bluuebunny/crossref_metadata_2025 huggingface dataset:
Humoral responses against the 85A and 85B antigens of Mycobacterium bovis BCG in patients with leprosy and tuberculosis
Authors: Van Vooren J P , Drowart A , De Bruyn J , Launois P , Millan J , Delaporte E , Develoux M , Yernault J C , Huygen K
Published: January 2020

Immunoglobulin G antibodies against the 85A and 85B components of the Mycobacterium bovis BCG antigen 85 complex separated by isoelectric focusing were investigated in serum samples from 129 patients representing the major forms of leprosy, 111 tuberculous patients, and 153 healthy subjects. For both of the antigens, a higher degree of staining was observed for lepromatous leprosy patients and patients with active tuberculosis than for the other groups. Because sera from some healthy subjects recognized the 85A antigen, we suggest that antigen 85B is the most useful component of the antigen 85 complex for the serodiagnosis of the multibacillary forms of leprosy or of the active forms of tuberculosis.
Antibodies to shiga holotoxin and to two synthetic peptides of the B subunit in sera of patients with Shigella dysenteriae 1 dysentery
Authors: Levine M M , McEwen J , Losonsky G , Reymann M , Harari I , Brown J E , Taylor D N , Donohue-Rolfe A , Cohen D , Bennish M
Published: January 2020

Acute- and convalescent-phase sera from 18 Thai patients and convalescent-phase sera from two Israeli patients and one Bangladeshi patient with Shigella dysenteriae 1 (Shiga) dysentery were tested by enzyme-linked immunosorbent assay to detect antibodies that bind S. dysenteriae lipopolysaccharide (LPS), Shiga holotoxin, or two synthetic peptides representing epitopes from the B subunit of Shiga toxin. Paired sera from 24 Maryland adults with Shigella flexneri 2a or Shigella sonnei diarrhea served as negative controls. Of the 16 paired Thai serum samples tested for immunoglobulin G LPS antibody, 10 had greater than or equal to 4-fold rises (the two subjects with the highest convalescent-phase titers exhibited toxin-neutralizing activity); acute-phase specimens from four of the remaining six individuals already had elevated Shiga LPS titers in their acute specimens ranging from 1:800 to 1:12,800. Similarly, convalescent-phase sera from the two Israeli patients and the Bangladeshi patient revealed LPS titers of 1:800 to 1:3,200. In contrast, none of the Maryland volunteers with S. flexneri or S. sonnei diarrhea manifested rises in Shiga anti-LPS (P less than 0.00001 versus 10 of 16 Thai patients). Only 4 of the 18 Thai patients had significant rise in antibody to purified Shiga toxin, while one of the two Israeli patients and the one Bangladeshi patient had elevated convalescent-phase titers. None of the sera that reacted with Shiga holotoxin had antibody that bound to the peptides. This report, which describes a search for serum antibodies that bind Shiga toxin in patients with Shiga dysentery, demonstrates such antibodies in only a minority of patients with bacteriologically confirmed disease. During Shiga dysentery, Shiga toxin may be elaborated in such small quantities in vivo that it fails to elicit an immune response in most patients even though it may exert biological effects. In this behavior Shiga toxin resembles tetanus toxin, another potent exotoxin that fails to elicit antitoxic responses in people who recover from clinical tetanus.
Molecular detection of sorbitol-fermenting Escherichia coli O157 in patients with hemolytic-uremic syndrome
Authors: Gunzer F , Böhm H , Rüssmann H , Bitzan M , Aleksic S , Karch H
Published: January 2020

Shiga-like toxin-producing Escherichia coli strains of serogroup O157 were identified in 26 of 104 patients with hemolytic-uremic syndrome and in 18 of 668 patients with diarrhea. All strains were identified by colony hybridization with DNA probes complementary to Shiga-like toxin I and Shiga-like toxin II gene sequences and characterized by biochemical tests and serotyping. Seventeen of these 44 patients had E. coli O157 strains which were unusual because they fermented sorbitol within 24 h of incubation and were positive for beta-glucuronidase activity. Culture filtrates of these sorbitol-fermenting strains were highly toxic to Vero cells in culture. Serological tests and DNA analysis performed by restriction endonuclease digestion of B-subunit toxin genes revealed that all 17 isolates produced Shiga-like toxin II. Although by using molecular probes we established a high frequency of sorbitol-fermenting E. coli O157 strains in the patients we examined, further studies on the prevalence of such isolates in other areas of endemic disease are clearly warranted.
Outer membrane protein subtypes of Haemophilus influenzae type b isolates causing invasive disease in Victoria, Australia, from 1988 to 1990
Authors: Clements D A , MacInnes S J , Gilbert G L
Published: January 2020

Outer membrane protein subtyping of 187 isolates of Haemophilus influenzae type b (Hib), isolated from children with invasive Hib disease in Victoria, Australia, showed that a single outer membrane protein subtype (1VA) was responsible for 83% of the infections. It was identical to that responsible for the majority of cases of invasive Hib disease in Europe.
Selective buffered charcoal-yeast extract medium for isolation of nocardiae from mixed cultures
Authors: Garrett M A , Holmes H T , Nolte F S
Published: January 2020

The recovery of Nocardia species from mixed cultures is facilitated by use of a selective medium. We show that buffered charcoal-yeast extract medium with polymyxin, anisomycin, and vancomycin can be used for the selective isolation of nocardiae from contaminated specimens.
Serological responses to various Coccidioides antigen preparations in a new enzyme immunoassay
Authors: Gade W , Ledman D W , Wethington R , Yi A
Published: January 2020

