bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular Analysis of Riboflavin Synthesis Genes in
Bartonella henselae
and Use of the
ribC
Gene for Differentiation of
Bartonella
Species by PCR
Authors: Bereswill Stefan , Hinkelmann Silke , Kist Manfred , Sander Anna 
Published: December 2019

ABSTRACT

The biosynthesis pathway for riboflavin (vitamin B
2
), the precursor of the essential cofactors flavin mononucleotide and flavin adenine dinucleotide, is present in bacteria and plants but is absent in vertebrates. Due to their conservation in bacterial species and their absence in humans, the riboflavin synthesis genes should be well suited either for detection of bacterial DNA in human specimens or for the differentiation of pathogenic bacteria by molecular techniques. A DNA fragment carrying the genes
ribD
,
ribC
, and
ribE
, which encode homologues of riboflavin deaminase (RibD) and subunits of riboflavin synthetase (RibC and RibE), respectively, was isolated from a plasmid-based DNA library of the human pathogen
Bartonella henselae
by complementation of a
ribC
mutation in
Escherichia coli
. Sequence analysis of the
ribC
gene region in strains of
B. henselae
, which were previously shown to be genetically different, revealed that the
ribC
gene is highly conserved at the species level. PCR amplification with primers derived from the
ribC
locus of
B. henselae
was used to isolate the corresponding DNA regions in
B. bacilliformis
,
B. clarridgeiae
, and
B. quintana
. Sequence analysis indicated that the riboflavin synthesis genes are conserved and show the same operon-like genetic organization in all four
Bartonella
species. Primer oligonucleotides designed on the basis of localized differences within the
ribC
DNA region were successfully used to develop species-specific PCR assays for the differentiation of
B. henselae
,
B. clarridgeiae
,
B. quintana
, and
B. bacilliformis
. The results obtained indicate that the riboflavin synthesis genes are excellent targets for PCR-directed differentiation of these emerging pathogens. The PCR assays developed should increase our diagnostic potential to differentiate
Bartonella
species, especially
B. henselae
and the newly recognized species
B. clarridgeiae.
Evaluation of the Tuberculin Gamma Interferon Assay: Potential To Replace the Mantoux Skin Test
Authors: Pottumarthy Sudha , Morris Arthur J. , Harrison Adrian C. , Wells Virginia C. 
Published: December 2019

ABSTRACT

We evaluated an in vitro test of cell-mediated immunity, the tuberculin gamma interferon assay,
Quanti
FERON-TB (QIFN), in 455 individuals from three groups: group I, 237 immigrants from high-risk countries; group II, 127 health care workers undergoing Mantoux testing; group III, 91 patients being investigated for possible active tuberculosis (79 patients) or
Mycobacterium avium-Mycobacterium intracellulare
complex infection (12 patients). The QIFN results were compared either to those of the Mantoux test or to microbiological and clinical diagnosis, as appropriate. In each group the correlation between the diameter of induration for the skin test and the magnitude of QIFN response was significant and of moderate strength (Spearman’s rank correlation coefficient; ρ = 0.59 to 0.61;
P
< 0.001). For group I, the agreement between QIFN and Mantoux results was 89% for Mantoux-negative and 64% for Mantoux-positive individuals. For group II, when ≥10-mm-diameter induration was taken as positive, the agreement was 81% for Mantoux-negative and 67% for Mantoux-positive individuals. For group III, agreement was 81% for Mantoux-negative and 86% for Mantoux-positive patients. For patients being evaluated for active tuberculosis, the performance of the Mantoux test was not statistically different from that of the QIFN assay. In patients with active tuberculosis, the assay had a sensitivity of 77%, not significantly higher for extrapulmonary than pulmonary cases (83% versus 74%). QIFN sensitivity was not significantly different for smear-negative or smear-positive cases (80% versus 71%). The QIFN assay is a potential replacement for the Mantoux test. The acceptability of these performance values and those of similar evaluations will determine the place this test will have in detecting evidence of mycobacterial infection.
Clinical Comparison of the
Treponema pallidum
CAPTIA Syphilis-G Enzyme Immunoassay with the Fluorescent Treponemal Antibody Absorption Immunoglobulin G Assay for Syphilis Testing
Authors: Halling V. W. , Jones M. F. , Bestrom J. E. , Wold A. D. , Rosenblatt J. E. , Smith T. F. , Cockerill F. R. 
Published: December 2019

