Is It Possible to Differentiate Pneumocystis jirovecii Pneumonia and Colonization in the Immunocompromised Patients with Pneumonia?

Respiratory sample staining is a standard tool used to diagnose Pneumocystis jirovecii pneumonia (PjP). Although molecular tests are more sensitive, their interpretation can be difficult due to the potential of colonization. We aimed to validate a Pneumocystis jirovecii (Pj) real-time PCR (qPCR) assay in bronchoscopic bronchoalveolar lavage (BAL) and oropharyngeal washes (OW). We included 158 immunosuppressed patients with pneumonia, 35 lung cancer patients who underwent BAL, and 20 healthy individuals. We used a SYBR green qPCR assay to look for a 103 bp fragment of the Pj mtLSU rRNA gene in BAL and OW. We calculated the qPCR cut-off as well as the analytical and diagnostic characteristics. The qPCR was positive in 67.8% of BAL samples from the immunocompromised patients. The established cut-off for discriminating between disease and colonization was Ct 24.53 for BAL samples. In the immunosuppressed group, qPCR detected all 25 microscopy-positive PjP cases, plus three additional cases. Pj colonization in the immunocompromised group was 66.2%, while in the cancer group, colonization rates were 48%. qPCR was ineffective at diagnosing PjP in the OW samples. This new qPCR allowed for reliable diagnosis of PjP, and differentiation between PjP disease and colonization in BAL of immunocompromised patients with pneumonia.

High transient colonization by Pneumocystis jirovecii between mothers and newborn.

The aim of the study was to explore the frequency and dynamics of acquisition and colonization of Pneumocystis jirovecii among neonates, as well as the epidemiological and genotypic characteristics in mother-child binomial. In a prospective enrolled cohort of women in their third trimester of pregnancy, nasopharyngeal swabs (NPS) and clinical and epidemiological data were collected at four different times: 17 days, 2nd, 4th, and 6th month of life of the newborn. P. jirovecii was detected by nested-PCR for the mtLSU-rRNA gene in each NPS; the genotypes were determined amplifying four genes. Forty-three pairs and 301 NPS were included. During the third trimester, 16.3% of pregnant women were colonized. The rate of colonization in mothers at delivery was 16, 6, 16, and 5% and in their children 28, 43, 42, and 25%, respectively. Within pregnant women, 53% remained negative throughout follow-up, and among these, 91% of their children were positive in at least one of their samples. In both, mothers and children, the most frequent genotype of P. jirovecii was 1. CONCLUSION: The frequency of colonization by P. jirovecii was higher in newborns than in their respective progenitors. Colonization of both mothers and children is transitory; however, the mother of the newborn is not necessarily the source of primary infection. What is Known: • We did not find studies comparing P. jirovecii colonization between mothers and children simultaneously, yet the frequency of colonization by serologic and molecular methods in pregnant women has been reported. What is New: • According to our findings, 3/4 of the children had transient colonization during the first 6 months of life, in only half in the mothers, without proof of mother-to-child transmission or vice versa.

Impact on Bacterial Resistance of Therapeutically Nonequivalent Generics: The Case of Piperacillin-Tazobactam.

Previous studies have demonstrated that pharmaceutical equivalence and pharmacokinetic equivalence of generic antibiotics are necessary but not sufficient conditions to guarantee therapeutic equivalence (better called pharmacodynamic equivalence). In addition, there is scientific evidence suggesting a direct link between pharmacodynamic nonequivalence of generic vancomycin and promotion of resistance in Staphylococcus aureus. To find out if even subtle deviations from the expected pharmacodynamic behavior with respect to the innovator could favor resistance, we studied a generic product of piperacillin-tazobactam characterized by pharmaceutical and pharmacokinetic equivalence but a faulty fit of Hill's Emax sigmoid model that could be interpreted as pharmacodynamic nonequivalence. We determined the impact in vivo of this generic product on the resistance of a mixed Escherichia coli population composed of ∼99% susceptible cells (ATCC 35218 strain) and a ∼1% isogenic resistant subpopulation that overproduces TEM-1 ß-lactamase. After only 24 hours of treatment in the neutropenic murine thigh infection model, the generic amplified the resistant subpopulation up to 20-times compared with the innovator, following an inverted-U dose-response relationship. These findings highlight the critical role of therapeutic nonequivalence of generic antibiotics as a key factor contributing to the global problem of bacterial resistance.

Usefulness of FTA® cards as a Pneumocystis-DNA extraction method in bronchoalveolar lavage samples.

BACKGROUND: FTA® cards (Fast Technology for Analysis of Nucleic Acids) are an alternative DNA extraction method in bronchoalveolar lavage (BAL) samples for Pneumocystis jirovecii molecular analyses. The goal was to evaluate the usefulness of FTA® cards to detect P. jirovecii-DNA by PCR in BAL samples compared to silica adsorption chromatography (SAC). METHODS: This study used 134 BAL samples from immunocompromised patients previously studied to establish microbiological aetiology of pneumonia, among them 15 cases of Pneumocystis pneumonia (PCP) documented by staining and 119 with other alternative diagnoses. The FTA® system and SAC were used for DNA extraction and then amplified by nested PCR to detect P. jirovecii. Performance and concordance of the two DNA extraction methods compared to P. jirovecii microscopy were calculated. The influence of the macroscopic characteristics, transportation of samples and the duration of the FTA® card storage (1, 7, 10 or 12 months) were also evaluated. RESULTS: Among 134 BAL samples, 56% were positive for P. jirovecii-DNA by SAC and 27% by FTA®. All 15 diagnosed by microscopy were detected by FTA® and SAC. Specificity of the FTA® system and SAC were 82.4% and 49.6%, respectively. Compared to SAC, positivity by FTA® decreased with the presence of blood in BAL (62% vs 13.5%). The agreement between samples at 7, 10 and 12 months was 92.5% for FTA®. Positive cases by FTA® remained the same after shipment by mail. CONCLUSIONS: Results suggest that FTA® is a practical, safe and economical method to preserve P. jirovecii-DNA in BAL samples for molecular studies.