A new enzyme immunoassay (EIA) was used to investigate immunoglobulin M (IgM) and IgG responses to various coccidioidal antigen preparations. Direct binding and inhibition assays both revealed that the IgG responses of many serum specimens were directed largely to the tube precipitin (TP) antigen even when the microwells were coated with a combination of coccidioidal antigens (CCAs). From a panel of 28 serum specimens containing antibodies to CCAs, 4 serum specimens yielded high IgG signals (absorbance, greater than 1.0) and were negative for IgM (absorbance, less than 0.2), yet all four IgG responses were inhibited by at least 90% with partially purified TP antigens (2.4 micrograms/ml). The IgM and IgG fractions of several serum specimens were separated by ion-exchange chromatography and assayed by using different antigen preparations adsorbed to the microwells. The binding of both IgM and IgG peaks to the microwells coated with the CCA preparation was inhibited significantly by preincubation with TP antigens. One serum specimen (specimen 26) yielded a large IgG response (absorbance, greater than 2.0) with the CCA preparation and also bound directly to microwells coated only with TP antigens. The IgG signal (absorbance) of serum specimen 26 was reduced by 98% when it was preincubated with TP antigens prior to the assay. Significant IgG signals from several other serum specimens were observed when microwells were coated with TP antigen preparations, but they were absent when periodate-treated preparations were used. Two cerebrospinal fluid specimens yielded IgG signals with CCA-coated microwells, which were not inhibited by TP antigens, and yielded no signal with microwells coated only with TP antigens. The results are consistent with the concept that the typical serologic response to TP antigens occurs early in disease progression, but they suggest that TP antigens stimulate both IgM and IgG responses.
BioArgos: a fully automated blood culture system
Authors: Courcol R J , Duhamel M , Decoster A , Lemaire V M , Rastorgoueff M L , Ochin D , Martin G R
Published: January 2020

BioArgos (Sanofi Diagnostics Pasteur, Marnes-la-Coquette, France) is a fully automated blood culture system that detects carbon dioxide production by infrared spectroscopy through a glass bottle. This hands-off system was compared with the BACTEC NR-660 system (Becton Dickinson Diagnostic Instrument Systems, Towson, Md.). A total of 336 microorganisms belonging to 74 taxa were tested in simulated blood cultures by both systems. Experimental data showed no significant differences between the two systems. The inclusive detection times (+/- the standard deviations) were 33.2 +/- 28.7 and 35.0 +/- 30.6 h with BioArgos and BACTEC, respectively. Anaerobes were detected earlier with BioArgos, whereas detection of some organisms that need oxygen to grow was slightly delayed. In conclusion, BioArgos is as reliable and accurate as BACTEC NR-660 and shows better practicability owing to noninvasive detection, reduction of vial manipulation, and absence of daily maintenance.
Development of revised quality control limits for disk diffusion susceptibility tests of selected cephem antibiotics with Haemophilus influenzae and description of a new control strain
Authors: Jorgensen J H , Barry A L , Doern G V , Ferraro M J , Murray P R
Published: January 2020

Inconsistent quality control results in disk diffusion testing of cefaclor, cefamandole, cefonicid, and cefuroxime with Haemophilus influenzae ATCC 49247 and Haemophilus test medium (HTM) prompted a search for an alternative control strain that would provide more reliable results. A five-laboratory study was conducted to evaluate two candidate H. influenzae strains as possible alternatives to the aforementioned strain. Repetitive testing of the candidate strains and H. influenzae ATCC 49247 over several days with a total of six different lots of HTM documented consistent performance of the two candidate strains and confirmed inconsistent results for some of the antibiotics with H. influenzae ATCC 49247. In particular, certain lots of HTM failed to yield cefaclor and cefamandole zone sizes within the quality control range advocated by the National Committee for Clinical Laboratory Standards. Because of the greater consistency offered by the new strains, one was selected (now designated H. influenzae ATCC 49766) to be recommended for routine quality control testing of cefaclor, cefamandole, cefonicid, cefuroxime, and the related carbacephem loracarbef. The new control strain and zone size ranges proposed here have been approved by the National Committee for Clinical Laboratory Standards in place of the previously recommended strain and zone size limits for testing of these five cephem antibiotics.
Use of aminotransferase, hepatitis C antibody, and hepatitis C polymerase chain reaction RNA assays to establish the diagnosis of hepatitis C virus infection in a diagnostic virology laboratory
Authors: Gretch D , Lee W , Corey L
Published: January 2020

Clinical and therapeutic decisions for hepatitis C virus (HCV) infection depend on factors that include documentation of past infection as well as identification of those who might benefit from antiviral chemotherapy with systemic interferon. To evaluate the ability of a diagnostic laboratory to accurately identify such patients, we compared results obtained with serum transaminase assays, two HCV antibody assays (enzyme immunoassay [EIA] and immunoblot), and a polymerase chain reaction (PCR)-based assay for HCV RNA using a group of consecutively submitted samples within our university-based diagnostic virology laboratory and sera from a population of random blood donors. One hundred percent of specimens with R values of greater than 3.0 in the HCV EIA were positive in the confirmatory immunoblot. However, 25% of specimens with EIA R values of between 1.0 and 3.0 were not confirmed by either recombinant immunoblot assay (RIBA) or RNA PCR assay (false-positive specimens). A significant correlation (P less than 0.01) between increasing reactivity in the RIBA and positivity in the RNA PCR assay was found. The incidence of HCV viremia, as determined by the RNA PCR assay, was 73% for confirmed seropositive specimens, 33% for seropositive specimens with indeterminate RIBA results, 12% for seronegative specimens obtained from infected patients, and 2.0% for seronegative specimens obtained from uninfected blood donors. In contrast, serum transaminase testing did not correlate with the RNA PCR assay for HCV. Use of the EIA and immunoblot assay followed by RNA PCR testing will identify most patients who are viremic with HCV.
Enterohemorrhagic Escherichia coli associated with hemolytic-uremic syndrome in Chilean children
Authors: Cordovéz A , Prado V , Maggi L , Cordero J , Martinez J , Misraji A , Rios R , Soza G , Ojeda A , Levine M M
Published: January 2020