ABSTRACT

Recently, a treponema-specific immunoglobulin G (IgG) enzyme immunoassay (EIA), the CAPTIA Syphilis-G (Trinity Biotech, Jamestown, N.Y.), has become available as a diagnostic test for syphilis. A total of 89 stored sera previously tested by the fluorescent treponemal antibody absorption (FTA-ABS) IgG assay were evaluated by the CAPTIA EIA. The FTA-ABS IgG procedure was performed by technologists unblinded to results of rapid plasmid reagin (RPR) testing of the same specimens. Borderline CAPTIA-positive samples (antibody indices of ≥0.650 and ≤0.900) were retested; if the second analysis produced an index of >0.900, the sample was considered positive. Thirteen of 89 (15%) samples had discrepant results. Compared to the FTA-ABS assay, the CAPTIA EIA had a sensitivity and specificity and positive and negative predictive values of 70.7, 97.9, 96.7, and 79.7%, respectively. In another analysis, discrepancies between results were resolved by repeated FTA-ABS testing (technologists were blinded to previous RPR results) and patient chart reviews. Seven CAPTIA-negative samples which were previously interpreted (unblinded) as minimally reactive by the FTA method were subsequently interpreted (blinded) as nonreactive. One other discrepant sample (CAPTIA negative and FTA-ABS positive [at an intensity of 3+], unblinded) was FTA negative with repeated testing (blinded). For the five remaining discrepant samples, chart reviews indicated that one patient (CAPTIA negative and FTA-ABS positive [minimally reactive], blinded) had possible syphilis. These five samples were also evaluated and found to be negative by another treponema-specific test, the
Treponema pallidum
microhemagglutination assay. Therefore, after repeated testing and chart reviews, 2 of the 89 (2%) samples had discrepant results; the adjusted sensitivity, specificity, and positive and negative predictive values were 96.7, 98.3, 96.7, and 98.3%, respectively. This study demonstrates that the CAPTIA IgG EIA is a reliable method for syphilis testing and that personnel performing tests which require subjective interpretation, like the FTA-ABS test, may be biased by RPR test results.
Slime Production and Expression of the Slime-Associated Antigen by Staphylococcal Clinical Isolates
Authors: Ammendolia M. G. , Di Rosa R. , Montanaro L. , Arciola C. R. , Baldassarri L. 
Published: December 2019

ABSTRACT

The ability to produce slime and to express a slime-associated antigen was examined in a collection of staphylococcal clinical isolates. Slime-producing strains were found among coagulase-negative staphylococci in percentages comparable to those reported in other studies; surprisingly, a high percentage of
Staphylococcus aureus
strains also were able to produce this extracellular material. In the latter case, this ability was strongly dependent on the presence of an additional carbohydrate source in the growth medium. Expression of the slime-associated antigen appeared to be species specific and confined to the
Staphylococcus epidermidis
sensu stricto isolates; its strong association with the ability of these strains to produce thicker biofilms indicated slime-associated antigen as a possible virulence marker for
S. epidermidis
.
Ciprofloxacin Resistance in
Campylobacter jejuni
Isolates: Detection of
gyrA
Resistance Mutations by Mismatch Amplification Mutation Assay PCR and DNA Sequence Analysis
Authors: Zirnstein Gerald , Li Yu , Swaminathan Bala , Angulo Frederick 
Published: December 2019

ABSTRACT

The
gyrA
quinolone resistance determining region was sequenced from 13 ciprofloxacin-resistant and 20 ciprofloxacin-susceptible
Campylobacter jejuni
isolates. All isolates resistant to ciprofloxacin had Thr-86-to-Ile mutations, a mutation frequently associated with the acquisition of resistance to fluoroquinolones. A mismatch amplification mutation assay (MAMA) PCR protocol was developed that detects this
gyrA
mutation in quinolone-resistant isolates. The MAMA PCR provides a means for routine detection of the
gyrA
mutation without the need for sequencing the
gyrA
gene.
Detection of Prosthetic Hip Infection at Revision Arthroplasty by Immunofluorescence Microscopy and PCR Amplification of the Bacterial 16S rRNA Gene
Authors: Tunney Michael M. , Patrick Sheila , Curran Martin D. , Ramage Gordon , Hanna Donna , Nixon James R. , Gorman Sean P. , Davis Richard I. , Anderson Neil 
Published: December 2019

