Alessandra Moroni

Evaluation of LIAISON Treponema Screen, a Novel Recombinant Antigen-Based Chemiluminescence Immunoassay for Laboratory Diagnosis of Syphilis

ABSTRACT The purpose of this study was to evaluate the diagnostic performance of LIAISON Treponema Screen (DiaSorin, Saluggia, Italy), a new automated chemiluminescence immunoassay (CLIA), in comparison with that of rapid plasma reagin (RPR) and the following currently used treponemal tests: hemagglutination test (TPHA), immunoenzymatic assay (EIA), and Western blot (WB). First, a retrospective study was performed with a panel of 2,494 blood donor sera, a panel of 131 clinical and serologically characterized syphilitic sera, and 96 samples obtained from subjects with potentially interfering diseases or conditions. A prospective study was also performed by testing 1,800 unselected samples submitted to the Microbiology Laboratory of the St. Orsola Hospital in Bologna, Italy, for routine screening for syphilis. As expected, RPR was the least specific method, especially when potentially cross-reacting sera were tested. On the contrary, all of the treponemal tests proved to be very specific (99.9%) and they performed with the following sensitivities: 100% (WB), 99.2% (CLIA), 95.4% (EIA), and 94.7% (TPHA).

Laboratory diagnosis of syphilis with automated immunoassays.

The serological detection of specific antibodies to Treponema pallidum is of particular importance in the diagnosis of syphilis. The purpose of this study was to evaluate diagnostic performances of automated immunoassays in comparison with T. pallidum hemagglutination test (TPHA) and Western Blot (WB). The retrospective study was performed with different panels of sera: 244 clinical and serological characterized syphilitic sera and 203 potentially interfering samples. All the sera were tested by Enzygnost Syphilis, ARCHITECT Syphilis TP, TPHA, and homemade WB. The diagnostic performances of the two assays were very similar: both Enzygnost Syphilis and ARCHITECT Syphilis TP performed with a sensitivity of 99.2%, whereas the specificity was 98.5 and 98.4%, respectively. Considering the suitability for automation, both immunoassays may represent a good choice as a screening test. However, the use of a confirmatory test, such as TPHA or WB, remains a must in order to avoid false‐positive results. J. Clin. Lab. Anal. 23:1–6, 2009.

Activity of Cathelicidin Peptides against Chlamydia spp

ABSTRACT The in vitro activity of six cathelicidin peptides against 25 strains of Chlamydia was investigated. SMAP-29 proved to be the most active peptide, reducing the inclusion numbers of all 10 strains of Chlamydia trachomatis tested by ≥50% at 10 μg/ml. This peptide was also active against C. pneumoniae and C. felis.

Prenatal Syphilis Infection Is A Possible Cause Of Preterm Delivery Among Immigrant Women From Eastern Europe

Objective:to evaluate the prevalence of maternal syphilis at delivery and neonatal syphilis infection in an Italian urban area, in connection with the increased flow of immigration. Study design: A prospective surveillance study was carried out in Bologna, Italy, from November 2000 to March 2006. All pregnant women were screened for syphilis at delivery. Infants born to seropositive mothers were enrolled in a prospective follow-up. Results: During the study period 19 205 women gave birth to 19 548 infants. A total of 85 women were seropositive for syphilis at delivery. The overall syphilis seroprevalence in pregnant women was 0.44%, but it was 4.3% in women from eastern Europe and 5.8% in women from Central–South America. Ten women were first found positive at delivery, as they did not receive any prenatal care. Nine of these were from eastern Europe. All their infants were asymptomatic, but six had both reactive immunoglobulin (Ig)M western blot and rapid plasma reagin tests and were considered prenatally infected. Three of six were preterm (gestational age <37 weeks). Conclusions: In Italy, congenital syphilis infection is strictly related to immigration from eastern Europe. Although it is asymptomatic, it could cause premature delivery. Therefore, it is necessary to perform serological tests during the third trimester in mothers coming from endemic areas to adequately treat syphilis in pregnancy and prevent congenital infection. If the mother’s test results are not available at delivery, it is necessary to investigate the newborn, especially if it is born prematurely.

Borrelia burgdorferi VlsE antigen for the serological diagnosis of Lyme borreliosis

In this study, raising and development of antibody response to Borrelia burgdorferi infection in 66 Italian patients suffering from culture-confirmed Lyme borreliosis erythema migrans (EM) was investigated. Sixty-two of 66 cultures obtained from biopsies were identified as B. afzelii by PCR. A total of 175 serially collected serum samples were tested by using two different sets of commercial assays: Enzygnost Lyme link VlsE/IgG and Enzygnost Borreliosis IgM (DADE Behring, Marburg, Germany) and LIAISON Borrelia IgG and IgM (Diasorin, Saluggia, Italy). Considering only samples obtained at first presentation when EM was clinically evident, 49/66 patients (72.4%) were IgG or IgM positive by Enzygnost, whereas 33/66 (50.0%) patients were IgG or IgM positive by LIAISON. Taking into account the follow-up period, eight patients sero-converted for IgG or IgM by Enzygnost and four by LIAISON. Similar and very good specificity values were obtained by all methods. Testing sera obtained from blood donors (n = 300) and from patients suffering from some of the most common biological conditions possibly resulting in false-positive reactivity in Lyme disease serology (n = 100) showed that Enzygnost Lyme link VlsE/IgG was the more specific (98.3%), followed by LIAISON Borrelia IgG (96.5%), and considering IgM tests, Enzygnost Borreliosis IgM showed to be 95.3%% specific, whereas the LIAISON Borrelia IgM was 92.8% specific. Recombinant VlsE antigens obtained from all three B.burgdorferi genospecies pathogenic to humans (included in Enzygnost Lyme link VlsE/IgG) greatly improved serodiagnosis of Lyme disease.