A clinicoepidemiological study was undertaken to determine if enterohemorrhagic Escherichia coli (EHEC) was associated with hemolytic-uremic syndrome (HUS) in children in Santiago, Valdivia, and Temuco, Chile. Prospective surveillance detected 20 hospitalized cases of HUS in children less than 4 years of age in these cities from March 1988 to March 1989. Each HUS patient was matched (by sex and age) with two control children (hospitalized elective-surgery patients). To detect EHEC, DNA from stool culture isolates of E. coli was detected by hybridization with biotin-labelled DNA probes specific for the EHEC virulence plasmid, Shiga-like toxin I (SLT-I) or SLT-II. Stool cultures from 6 of 20 cases (30%) and from 2 of 38 controls (5.3%) yielded EHEC (P = 0.0158). EHEC isolates from all HUS cases hybridized with the EHEC plasmid probe and with probes for SLT-I or -II (or both). The serogroups of the isolates included O157, O26, and O111. EHEC causes HUS in Chile, and the biotinylated gene probes are practical diagnostic tools for epidemiologic studies.
Evaluation of two colored latex kits, the Wellcolex Colour Salmonella Test and the Wellcolex Colour Shigella Test, for serological grouping of Salmonella and Shigella species
Authors: Bouvet P J , Jeanjean S
Published: January 2020

Two colored latex kits (the Wellcolex Colour Salmonella Test [WCT-Salmonella] and the Wellcolex Colour Shigella Test [WCT-Shigella]; Division Diagnostics, Laboratories Wellcome S.A., Paris, France), which allow identification of the most frequently encountered Salmonella serogroups and Shigella species, respectively, were evaluated. WCT-Salmonella and WCT-Shigella yielded sensitivities of 98.4 and 98%, respectively, and a specificity of 100% when they were tested on pure cultures received at a reference laboratory.
Primary isolation of human T-cell leukemia-lymphoma virus types I and II: use for confirming infection in seroindeterminate blood donors
Authors: Hjelle B , Mills R , Goldsmith C , Swenson S G , Cyrus S
Published: January 2020

We describe the use of an immunofluorescence assay and coculture to confirm human T-cell leukemia-lymphoma virus (HTLV) infection. Peripheral blood mononuclear cells from 32 of 32 seropositive donors were positive in the immunofluorescence assay, and 63% of their cocultures produced p24 antigen. Specific antibodies distinguished HTLV type I (HTLV-I) from HTLV-II. HTLV-I or HTLV-II was isolated from donors with indeterminate serologic test results.
Comparison of the MEASLESTAT M test kit with the sucrose density gradient centrifugation-hemagglutination inhibition method for detection of measles virus-specific immunoglobulin M
Authors: Withers G S , Sweimler W I , Kleger B
Published: January 2020

A new commercial kit, MEASLESTAT M (Whittaker Bioproducts, Inc.), was compared with the sucrose density gradient centrifugation-hemagglutination inhibition method for the detection of measles virus-specific immunoglobulin M. Overall agreement between the two procedures was 97.1% for 104 single and paired serum samples tested. The sensitivity and specificity of MEASLESTAT M were 98.4 and 95.2%, respectively.
Clamped homogeneous electric field gel electrophoresis typing of Torulopsis glabrata isolates causing nosocomial infections
Authors: Khattak M N , Burnie J P , Matthews R C , Oppenheim B A
Published: January 2020

Thirty isolates of Torulopsis glabrata were examined by pulsed-field gel electrophoresis, which resolved 13 DNA pieces, allowing the identification of 12 types. Bands at 1,400, 1,200, 1,070, 1,025, 681, and 500 kbp were conserved. When applied to 18 isolates from an outbreak, 10 distinct types were identified by this technique. Seven patients had isolates which were identical.
Comparison of indirect immunofluorescence and membrane fluorescence assays for the differentiation of antibodies to human immunodeficiency virus types 1 and 2
Authors: Gallo D , Hoffman M N , Yeh E T , George J R , Hanson C V
Published: January 2020

Serum samples from 20 human immunodeficiency virus type 1 (HIV-1)- and 30 HIV-2-infected and 7 dually infected individuals were reacted by using the indirect immunofluorescence assay (IFA) and membrane fluorescence assay in order to determine whether these methods were useful for typing HIV-1 and HIV-2 antibodies. Although 41 of 50 (82%) of the HIV-1- and HIV-2-positive specimens cross-reacted to some extent with the heterologous antigen in the IFA, the antigen with the higher titer correlated completely with the infecting type. The IFA could not distinguish single from dual infections, however. In contrast, only 4 of the 50 (8%) serum samples cross-reacted in the membrane fluorescence test. All seven of the specimens from patients with mixed infections reacted with both antigens. The membrane fluorescence test appears to be reliable for serodifferentiation of HIV-1 and HIV-2 infections and may be useful for laboratories with low-volume typing requirements.
Effect of interstrain variation on diagnostic DNA amplification of the cytomegalovirus major immediate-early gene region
Authors: Chou S
Published: January 2020

