Aspergillus PCR in Bronchoalveolar Lavage Fluid for the Diagnosis and Prognosis of Aspergillosis in Patients With Hematological and Non-hematological Conditions

Objectives: We evaluated the usefulness of an Aspergillus fumigatus quantitative PCR assay performed in bronchoalveolar lavage fluid (BAL) for the diagnosis and prognosis of both invasive and non-invasive aspergillosis.Methods: This 4-year retrospective study involved 613 at-risk patients who had either hematological disorders or other immunosuppressive conditions, notably solid organ transplants. Thirty-five patients had proven/probable aspergillosis and thirteen had chronic non-invasive aspergillosis. We compared PCR, galactomannan index and mycological analysis of BAL.Results: For invasive aspergillosis (IA), PCR performed in BAL yielded 88.6% sensitivity and 95.5% specificity. Comparatively, galactomannan index and mycological examination yielded only 56.3 and 63.6% sensitivity and 97.6 and 94.5% specificity, respectively. Considering the 13 chronic aspergillosis cases, PCR, galactomannan index and mycological examination yielded 76.9, 15.4, and 84.6% sensitivity and 92.2, 94.9, and 93% specificity, respectively. Fungal load in BAL evaluated by PCR was able to discriminate between aspergillosis and contamination, but not between invasive and non-invasive forms. Finally, fungal load was predictive of 90-day mortality, with 23.1% mortality for patients with less than 500 copies/mL versus 68.4% for patients above that cut-off (p < 0.05).Conclusion: Our results indicate that Aspergillus PCR in BAL is of particular interest for both the diagnosis and the prognosis of IA. It is likewise an interesting tool for the diagnosis of non-invasive forms

Comparaison de deux témoins d’inhibition de la PCR diagnostique

National audienc

The impact of lowering the cut-off value on the sensitivity of the Platelia Elisa IgG (Bio-Rad) test for toxoplasmosis diagnosis

Determining specific immune status against Toxoplasma gondii is essential for assessing the risk of reactivation in immunocompromised patients or defining serological monitoring and appropriate prophylactic measures during pregnancy. In France, toxoplasmosis serological screening requires systematic testing for IgM and IgG antibodies. The Platelia Toxo IgG and IgM test (Bio-Rad) is one of the most widely used tests for anti-toxoplasmic antibody detection. We performed a study on 384 sera, including 123 IgG negative (<6 IU/mL) and 261 IgG equivocal (6–9 IU/mL) sera tested with Platelia Toxo IgG and collected during routine screening at Pitié-Salpêtrière Hospital, Paris, France to determine the best-performing IgG titer cut-off value. Out of these 383 sera, 298 were IgM negative by Platelia Toxo IgM and 86 were IgM positive. All sera were also tested against Toxo IgG II LD BIO western blot test as confirmation. Our results indicated that an IgG titer cut-off value of ≥4.4 IU/mL for the Platelia Toxo IgG met the definition of positivity, a value significantly lower than that indicated by the manufacturers. In the presence of IgM antibodies, the IgG titer cut-off decreased significantly to a value ≥0.2 IU/mL. This latter cut-off also allowed adequate diagnosis of proven toxoplasmosis seroconversion in 76.7% of cases (33/43). Our findings may improve toxoplasmosis care by reducing therapeutic intervention time and eliminating the need for further serological monitoring

The impact of lowering the cut-off value on the sensitivity of the Platelia Elisa IgG (Bio-Rad) test for toxoplasmosis diagnosis

Determining specific immune status against Toxoplasma gondii is essential for assessing the risk of reactivation in immunocompromised patients or defining serological monitoring and appropriate prophylactic measures during pregnancy. In France, toxoplasmosis serological screening requires systematic testing for IgM and IgG antibodies. The Platelia Toxo IgG and IgM test (Bio-Rad) is one of the most widely used tests for anti-toxoplasmic antibody detection. We performed a study on 384 sera, including 123 IgG negative (<6 IU/mL) and 261 IgG equivocal (6–9 IU/mL) sera tested with Platelia Toxo IgG and collected during routine screening at Pitié-Salpêtrière Hospital, Paris, France to determine the best-performing IgG titer cut-off value. Out of these 383 sera, 298 were IgM negative by Platelia Toxo IgM and 86 were IgM positive. All sera were also tested against Toxo IgG II LD BIO western blot test as confirmation. Our results indicated that an IgG titer cut-off value of ≥4.4 IU/mL for the Platelia Toxo IgG met the definition of positivity, a value significantly lower than that indicated by the manufacturers. In the presence of IgM antibodies, the IgG titer cut-off decreased significantly to a value ≥0.2 IU/mL. This latter cut-off also allowed adequate diagnosis of proven toxoplasmosis seroconversion in 76.7% of cases (33/43). Our findings may improve toxoplasmosis care by reducing therapeutic intervention time and eliminating the need for further serological monitoring