ABSTRACT

In this study the detection rates of bacterial infection of hip prostheses by culture and nonculture methods were compared for 120 patients with total hip revision surgery. By use of strict anaerobic bacteriological practice during the processing of samples and without enrichment, the incidence of infection by culture of material dislodged from retrieved prostheses after ultrasonication (sonicate) was 22%. Bacteria were observed by immunofluorescence microscopy in 63% of sonicate samples with a monoclonal antibody specific for
Propionibacterium acnes
and polyclonal antiserum specific for
Staphylococcus
spp. The bacteria were present either as single cells or in aggregates of up to 300 bacterial cells. These aggregates were not observed without sonication to dislodge the biofilm. Bacteria were observed in all of the culture-positive samples, and in some cases in which only one type of bacterium was identified by culture, both coccoid and coryneform bacteria were observed by immunofluorescence microscopy. Bacteria from skin-flake contamination were readily distinguishable from infecting bacteria by immunofluorescence microscopy. Examination of skin scrapings did not reveal large aggregates of bacteria but did reveal skin cells. These were not observed in the sonicates. Bacterial DNA was detected in 72% of sonicate samples by PCR amplification of a region of the bacterial 16S rRNA gene with universal primers. All of the culture-positive samples were also positive for bacterial DNA. Evidence of high-level infiltration either of neutrophils or of lymphocytes or macrophages into associated tissue was observed in 73% of patients. Our results indicate that the incidence of prosthetic joint infection is grossly underestimated by current culture detection methods. It is therefore imperative that current clinical practice with regard to the detection and subsequent treatment of prosthetic joint infection be reassessed in the light of these results.
Comparison of Agar Dilution, Microdilution, E-Test, and Disk Diffusion Methods for Testing Activity of Cefditoren against
Streptococcus pneumoniae
Authors: Kelly Linda M. , Jacobs Michael R. , Appelbaum Peter C. 
Published: December 2019

ABSTRACT

This study evaluated the susceptibility of pneumococci to cefditoren by agar dilution and microdilution methods (both in air) and by E-test (AB Biodisk, Solna, Sweden) and disk diffusion methods (both in CO
2
). By the three MIC tests, the MICs at which 50 and 90% of isolates were inhibited (MIC
50
s and MIC
90
s) were, respectively, as follows (in micrograms per milliliter): for the 65 penicillin-susceptible strains tested, 0.016 and 0.03 (by agar dilution), 0.016 and 0.03 (by microdilution), and 0.016 and 0.03 (by E test); for the 68 penicillin-intermediate strains tested, 0.125 and 0.5 (by agar dilution), 0.125 and 0.5 (by microdilution), and 0.25 and 0.5 (by E test); and for the 67 penicillin-resistant strains tested, 1.0 and 1.0 (by agar dilution), 0.5 and 1.0 (by microdilution), and 1.0 and 1.0 (by E test). With tentative cefditoren breakpoints (in micrograms per milliliter) of ≤2.0 (susceptible), 4.0 (intermediate), and ≥8.0 (resistant), all strains were susceptible to cefditoren by agar, microdilution, and E-test results; with breakpoints of ≤1.0, 2.0, and ≥4.0 μg/ml, 97% of strains were cefditoren susceptible by agar dilution results, 98% were susceptible by microdilution results, and 99% were susceptible by E-test results. When microdilution and E-test results were compared to those from the reference agar dilution method, 191 (95.5%) and 183 (91.5%) of strains gave essential agreement (±1 log
2
dilution); 8 (2.7%) minor discrepancies were found for both methods with a breakpoint of ≤1.0 μg/ml, and no discrepancies were found with a breakpoint of ≤2.0 μg/ml. Disk test results (breakpoint, ≤1.0 μg/ml) produced 2 major and 30 minor errors, with corresponding zone diameters (in millimeters) of ≥20 (susceptible), 17 to 19 (intermediate), and ≤16 (resistant); a ≤2.0-μg/ml breakpoint yielded zone diameters of ≥16 mm (susceptible). All three methods for testing the MIC of cefditoren showed excellent correlation.
Is There an Ecological Relationship between Rates of Antibiotic Resistance of Species of the Genus
Streptococcus
?
Authors: Gómez-Lus Rafael , Granizo Juan J. , Aguilar Lorenzo , Bouza Emilio , Gutierrez Avelino , García-de-Lomas Juan ,   the Spanish Surveillance Group for Respiratory Pathogens
Published: December 2019

ABSTRACT

The relationship between resistance to antibiotics on the part of
Streptococcus pneumoniae
and
Streptococcus pyogenes
was studied by comparing different prevalences of resistance among hospitals obtained from a recent microbiological surveillance of community-acquired respiratory tract infections. A high correlation for erythromycin resistance was found between
S. pneumoniae
isolates from lower respiratory tract infections and
S. pyogenes
isolates collected from pharyngeal swabs.
Elimination of Bacterial DNA from
Taq
DNA Polymerases by Restriction Endonuclease Digestion
Authors: Carroll Nora M. , Adamson Peter , Okhravi Narciss 
Published: December 2019