The immediate-early region exon 4 sequences of six clinical cytomegalovirus strains were determined and compared with those of laboratory strains AD169 and Towne. Of 407 codons in exon 4, 33 (8.1%) showed interstrain variation at the peptide level and 74 (18%) showed interstrain variation at the nucleotide level. Variation occurred sporadically throughout the exon, and no grouping of strains was apparent. Published oligonucleotide primers proposed for diagnostic detection of cytomegalovirus by polymerase chain reaction have often been based on exon 4 sequences. Some of these primers show sequence mismatches with strains sequenced here. Amplification sensitivity for mismatched strains was reduced up to 100-fold. More-uniform detection sensitivity was achieved with primers of conserved sequence.
Evidence that Lo's mycoplasma (Mycoplasma fermentans incognitus) is not a unique strain among Mycoplasma fermentans strains
Authors: Sasaki T , Sasaki Y , Kita M , Suzuki K , Watanabe H , Honda M
Published: January 2020

Mycoplasma fermentans incognitus has attracted much interest either as a cofactor for the progression of AIDS or as a pathogenic agent in non-AIDS-related diseases (S.-C. Lo, M. S. Dawson, P. B. Newton III, M. A. Sonoda, J. W.-K. Shih, W. F. Engler, R. Y.-H. Wang, and D. J. Wear, Am. J. Trop. Med. Hyg. 41:364-376, 1989; S.-C. Lo, M. S. Dawson, M. Wong, P. B. Newton III, M. A. Sonoda, W. F. Engler, R.Y.-H Wang, J. W.-K. Shih, H. J. Alter, and D. J. Wear, Am. J. Trop. Med. Hyg. 41:601-616, 1989; S.-C. Lo, J.W.-K. Shih, N.-Y. Yang, C.-Y. Ou, and R. Y.-H. Wang, Am. J. Trop. Med. Hyg. 40:213-226, 1989). In the present study, the genetic and serologic properties of the incognitus strain and other M. fermentans strains were compared. Furthermore, the replication of human immunodeficiency virus type 1 (HIV-1), determined by reverse transcriptase activity and HIV-1 p24 antigen level, in peripheral blood mononuclear cells was evaluated after stimulation with mycoplasma cell lysates. The psb-2.2 viruslike infectious agent DNA probe, used to identify the incognitus strain in the tissues of AIDS and non-AIDS patients by Lo et al. (Am. J. Trop. Med. Hyg. 41:364-376 and 40:213-226, 1989), showed reaction patterns similar to those of two M. fermentans strains isolated from cell cultures but not to that of the type strain PG18. Restriction enzyme patterns of the incognitus strain with EcoRI and HindIII were also similar to those of M. fermentans strains isolated from cell cultures. There were no remarkable differences in the immunoblot profiles between the incognitus strain and the other M. fermentans strains. These results suggest that the incognitus strain is not a unique strain among M. fermentans strains. Further, cell lysates of each M. fermentans strain could also enhance the replication of HIV-1 to a level similar to that of the incognitus strain as determined by the reverse transcriptase activity and the amount of the p24 antigen.
Surface immunofluorescence assay for diagnosis of Lyme disease
Authors: Cevenini R , Sambri V , Massaria F , Franchini R , D'Antuono A , Borda G , Negosanti M
Published: January 2020

A surface immunofluorescence assay (SIFA) was analyzed and compared with a conventional indirect immunofluorescence assay (IFA) and whole-cell enzyme-linked immunosorbent assay (ELISA) for detecting immunoglobulin G (IgG) antibodies to Borrelia burgdorferi in sera from patients with Lyme disease. Fifty-five patients with syphilis and 33 patients with rheumatoid arthritis were used as disease controls. The sensitivity of the SIFA was low during the acute phase of Lyme disease (sera from seven of nine patients presenting with erythema chronicum migrans were negative during the first 2 months of illness); later, seroconversion was observed in all patients at various times during convalescence. Sera from five patients with complicated Lyme disease were strongly positive. SIFA was found to be highly specific, since sera from all patients with secondary or latent syphilis and patients with rheumatoid arthritis did not react in the test. Strong cross-reactivity occurred when these sera were tested in conventional IFA and ELISA; sera from 38 (69%) patients with syphilis were positive by IFA and sera from 51 (93%) patients were positive by ELISA, whereas 7 (21%) and 12 (36%) of the serum samples from patients with rheumatoid arthritis were positive by IFA and ELISA, respectively. Immunoblot analysis of SIFA-positive sera showed that the 31- and 34-kDa outer surface proteins (proteins A and B, respectively) of B. burgdorferi were the major reactive antigens involved in the test. The results support a role for SIFA in the investigation of complicated Lyme disease as well as in the differentiation of Lyme disease from other diseases associated with B. burgdorferi cross-reactive antibodies.
Isolation of Kingella kingae from a corneal ulcer
Authors: Mollee T , Kelly P , Tilse M
Published: January 2020

Kingella kingae was isolated from a corneal ulcer in an 11-month-old male. This organism has been previously reported from normally sterile sites, including sites of endophthalmitis, but this is the first time isolation from the cornea has been reported.
Clinical and bacteriological study of nosocomial infections due to Enterobacter aerogenes resistant to imipenem
Authors: de Champs C , Henquell C , Guelon D , Sirot D , Gazuy N , Sirot J
Published: January 2020

