven C. pneumoniae infections, reactivities were seen with 98-, 68-, 60-, 39.5-, and 30-kilodalton proteins. Similar patterns were seen in sera from patients who were serologically positive and isolation negative. The onset of seropositivity for C. pneumoniae was accompanied by reactivities against presumably shared chlamydial antigens and a C. pneumoniae-specific 98-kilodalton protein.
Comparison of improved BACTEC and Lowenstein-Jensen media for culture of mycobacteria from clinical specimens
Authors: Anargyros P , Astill D S , Lim I S
Published: January 2020

A 4-month trial involving 2,563 routine clinical specimens was conducted to compare the improved BACTEC TB system (12B medium) with the conventional Lowenstein-Jensen (LJ) media for the isolation, identification, and susceptibility testing of mycobacteria. One hundred sixty-two mycobacterial isolates were recovered, 147 (91%) with BACTEC and 118 (73%) with LJ media. Of these, 62 were Mycobacterium tuberculosis complex strains, 59 (95%) of which were isolated with BACTEC and 54 (87%) of which were isolated with LJ media. Of the remaining 100 isolates, which were mycobacteria other than tuberculosis (MOTT), BACTEC and LJ media detected 88 and 64%, respectively. The contamination rate was significantly higher in BACTEC (5%) than in LJ media (3.3%). The mean isolation time for M. tuberculosis complex with BACTEC was 15.5 days, compared with 25.6 days with LJ. For MOTT, the mean isolation times were 5.8 and 21.4 days, respectively. Identification of 32 M. tuberculosis complex isolates and 38 isolates of MOTT by the BACTEC NAP (p-nitro-alpha-acetylamino-beta-hydroxypropiophenone) inhibition test gave 100% agreement with conventional biochemical identifications. The results of susceptibility testing of 18 M. tuberculosis complex isolates with BACTEC agreed completely with those obtained by the resistance ratio method.
Staphylococcal slime: a cautionary tale
Authors: Drewry D T , Galbraith L , Wilkinson B J , Wilkinson S G
Published: January 2020

Slime production by Staphylococcus epidermidis may be important in the adherence to and colonization of biomedical devices, and slime has been proposed to have various effects on the immune system. Attempts were made to isolate, purify, and chemically characterize slime from S. epidermidis cultivated under fluid on tryptic soy broth-agar medium. "Crude slime" from slime-producing strain RP-12 was characterized by a high galactose content. Similar materials in similar yields were isolated from slime-producing strain Kaplan, a non-slime-producing mutant, Kaplan-6A, and sterile medium controls, suggesting that crude slime was derived mainly from the medium. The occurrence of D- and L-galactose and pyruvate and sulfate residues and methylation analysis of these crude slime preparations, monitored by gas-liquid chromatography and mass spectrometry, showed that the agar was the main source of crude slime, suggesting that the preparation was largely an artifact of the growth and isolation procedures. Similar high-galactose-content preparations from both S. epidermidis and Staphylococcus aureus, assumed to be bacterial products and with a variety of biological activities, have been described by other investigators. Growth attached to a solid surface appears to be important for slime production. An accumulation of turned-over cell surface molecules and released macromolecules such as DNA may contribute to slime production. Avoidance of agar and development of a chemically defined medium for slime production are recommended for further studies.
Isolation and identification of autoagglutinating serogroup O:11 Aeromonas strains in the clinical laboratory
Authors: Kokka R P , Janda J M
Published: January 2020

We evaluated the extent to which serogroup O:11 Aeromonas strains could be recovered from both clinical and environmental specimens and the cultural parameters that affected the phenotypic marker (autoagglutination) associated with this group. Of over 200 Aeromonas strains screened, serogroup O:11 was identified only among the phenospecies A. hydrophila and A. sobria and was associated with clinical isolates more frequently than with environmental strains. Blood and wound isolates accounted for almost 50% of all O:11 strains identified. The autoagglutination phenotype associated with O:11 strains could be detected in most commercial liquid media, under a wide range of growth temperatures, and within 15 min of incubation at 100 degrees C. The results suggest that clinical laboratories can recognize this important group of Aeromonas strains by two simple tests.
Removal of inhibitory substances from human fecal specimens for detection of group A rotaviruses by reverse transcriptase and polymerase chain reactions
Authors: Wilde J , Eiden J , Yolken R
Published: January 2020

A method was developed for the purification of rotavirus RNA from fecal extracts in order to permit the sensitive identification of group A rotavirus in fecal specimens by the polymerase chain reaction. Sequential reactions with reverse transcriptase and Taq polymerase with directed primers from rotavirus gene 6 yielded characteristic 259-base-pair fragments that were then visualized by silver stain on a polyacrylamide gel. As few as 500 genomic copies of purified rotavirus RNA could be detected in this manner. However, when the method was applied to fecal samples with added rotavirus virions, inhibition was noted in many of the fecal extracts which were tested. The inhibition could be reversed by dilution of the fecal extract, but sensitivity was also reduced by a corresponding dilutional factor. The inhibition was quantitatively removed by an added step in the extraction process that utilized chromatographic cellulose fiber powder (CF11 powder) to purify the rotavirus RNA during a series of rapid washing and elution steps. After CF11 purification, rotavirus RNA could be detected in experimental fecal samples at dilutions 1,000- to 10,000-fold beyond the detection limits of standard techniques such as enzyme immunoassay and the direct visualization of RNA following polyacrylamide gel electrophoresis. Furthermore, following purification by CF11, rotavirus RNA could be detected in all of seven enzyme-linked immunosorbent assay-positive fecal samples obtained from a child with rotavirus gastroenteritis; when CF11 purification was not performed, rotavirus RNA could be detected in only four of these samples, even after the removal of inhibitors by dilution of the extracts. Large-scale identification of rotavirus in fecal specimens may be possible by use of CF11 purification of viral RNA prior to sequential reactions with reverse transcriptase and Taq polymerase in a modified polymerase chain reaction.