ABSTRACT

The incidence of false positives due to the presence of bacterial DNA in
Taq
DNA polymerase is an obstacle to the use of PCR in the diagnosis of infection. We describe a method that uses a restriction enzyme to destroy the ability of contaminating sequences to act as templates for a nested PCR which uses primers based on the 16S rRNA genes. The method was used prior to a PCR that amplified 10 fg of bacterial DNA. This method can be readily adapted to suit other sensitive PCRs required for clinical applications.
Prevalence of
Borrelia burgdorferi
and Granulocytic and Monocytic Ehrlichiae in
Ixodes ricinus
Ticks from Southern Germany
Authors: Baumgarten Birgit U. , Röllinghoff Martin , Bogdan Christian 
Published: December 2019

ABSTRACT

A total of 287 adult
Ixodes ricinus
ticks, collected in two regions of southern Germany (Frankonia and Baden-Württemberg) where
Borrelia burgdorferi
infections are known to be endemic, were examined for the presence of 16S ribosomal DNA specific for the
Ehrlichia phagocytophila
genogroup,
E. chaffeensis
,
E. canis
, and
B. burgdorferi
by nested PCR. Totals of 2.2% (6 of 275) and 21.8% (65 of 275) of the ticks were positive for the
E. phagocytophila
genogroup and
B. burgdorferi
, respectively. Two ticks (0.7%) were coinfected with both bacteria. Of 12 engorged
I. ricinus
ticks collected from two deer, 8 (67%) were positive for the
E. phagocytophila
genogroup and one (8%) was positive for
B. burgdorferi
. There was no evidence of infection with
E. canis
or
E. chaffeensis
in the investigated tick population. The nucleotide sequences of the 546-bp
Ehrlichia
PCR products differed at one or two positions from the original sequence of the human granulocytic ehrlichiosis (HGE) agent (S.-M. Chen, J. S. Dumler, J. S. Bakken, and D. H. Walker, J. Clin. Microbiol. 32:589–595, 1994). Three groups of sequence variants were detected; two of these were known to occur in other areas in Europe or the United States, whereas one has not been reported before. Thus, in the German
I. ricinus
tick population closely related granulocytic ehrlichiae are prevalent, which might represent variants of
E. phagocytophila
or the HGE agent.
Value of PCR for Detection of
Toxoplasma gondii
in Aqueous Humor and Blood Samples from Immunocompetent Patients with Ocular Toxoplasmosis
Authors: Bou Germán , Figueroa Marta S. , Martí-Belda Paloma , Navas Enrique , Guerrero Antonio 
Published: December 2019

ABSTRACT

Toxoplasma gondii
infection is an important cause of chorioretinitis in the United States and Europe. Most cases of
Toxoplasma
chorioretinitis result from congenital infection. Patients are often asymptomatic during life, with a peak incidence of symptomatic illness in the second and third decades of life. Diagnosis is mainly supported by ophthalmological examination and a good response to installed therapy. However, establishment of a diagnosis by ophthalmological examination alone can be difficult in some cases. To determine the diagnostic value of PCR for the detection of
T. gondii
, 56 blood and 56 aqueous humor samples from 56 immunocompetent patients were examined. Fifteen patients with a diagnosis of ocular toxoplasmosis had increased serum anti-
T. gondii
immunoglobulin G levels but were negative for anti-
T. gondii
immunoglobulin M (group 1), and 41 patients were used as controls (group 2). Samples were taken before antiparasitic therapy was initiated, and only one blood sample and one aqueous humor sample were obtained for each patient. Single nested PCRs and Southern blot hybridization were performed with DNA extracted from these samples. The results obtained showed sensitivity and specificity values of 53.3 and 83%, respectively. Interestingly, among all patients with ocular toxoplasmosis, a positive PCR result with the aqueous humor sample was accompanied by a positive PCR result with the blood sample. This result suggests that ocular toxoplasmosis should not be considered a local event, as PCR testing of blood samples from patients with ocular toxoplasmosis yielded the same result as PCR testing of aqueous humor samples. PCR testing may be useful for discriminating between ocular toxoplasmosis and other ocular diseases, and also can avoid the problems associated with ocular puncture.
Biotyping and Virulence Properties of Skin Isolates of
Candida parapsilosis
Authors: De Bernardis Flavia , Mondello Francesca , San Millàn Rosario , Pontòn Josè , Cassone Antonio 
Published: December 2019