Enterobacter aerogenes strains resistant to imipenem were isolated in 10 patients, 7 of whom had received imipenem-cilastatin. The strains were differentiated by biotype, antibiotype, and plasmid content. All of the strains overproduced a chromosomal cephalosporinase and lost a major outer membrane protein with a size of about 40 kDa. In 5 of the 10 patients, E. aerogenes strains resistant to extended-spectrum cephalosporin were isolated during the same stay. In three patients, the similarity between the imipenem-susceptible and -resistant strains suggests the occurrence of mutation and reversion in vivo. The combination imipenem-cilastatin has been critically important for use with multiresistant strains of Enterobacter spp., but its use increases the risk of selection of imipenem-resistant strains.
Growth and phenotypic characterization of Legionella species on semisolid media made with washed agar
Authors: Rogers J E , Jones G W , Engleberg N C
Published: January 2020

Legionella pneumophila (and 28 Legionella species) grew efficiently on charcoal-free, buffered yeast extract medium made with washed agar and without apparent loss of infectivity for U937 cells. Because charcoal-free, buffered yeast extract is transparent, it is a suitable base for indicator media and pigment detection. In standard media, charcoal apparently prevents agar contaminants from inhibiting Legionella growth.
Growth of Helicobacter pylori in media containing cyclodextrins
Authors: Olivieri R , Bugnoli M , Armellini D , Bianciardi S , Rappuoli R , Bayeli P F , Abate L , Esposito E , de Gregorio L , Aziz J
Published: January 2020

We show that solid and liquid media, supplemented only with cyclodextrins and free of blood and its derivatives, support the growth of Helicobacter pylori. These media can be used for primary isolation of the bacteria from biopsy samples, routine laboratory growth, and large-scale industrial fermentation.
Comparison of two antigen assays for rapid intrapartum detection of vaginal group B streptococcal colonization
Authors: Green M , Dashefsky B , Wald E R , Laifer S , Harger J , Guthrie R
Published: January 2020

As part of a clinical investigation evaluating the efficacy of intrapartum antigen detection for screening for heavy vaginal colonization with group B streptococci (GBS), we compared the performance of modified Bactigen and Directigen GBS latex particle agglutination (LPA) kits. Paired vaginal swabs obtained from women in labor were rapidly transported to the laboratory and used for culturing (both swabs) and LPA testing (one swab by each method). GBS growth was estimated semiquantitatively and further designated as light or heavy growth. Performance specifications for each method were determined by comparing LPA and culture results from the same swab. A total of 4,251 paired swabs were evaluated during the study period. The performance specifications for detecting GBS growth of any degree for Bactigen and Directigen, respectively, were as follows: sensitivity, 20 and 24%; specificity, 99 and 99%. The performance specifications for heavy colonization for Bactigen and Directigen, respectively, were as follows: sensitivity, 57 and 62%; specificity, 99 and 99%. Neither LPA kit was a sensitive indicator of vaginal colonization with GBS or neonatal infection.
Genotypic and antigenic identification of two new strains of spotted fever group rickettsiae isolated from China
Authors: Yu X , Jin Y , Fan M , Xu G , Liu Q , Raoult D
Published: January 2020

Four isolates of spotted fever group rickettsiae isolated from ticks in China were compared with all known species and strains of spotted fever group rickettsiae by immunofluorescence assay, DNA polymerase chain reaction followed by restriction endonuclease fragment length polymorphism analysis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and Western immunoblot. The Chinese isolates belonged to three types, including a novel serotype which has not been described before. One isolate obtained from tick ova of Dermacentor nuttallii in Inner Mongolia was antigenically and genotypically identical to Rickettsia sibirica. Two isolates obtained from Dermacentor sinicus collected from Beijing were identical, different from other members of spotted fever group rickettsiae but apparently closely related to R. sibirica. HA-91, a strain isolated from Hyalomma asiaticum bv. kozlovi olenew, was antigenically and genotypically unique among spotted fever group rickettsiae, and we feel that data presented here should prompt consideration of it as a new species on the basis of current rickettsial taxonomy.
Discrimination among thermophilic Campylobacter species by polymerase chain reaction amplification of 23S rRNA gene fragments
Authors: Eyers M , Chapelle S , Van Camp G , Goossens H , De Wachter R
Published: January 2020

By comparing nucleic acid sequences determined for one of the most variable areas of 23S rRNA genes of 23 Campylobacter strains, we were able to identify regions specific for thermophilic Campylobacter strains. Oligonucleotide primers corresponding to these unique regions were synthesized and used in the polymerase chain reaction. One primer pair selectively detected all thermophilic Campylobacter species, while four other primer pairs allowed discrimination among the thermophilic species Campylobacter coli, Campylobacter jejuni subsp. jejuni, Campylobacter lari, and Campylobacter upsaliensis. All primer sets were tested successfully on a large number of clinical isolates.
Rapid identification of mycobacteria to the species level by polymerase chain reaction and restriction enzyme analysis
Authors: Telenti A , Marchesi F , Balz M , Bally F , Böttger E C , Bodmer T
Published: January 2020

A method for the rapid identification of mycobacteria to the species level was developed on the basis of evaluation by the polymerase chain reaction (PCR) of the gene encoding for the 65-kDa protein. The method involves restriction enzyme analysis of PCR products obtained with primers common to all mycobacteria. Using two restriction enzymes, BstEII and HaeIII, medically relevant and other frequent laboratory isolates were differentiated to the species or subspecies level by PCR-restriction enzyme pattern analysis. PCR-restriction enzyme pattern analysis was performed on isolates (n = 330) from solid and fluid culture media, including BACTEC, or from frozen and lyophilized stocks. The procedure does not involve hybridization steps or the use of radioactivity and can be completed within 1 working day.
Evaluation of an immunoblot assay for serological confirmation and differentiation of human T-cell lymphotropic virus types I and II
Authors: Roberts B D , Foung S K , Lipka J J , Kaplan J E , Hadlock K G , Reyes G R , Chan L , Heneine W , Khabbaz R F
Published: January 2020