Evaluation of the BioPlex 2200 Syphilis System as a First-Line Method of Reverse-Sequence Screening for Syphilis Diagnosis

ABSTRACT Despite recent technological advances, the diagnosis of syphilis remains a challenging enterprise. Actually, most high-volume laboratories have adopted the “reverse algorithm” due several factors, including the potential to automate testing. Recently, immunoassays processed on random-access systems have been proposed as screening tests. The purpose of this study was to evaluate diagnostic performances of BioPlex 2200 Syphilis IgG and BioPlex 2200 Syphilis IgM, tests based on Multiplex Flow technology, in comparison with the performance of Architect Syphilis TP, a chemiluminescent immunoassay for the detection of IgG and/or IgM anti-Treponema pallidum antibodies. A retrospective study was performed with a panel of 100 blood donor sera, a panel of 350 clinical and laboratory-characterized syphilitic sera, and 170 samples obtained from subjects with potentially interfering conditions. Moreover, 200 unselected samples submitted to the Microbiology Laboratory of St. Orsola Hospital in Bologna for routine screening for syphilis were evaluated. As confirmatory tests, T. pallidum hemagglutination and Western blot assays were used. Considering the IgG Western blot (WB) assay to be the gold standard method, BioPlex 2200 Syphilis IgG specificity was far higher than Architect Syphilis TP specificity (89.7% versus 78.4%, respectively), whereas the sensitivity was 100% for both automated methods. Compared to the IgM WB assay, BioPlex 2200 Syphilis IgM performed with a specificity of 94.9%, whereas the sensitivity was 84.8%. Considering the excellent ease of use and automation, the high sample throughput and its valuable analytical performances, BioPlex Syphilis 2200 IgG could represent a suitable choice for high-volume laboratories. BioPlex Syphilis 2200 IgM could be considered a good addition to IgG testing for uncovering active infections.

Detection of Chlamydia trachomatis DNA in patients with non-gonococcal urethritis using the polymerase chain reaction.

A practical protocol using the polymerase chain reaction (PCR) was designed for detecting Chlamydia trachomatis in clinical samples. DNA was extracted from material collected on urethral swabs and used as substrate for the PCR. The target was a 600 basepair DNA segment of the multicopy plasmid that is common to all strains of the bacterium. Negative samples were checked for loss of DNA or presence of polymerase inhibitors by a second PCR, targeted to a conserved segment of the human genome. The whole procedure was tested on 216 men with non-gonococcal urethritis (NGU). All patients were independently assessed by tissue culture isolation (60 positive samples) and a commercial immunoenzymatic assay. The PCR protocol, while sufficiently simple for routine application, was reliable and, for the diagnosis of urethritis, at least as good as tissue culture isolation.

A Decrease in the Immunoglobulin G Antibody Response against the VlsE Protein of Borrelia burgdorferi Sensu Lato Correlates with the Resolution of Clinical Signs in Antibiotic-Treated Patients with Early Lyme Disease

ABSTRACT The purpose of this study was to evaluate the diagnostic performance of the LIAISON Borrelia Screen (Diasorin, Saluggia, Italy), a new automated immunoassay based on the chemiluminescent technology (chemiluminescence immunoassay). To assess whether a decrease in a negative value in the anti-VlsE immunoglobulin G (IgG) antibody titer was correlated with a positive response to treatment, a group of serially collected serum samples from 67 patients with culture-confirmed erythema migrans was retrospectively studied. All the patients had been treated with antibiotics and were free of disease within 3 to 6 months of follow-up. All the 15 patients who were found to be IgG positive at the time of enrollment and who were bled at least four times during the follow-up became IgG seronegative at 2 to 6 months posttreatment. These results indicate that a decline in the anti-VlsE antibody titer coincides with effective antimicrobial therapy in patients with early localized Lyme disease.

In Vitro Activities of Several Antimicrobial Agents against Recently Isolated and Genotyped Chlamydia trachomatis Urogenital Serovars D through K

ABSTRACT A systematic evaluation of the susceptibility of all Chlamydia trachomatis urogenital serovars (D through K) to levofloxacin, erythromycin, doxycycline, clarithromycin, and azithromycin was performed. All C. trachomatis serovars had comparable susceptibilities with respect to the various antimicrobials tested, thus confirming the homogeneous data so far obtained regarding the susceptibility of C. trachomatis to antimicrobial agents.

Serological response to chlamydial infection in sheep, studied by enzyme‐linked immunosorbent assay and immunoblotting

Two enzyme-linked immunosorbent assays (ELISA) using highly purified elementary bodies (EB) of Chlamydia psittaci A/22 strain (ovine) or 6BC strain (psittacine) were set up for the detection of antichlamydial antibodies in sheep. No significant differences were observed between the two ELISAs, whereas these tests proved to be more sensitive than complement fixation test and showed a good correlation (r = 0.75) with immunofluorescence assay. The periodate treatment of chlamydial antigens modified the results of serological responses studied by ELISA, depending on the sera. The average reduction of ELISA values by periodate was 28%, ranging between 5% and 65%. The immunoblot analysis of sheep sera showed high cross reactivity between the polypeptides of A/22 and 6BC strains. However, some differences were observed. The major outer membrane protein (MOMP) of 6BC strain was recognized at different molecular weight position (40,000 kDa) in comparison with the MOMP of A/22 strain (38,000 kDa). In addition, a clear band of 97,000 kDa was detected by all sheep sera tested with A/22 strain. This band was undetectable in the blots performed with 6BC strain.