Negativation of <i>Trypanosoma cruzi</i> PCR within Six Months after Treatment of a Child with Nifurtimox - Table 1

Negativation of <i>Trypanosoma cruzi</i> PCR within Six Months after Treatment of a Child with Nifurtimox - Table

Aspergillus PCR in serum for the diagnosis, follow-up and prognosis of invasive aspergillosis in neutropenic and non-neutropenic patients

International audienceWe evaluated the usefulness of a serum Aspergillus PCR assay for the diagnosis and prognosis of invasive aspergillosis in a study involving 941 patients for a total of 5,146 serum samples. Fifty-one patients had proven/probable aspergillosis. We compared galactomannan, PCR, and mycological analysis of pulmonary samples in both neutropenic and non-neutropenic patients. PCR performed in serum yielded 66.7% sensitivity, 98.7% specificity, 75.6% positive predictive value and 98.0% negative predictive value while the galactomannan index yielded 78.4% sensitivity, 87.5% specificity, 27% positive predictive value and 98.6% negative predictive value. The inclusion of PCR in the EORTC/MSG mycological criteria permitted the reclassification of nine other cases from possible to probable aspergillosis and increased sensitivity to 71.7%. Combining the galactomannan index with serum PCR increased the detection rate of invasive aspergillosis with 88.2% sensitivity. PCR was systematically negative in 16 patients with non-invasive forms of aspergillosis (namely aspergilloma and chronic aspergillosis). Remaining PCR-positive after a period of 14 to 20 days of treatment was related to poor outcome at 30 and 90 days. Our results also indicate that, unlike the determination of the galactomannan index, the initial fungal load as determined by PCR was highly predictive of 90-day mortality, with the rate of this latter being 15.8% for patients with less than 150 copies/mL vs 73.2% for patients at or above that cut-off (p<0.0001). Therefore, PCR appears to be a very powerful and interesting tool for the identification of patients with invasive aspergillosis who might benefit from more intense care

Freezing and storage at −20 °C provides adequate preservation of Toxoplasma gondii DNA for retrospective molecular analysis

Equipe EA MERSInternational audienceNucleic acid-based testing has become crucial for toxoplasmosis diagnosis. For retrospective (forensic or scientific) studies, optimal methods must be employed for DNA long-term storage. We compared Toxoplasma gondii detection before and after DNA storage using real-time PCR. No significant differences were found depending on duration or storage conditions at -20 °C or -80 °C

Multicentric Comparative Assessment of the Bio-Evolution Toxoplasma gondii Detection Kit with Eight Laboratory-Developed PCR Assays for Molecular Diagnosis of Congenital Toxoplasmosis

Pôle MERS CHU (EA)International audienceThe detection of Toxoplasma gondii in amniotic fluid is an essential tool for the prenatal diagnosis of congenital toxoplasmosis and is currently essentially based on the use of PCR. Although some consensus is emerging, this molecular diagnosis suffers from a lack of standardization and an extreme diversity of laboratory-developed methods. Commercial kits for the detection of T. gondii by PCR were recently developed and offer certain advantages; however, they must be assessed in comparison with optimized reference PCR assays. The present multicentric study aimed to compare the performances of the Bio-Evolution T. gondii detection kit and laboratory-developed PCR assays set up in eight proficient centers in France. The study compared 157 amniotic fluid samples and found concordances of 99% and 100% using 76 T. gondii-infected samples and 81 uninfected samples, respectively. Moreover, taking into account the classification of the European Research Network on Congenital Toxoplasmosis, the overall diagnostic sensitivity of all assays was identical and calculated to be 86% (54/63); specificity was 100% for all assays. Finally, the relative quantification results were in good agreement between the kit and the laboratory-developed assays. The good performances of this commercial kit are probably in part linked to the use of a number of good practices: detection in multiplicate, amplification of the repetitive DNA target rep529, and the use of an internal control for the detection of PCR inhibitors. The only drawbacks noted at the time of the study were the absence of uracil-N-glycosylase and small defects in the reliability of the production of different reagents