The confirmation of infection with human T-cell lymphotropic virus type I (HTLV-I) and type II (HTLV-II) currently involves multiple assays. These include Western blot (immunoblot) (WB) and/or radioimmunoprecipitation assay for detection of antibodies to HTLV-specific viral proteins and polymerase chain reaction and/or peptide-based enzyme immunoassays for differentiating between the two viruses. We undertook an evaluation of a modified WB assay that includes native HTLV-I viral proteins from MT-2 cells spiked with an HTLV-I transmembrane glycoprotein (recombinant p21e) and the HTLV-I- and HTLV-II-specific recombinant proteins MTA-1 and K55. The test panel consisted of well-characterized sera from U.S. blood donors, American Indians, intravenous drug users, and patients seen in sexually transmitted disease clinics. Of 158 HTLV-I/II-seropositive serum specimens tested, 156 (98.7%) were confirmed and typed as HTLV-I or HTLV-II. Of 82 HTLV-I/II-seroindeterminate or -seronegative serum specimens, only 1 was classified as HTLV-II positive: the sample had weak gag p19 and strong gag p24 and env p21e reactivity and was radioimmunoprecipitation assay negative for env gp61/68 but polymerase chain reaction positive for HTLV-II. The specificity of the modified WB for confirming and typing serum samples was therefore 100%. We conclude that this WB assay is useful for confirming and typing HTLV infection and can help simplify HTLV-I/II testing algorithms.
Improved isolation of Chlamydia trachomatis from a low-prevalence population by using polyethylene glycol
Authors: Gibson J P , Egerer R M , Wiedbrauk D L
Published: January 2020

The effect of polyethylene glycol (PEG) on the isolation of Chlamydia trachomatis was evaluated in our laboratory. Initial range-finding experiments demonstrated that the number of chlamydial inclusion bodies increased with increasing PEG concentrations. However, PEG concentrations above 10.5% became progressively more toxic to the McCoy cell monolayers. When 50 frozen clinical Chlamydia isolates were inoculated onto McCoy cell cultures with and without 7% PEG, the PEG-treated cultures produced three- to fivefold more chlamydial inclusions than cultures without PEG. This enhancement was also observed when 1,144 fresh clinical specimens from a low-prevalence population were tested. With fresh clinical specimens, PEG-treated cultures produced two- to sixfold more inclusions than standard cultures. The addition of 7% PEG to the chlamydial overlay medium significantly increased the number of inclusions in each culture, improved the sensitivity of the culture, and decreased the probability of missing a weakly positive specimen.
Two-way cross-neutralization mediated by a shared P (VP4) serotype between bovine rotavirus strains with distinct G (VP7) serotypes
Authors: Matsuda Y , Isegawa Y , Woode G N , Zheng S , Kaga E , Nakagomi T , Ueda S , Nakagomi O
Published: January 2020

Among bovine rotavirus strains, there are three G serotypes (G6, G8, and G10) and three P (VP4) serotypes (PB1, PB2, and PB3, which are defined on the basis of strains NCDV, UK, and B223, respectively). Plaque reduction neutralization assays with hyperimmune antisera disclosed two-way antigenic relationships of strain KN-4 with strain KK-3 (G10, PB3) as well as with strains NCDV (G6, PB1) and 0510 (G6, PB2). Neutralization assays with monoclonal antibodies specific for G6, G10, and PB3 revealed that KK-3 and KN-4 had the same P serotype (PB3) but that neither G6- nor G10-specific monoclonal antibody neutralized KN-4. Comparison of the VP7 gene sequence of KN-4 with those of other bovine rotavirus strains indicated that KN-4 was more similar to G6 bovine strains than to KK-3 and other G10 strains, suggesting that the G serotype of KN-4 was G6. From these results, we concluded that the two-way cross-neutralization between KN-4 and NCDV or 0510 was mediated by shared G6 serotype specificity, whereas the two-way cross-neutralization between KN-4 and KK-3 was mediated by shared P serotype specificity (PB3). Thus, KN-4 and KK-3 represent the first reported example of a two-way antigenic relationship mediated only by the P serotype. This article emphasizes the need for adopting a binary serotyping system and development of reagents which will enable classification of rotaviruses based on their G and P serotype specificities.
Large DNA restriction fragment polymorphism in the Mycobacterium avium-M. intracellulare complex: a potential epidemiologic tool
Authors: Mazurek G H , Hartman S , Zhang Y , Brown B A , Hector J S , Murphy D , Wallace R J
Published: January 2020

Mycobacterium avium-M. intracellulare complex (MAI) isolates were studied by comparing the large restriction fragment (LRF) patterns produced by digesting their DNAs with infrequently cutting restriction endonucleases and separating the resultant large fragments by pulsed-field gel electrophoresis. Four reference strains and 35 randomly selected clinical MAI isolates gave highly diverse LRF patterns when their DNAs were digested with XbaI or AsnI. The LRF patterns of random isolates identified to be the same species by DNA probe analysis were not similar. The LRF patterns of random isolates of the same serotype were also different. In contrast, all isolates recovered from the same patient gave identical patterns. This included 28 isolates from nine patients. One isolate from sputum, one isolate from bone marrow, and two isolates from blood recovered over a 27-month period from a patient with AIDS were identical. Seven isolates recovered from the sputum of a second patient over 37 months also had identical patterns. The LRF patterns of unrelated MAI strains are highly polymorphic, appear to be strain specific, are relatively stable, and offer exciting promise as epidemiologic markers for the study of MAI infections.
Strain identification of Mycobacterium tuberculosis by DNA fingerprinting: recommendations for a standardized methodology
Authors: van Embden J D , Cave M D , Crawford J T , Dale J W , Eisenach K D , Gicquel B , Hermans P , Martin C , McAdam R , Shinnick T M
Published: January 2020