ABSTRACT

The biotype and virulence of skin isolates of
Candida parapsilosis
were compared with blood isolates of the same fungus. Morphotype, resistotype, and electrophoretic karyotype determinations did not reveal any special cluster with a unique or dominant pathogenic feature among all of the isolates, regardless of their source. However, all cutaneous isolates had uniformly elevated secretory aspartyl-protease (Sap) activity, more than four times higher than the enzyme activity of the blood isolates. They were also highly vaginopathic in a rat vaginitis model, being significantly more virulent than blood isolates in this infection model. In contrast, skin isolates were nonpathogenic in systemic infection of cyclophosphamide-immunodepressed mice, while some blood isolates were, in this model, highly pathogenic (median survival time, 2 days, with internal organ invasion at autopsy). Finally, skin isolates did not differ, as a whole, from blood isolates in their adherence to plastic. This property was associated with a morphotype, as defined by a colony with continuous fringe, which was present among both skin and blood isolates. While confirming the genetic heterogenicity of
C. parapsilosis
, our data strongly suggest that the potential of this fungus to cause mucosal disease is associated with Sap production and is substantially distinct from that of systemic invasion.
Detection of Specific Immunoglobulin E during Maternal, Fetal, and Congenital Toxoplasmosis
Authors: Villena I. , Aubert D. , Brodard V. , Quereux C. , Leroux B. , Dupouy D. , Remy G. , Foudrinier F. , Chemla C. , Gomez-Marin J. E. , Pinon J. M. 
Published: December 2019

ABSTRACT

Toxoplasma immunoglobulin E (IgE) antibodies in 664 serum samples were evaluated by using an immunocapture method with a suspension of tachyzoites prepared in the laboratory in order to evaluate its usefulness in the diagnosis of acute
Toxoplasma gondii
infection during pregnancy, congenital infection, and progressive toxoplasmosis. IgE antibodies were never detected in sera from seronegative women, from patients with chronic toxoplasma infection, or from infants without congenital toxoplasmosis. In contrast, they were detected in 86.6% of patients with toxoplasmic seroconversion, and compared with IgA and IgM, the short kinetics of IgE was useful to date the infection precisely. For the diagnosis of congenital toxoplasmosis, specific IgE detected was less frequently than IgM or IgA (25 versus 67.3%), but its detection during follow-up of children may be interesting, reflecting an immunological rebound. Finally, IgE was detected early and persisted longer in progressive toxoplasmosis with cervical adenopathies, so it was also a good marker of the evolution of toxoplasma infection.
Degenerate and Nested PCR: a Highly Sensitive and Specific Method for Detection of Human Papillomavirus Infection in Cutaneous Warts
Authors: Harwood Catherine A. , Spink Patricia J. , Surentheran T. , Leigh Irene M. , de Villiers Ethel-Michele , McGregor Jane M. , Proby Charlotte M. , Breuer Judith 
Published: December 2019

ABSTRACT
The role of human papillomavirus (HPV) in anogenital carcinogenesis is firmly established, but evidence that supports a similar role in skin remains speculative. Immunosuppressed renal transplant recipients have an increased incidence of viral warts and nonmelanoma skin cancer, and the presence of HPV DNA in these lesions, especially types associated with the condition epidermodysplasia verruciformis (EV), has led to suggestions that HPV may play a pathogenic role. However, differences in the specificities and sensitivities of techniques used to detect HPV in skin have led to wide discrepancies in the spectrum of HPV types reported. We describe a degenerate nested PCR technique with the capacity to detect a broad spectrum of cutaneous, mucosal, and EV HPV types. In a series of 51 warts from 23 renal transplant recipients, this method detected HPV DNA in all lesions, representing a significant improvement over many previously published studies. Cutaneous types were found in 84.3% of warts and EV types were found in 80.4% of warts, whereas mucosal types were detected in 27.4% of warts. In addition, the method allowed codetection of two or more distinct HPV types in 94.1% of lesions. In contrast, single HPV types were detected in all but 1 of 20 warts from 15 immunocompetent individuals. In summary, we have established a highly sensitive and comprehensive degenerate PCR methodology for detection and genotyping of HPV from the skin and have demonstrated a diverse spectrum of multiple HPV types in cutaneous warts from transplant recipients. Studies designed to assess the significance of these findings to cutaneous carcinogenesis are under way.