DNA fingerprinting of Mycobacterium tuberculosis has been shown to be a powerful epidemiologic tool. We propose a standardized technique which exploits variability in both the number and genomic position of IS6110 to generate strain-specific patterns. General use of this technique will permit comparison of results between different laboratories. Such comparisons will facilitate investigations into the international transmission of tuberculosis and may identify specific strains with unique properties such as high infectivity, virulence, or drug resistance.
Incidental finding of a microsporidian parasite from an AIDS patient
Authors: McDougall R J , Tandy M W , Boreham R E , Stenzel D J , O'Donoghue P J
Published: January 2020

Light microscopic examination of feces from a human immunodeficiency virus-positive patient with chronic diarrhea, anorexia, and lethargy revealed the presence of numerous refractile bodies resembling microsporidian spores. They were subsequently identified as belonging to the genus Nosema on the basis of their ultrastructural characteristics. However, the microsporidia were enclosed within striated muscle cells, suggesting that they were probably ingested in food; thus, this represented an incidental finding rather than a true infection.
Acridine orange staining as a replacement for subculturing of false-positive blood cultures with the BACTEC NR 660
Authors: Hunter J S
Published: January 2020

Despite the customization of growth index thresholds within individual laboratories, use of the BACTEC NR 660 automated blood culture system results in a number of false-positive cultures. The results of Gram staining, acridine orange staining, and subculturing to agar media were evaluated on 210 false-positive blood cultures over a 6-month period. Inclusion of acridine orange staining in the routine workup of false-positive blood cultures can eliminate the need for subculturing.
Genomic fingerprinting of "Haemophilus somnus" isolates by using a random-amplified polymorphic DNA assay
Authors: Myers L E , Silva S V , Procunier J D , Little P B
Published: January 2020

The random-amplified polymorphic DNA (RAPD) assay was used to generate DNA fingerprints for 16 isolates of "Haemophilus somnus," and one isolate each of "Haemophilus agni," "Histophilus ovis," "Actinobacillus seminis," Pasteurella haemolytica, and Escherichia coli. The RAPD assay differentiated among "H. somnus" isolates, which shared similarity coefficients of 0.46 to 1.00 on the basis of pairwise comparisons of RAPD markers produced with nine random decamer primers. Three virulent encephalitic "H. somnus" isolates exhibited identical banding patterns, suggesting a common clonal ancestry. The RAPD assay clearly distinguished between the "H. somnus"-"H. agni"-"H. ovis" group and the other bacterial species tested. The results of the present study suggest that DNA fingerprinting of "H. somnus" isolates by the RAPD assay could be valuable in revealing subspecific divisions within this largely unexplored species.
Detection of Mycoplasma pulmonis in experimentally infected laboratory rats by 16S rRNA amplification
Authors: van Kuppeveld F J , Melchers W J , Willemse H F , Kissing J , Galama J M , van der Logt J T
Published: January 2020

Recently, an rRNA-based polymerase chain reaction (PCR) has been developed for the detection of murine mycoplasmas at both the genus and species level (F. J. M. van Kuppeveld, J. T. M. van der Logt, A. F. Angulo, M. J. van Zoest, W. G. V. Quint, H. G. Niesters, J. M. D. Galama, and W. J. G. Melchers, Appl. Environ. Microbiol. 58:2606-2615, 1992). In this study, the diagnostic value of this PCR assay for the detection of Mycoplasma pulmonis in infected rats was studied. For this purpose, 25 Wistar rats were infected intranasally with M. pulmonis strain M72-138 and investigated for the presence of this pathogen by both in vitro isolation and PCR. Five rats were monitored longitudinally by screening of throat swabs at several time points for up to 248 days postinfection. The remaining 20 rats were killed between 3 and 87 days postinfection, and organism recovery from both throat and urogenital tract specimens was attempted. M. pulmonis could be detected in the throat for up to 248 days postinfection but not in the urogenital tract, either by culture or by PCR. PCR proved to be the optimal method for testing throat samples. All samples in which M. pulmonis was detected by culture were also positive by PCR. By PCR, M. pulmonis was also detected in 3.7% of the samples which were culture negative and in 9.9% of the samples from which cultures were overgrown with bacteria. The results of this study demonstrate the suitability of PCR for the detection of mycoplasmal infection in rodents.
Rotavirus-inhibitory activity in serial milk samples from Mexican women and rotavirus infections in their children during their first year of life
Authors: Brüssow H , Benitez O , Uribe F , Sidoti J , Rosa K , Cravioto A
Published: January 2020

A total of 75 children born in rural Mexico were followed for diarrheal diseases and rotavirus (RV) excretion during the first year of life. For 18 children, an average of 14 serial breast milk samples were obtained between days 2 and 360 after delivery and were tested for RV-inhibitory activity. Of these samples, 70, 62, and 85% showed inhibitory activity against serotype (ST) 1 human RV, ST4 human RV, and ST3 simian RV, respectively; the median titers were 10, 10, and 20, respectively. Some 89% of the milk samples showed RV-specific antibodies in an enzyme-linked immunosorbent assay (median titer, 20). Surprisingly, 98% of the milk samples inhibited ST6 bovine RV. ST6, but not ST1, RV-inhibitory activity survived heat treatment (10 min at 80 degrees C). Of the 18 children tested, 13 children experienced 23 episodes of diarrhea (enterotoxigenic [n = 8] and enteropathogenic [n = 3] Escherichia coli, Campylobacter jejuni [n = 4], Shigella flexneri [n = 2], RV [n = 1]) and 5 children experienced 6 RV infections. Only one RV infection was associated with diarrhea. The five RV excretors did not differ from the nonexcretors with respect to the RV-inhibitory activity in the breast milk fed to them. The RV-inhibitory titers were too low in the majority of the studied Mexican milk samples to indicate an important effect of breast-feeding on the take rate of oral human, simian, or reassortant RV vaccines. Breast-feeding might, however, inhibit the take rate of a bovine RV vaccine.
Comparison of polymerase chain reaction with culture and enzyme immunoassay for diagnosis of pertussis
Authors: He Q , Mertsola J , Soini H , Skurnik M , Ruuskanen O , Viljanen M K
Published: January 2020

A polymerase chain reaction (PCR) assay amplifying a segment of a repeated gene element of Bordetella pertussis was compared with culture and enzyme immunoassay (EIA) for the diagnosis of pertussis. The PCR assay was specific for B. pertussis in tests with a panel of other bacteria and with an extensive collection of specimen material from healthy persons and children with respiratory infections other than pertussis. The PCR assay was used in the analysis of 117 nasopharyngeal swabs collected from children at an elementary school at which a pertussis outbreak occurred. Fifty-six (48%) of the 117 swabs were positive, including those for all six culture-positive cases. The PCR method was then applied to analyze another pertussis outbreak. Of 40 nasopharyngeal aspirates taken from 37 clinically susceptible pertussis patients and from three asymptomatic contacts, the PCR identified 18 (45%), including all 3 culture-positive and 5 (35%) of the 14 seropositive patients. The most consistent and reliable diagnosis by positive PCR result was observed with those patients experiencing symptoms within 1 to 6 weeks of sample collection. We conclude that PCR is a rapid, sensitive, and specific means of diagnosing pertussis, especially during the first weeks of disease. The assay can be performed with both nasopharyngeal swabs and aspirates.
Endocarditis caused by Rochalimaea quintana in a patient infected with human immunodeficiency virus
Authors: Spach D H , Callis K P , Paauw D S , Houze Y B , Schoenknecht F D , Welch D F , Rosen H , Brenner D J
Published: January 2020

Rochalimaea quintana and Rochalimaea henselae are closely related, fastidious, gram-negative rickettsiae. Thus far, the spectrum of human Rochalimaea sp. infections has not included endocarditis. We describe a 50-year-old human immunodeficiency virus-positive man who developed endocarditis caused by R. quintana. DNA relatedness studies, which compared our patient's blood culture isolate with known Rochalimaea species, identified the organism as R. quintana. Our report expands the spectrum of Rochalimaea sp. infections and identifies a new infectious cause of endocarditis.
Three supplementary diagnostic tests for Campylobacter species and related organisms
Authors: Burnens A P , Nicolet J
Published: January 2020

We analyzed 307 recent clinical isolates and reference strains of Campylobacter spp., Helicobacter spp., and Arcobacter spp. for arylsulfatase and pyrazinamidase activities and susceptibility to polymyxin B. All isolates studied were identified by DNA dot hybridization prior to testing. Campylobacter concisus, C. sputorum, and Helicobacter fennelliae were positive for arylsulfatase. C. jejuni, C. coli, C. lari, C. upsaliensis, and C. concisus were positive for pyrazinamidase. Only Helicobacter spp. were resistant to polymyxin B. C. fetus subsp. fetus, C. hyointestinalis, C. mucosalis, and Arcobacter spp. were negative in all three tests. The tests were simple to perform and read, and with the exception of C. sputorum biovars, all isolates within a species gave consistently positive or negative results in all the assays. These three phenotypic tests may help refine current methods for phenotypic identification of Campylobacter spp. and related organisms in a reference laboratory setting.
Amplification of Chlamydia trachomatis DNA by ligase chain reaction
Authors: Dille B J , Butzen C C , Birkenmeyer L G
Published: January 2020

Amplification of Chlamydia trachomatis DNA by polymerase chain reaction was compared with amplification by ligase chain reaction (LCR). Both amplification procedures were able to consistently amplify amounts of DNA equivalent to three C. trachomatis elementary bodies. All 15 C. trachomatis serovars were amplified to detectable levels by LCR, and no DNA from 16 organisms potentially found in clinical specimens or from Chlamydia psittaci and Chlamydia pneumoniae was amplified by LCR.
Comparison of dot filter hybridization, Southern transfer hybridization, and polymerase chain reaction amplification for diagnosis of anal human papillomavirus infection
Authors: Kuypers J M , Critchlow C W , Gravitt P E , Vernon D A , Sayer J B , Manos M M , Kiviat N B
Published: January 2020

The detection and classification of human papillomavirus (HPV) by a consensus primer polymerase chain reaction (PCR) technique were compared with detection and classification by dot filter hybridization (DFH) and Southern transfer hybridization (STH). PCR detected HPV in 87% of specimens; the detection rates for DFH and STH were 51% and 49%, respectively. The specific HPV types detected by STH were also detected by PCR in 90% of specimens. However, 75% of the samples positive for unclassified HPV by STH were typed by PCR. PCR results were reproducible, as assessed by repeat analysis (96% agreement), by analysis of paired same-day specimens (89% agreement), and by interlaboratory analysis (88% agreement). PCR is a sensitive, specific, and reproducible test for HPV detection and classification in clinical and epidemiologic studies.