Pneumocystis jirovecii haplotypes at the internal transcribed spacers of the rRNA operon in French HIV-negative patients with diverse clinical presentations of Pneumocystis infections.

Pneumocystis jirovecii, a transmissible fungus, is the causative agent of pulmonary infections. Its genomic diversity has appeared in reports from around the world but data on P. jirovecii genotypes in France are still limited. This study describes the typing of P. jirovecii isolates from 81 HIV-negative patients monitored at Brest University Hospital, Brittany, France, 40 of whom developed Pneumocystis pneumonia (PcP), and remaining 41 patients were colonized by the fungus. The isolates were assayed at the internal transcribed spacer (ITS)1 and ITS2 under improved amplification conditions to avoid in vitro ITS recombination. P. jirovecii ITS haplotypes were identified in 56/81 patients (31 PcP patients and 25 patients who were colonized) which revealed a high diversity in that 27 different haplotypes were identified. Eg was the most frequent haplotype (31/56, 55.3%), followed by Ec and Ai (5/56, 8.9% each). In contrast, Ne, usually the second most frequent haplotype in Europe and the USA, was observed in only 2/56 patients (3.6%). Mixed infections were detected in 18/56 patients (32.1%; 12 PcP patients and six who were colonized). No significant differences were observed in haplotype diversity, frequency of peculiar haplotypes, and mixed infection occurrence, between the two patient populations. The study, conducted with the largest HIV-negative patient population investigated so far, shows that ITS typing remains an efficient method for characterizing P. jirovecii among human populations, whatever their clinical presentation of Pneumocystis infections.

A cluster of Pneumocystis infections among renal transplant recipients: molecular evidence of colonized patients as potential infectious sources of Pneumocystis jirovecii.

BACKGROUND: Eighteen renal transplant recipients (RTRs) developed Pneumocystis jirovecii infections at the renal transplantation unit of Brest University Hospital (Brest, Brittany, France) from May 2008 through April 2010, whereas no cases of P. jirovecii infection had been diagnosed in this unit since 2002. This outbreak was investigated by identifying P. jirovecii types and analyzing patient encounters. METHODS: The identification of P. jirovecii internal transcribed spacer (ITS) types was performed on P. jirovecii isolates from the 18 RTRs (12 patients with Pneumocystis pneumonia [PCP], 6 colonized patients), 22 unlinked control patients (18 patients with PCP, 4 colonized patients), and 69 patients (34 patients with PCP, 35 colonized patients) with contemporaneously diagnosed P. jirovecii infections in the Brest geographic area. A transmission map was drawn up. Its analysis was combined with the results of P. jirovecii typing. RESULTS: P. jirovecii ITS type identification was successful in 14 of 18 RTRs, 15 of 22 control patients, and 48 of the 69 patients. Type Eg was the most frequent type in the 3 patient groups. However, its frequency was significantly higher in the first patient group than in the 2 other groups (P < .05 and P < .01, respectively). Fourteen encounters between RTRs who harbored an identical type were observed. Ten patients were considered as possible index patients, of whom 3 were colonized by the fungus, and 7 presented PCP. CONCLUSIONS: The results provide to our knowledge the first data on the role of colonized patients as potential sources of P. jirovecii in a context of nosocomial acquisition of the fungus.

Systematic overestimation of human serum albumin by capillary zone electrophoresis method due to monoclonal immunoglobulin interferences.

BACKGROUND: The capillary zone electrophoresis method of albumin measurement is frequently used in monoclonal gammopathy patients but some studies suggest poor performances of the method in this population. The aim of this study was to analyse the impact of serum monoclonal immunoglobulins on human serum albumin determination by capillary zone electrophoresis method compared to other available methods. METHOD: We prospectively measured albumin in 100 freshly collected non-frozen serum samples in a monoclonal gammopathy patients population, by using four different methods: the capillary zone electrophoresis method, the bromocresol purple dye method, the nephelometric method and the turbidimetric method. Differences in albumin values between the different methods were analysed with respect to serum monoclonal immunoglobulin concentration. These differences were further investigated by measuring albumin levels in human serum samples spiked with exogenous monoclonal immunoglobulins. RESULTS: Human serum albumin difference values between capillary zone electrophoresis compared to immunonephelometry method are significantly correlated with increasing monoclonal immunoglobulins concentrations: regression analyses revealed a correlation coefficient r2 = 0.60 and a slope of 0.14 (0.12-0.17, 95% confidence interval). The capillary zone electrophoresis method overestimated serum albumin levels by up to 67% (12 g/L) when monoclonal immunoglobulin level was 63 g/L. The determination of albumin levels in human serum samples spiked with exogenous monoclonal immunoglobulins showed an overestimation of human serum albumin measurement by the capillary zone electrophoresis method proportional to the amount of monoclonal immunoglobulin added in the serum with a slope of 0.19 (0.18-0.20, 95% confidence interval). CONCLUSION: Monoclonal immunoglobulins directly interfere with serum albumin measurement by the capillary zone electrophoresis method leading to a systematic overestimation of serum albumin concentrations proportional to the serum monoclonal immunoglobulin level.

Is capillary zone electrophoresis a suitable method for estimating serum albumin: A comparison of four methods.

BACKGROUND: The capillary zone electrophoresis method of albumin measurement is frequently used for oncologic and haematologic patients but few data exist about the agreement between the albumin measurements performed by capillary zone electrophoresis and other methods. The aim of this study was to analyse the agreement between human serum albumin measurements by capillary zone electrophoresis and by the nephelometry, bromocresol purple and turbidimetry methods. METHOD: We prospectively measured 100 freshly collected non-frozen patient serum samples, by using four different methods: the capillary zone electrophoresis method performed with a CAPILLARYS 2 instrument, the bromocresol purple dye method performed with an Advia XPT analyser, the nephelometric method performed with a BN ProSpec analyser and the turbidimetric method with reagents from DiAgam and performed with the Advia XPT analyser. RESULTS: A bias towards higher values in the lower range of albumin concentrations was observed with capillary zone electrophoresis compared to immunonephelometry: correlation coefficient r2 = 0.925; slope of 0.86 (0.82-0.89, 95% confidence interval), which is significantly different from 1; and an intercept of 4.94 g/L (3.67-6.16, 95% confidence interval). Similar results were observed when comparing capillary zone electrophoresis to the bromocresol purple and immunoturbidimetry methods. The capillary electrophoresis method overestimated low albumin levels by up to 25% (5 g/L). CONCLUSION: Compared to the nephelometry, turbidimetry and bromocresol purple methods, the capillary zone electrophoresis method tends to overestimate human serum albumin concentrations for levels below 30 g/L. This discrepancy could lead to an overestimation of the nutritional status, an inappropriate scoring of the disease and a delay in nutritional treatment.

Absence of Pneumocystis dihydropteroate synthase mutants in Brittany, France.

Archival Pneumocystis jirovecii specimens from 84 patients monitored at Rennes University Hospital (Rennes, France) were assayed at the dihydropteroate synthase (DHPS) locus. No patient was infected with mutants. The results provide additional data showing that P. jirovecii infections involving DHPS mutants do not represent a public health issue in Brittany, western France.

Circulation of Pneumocystis dihydropteroate synthase mutants in France.

Data on the prevalence of Pneumocystis jirovecii (P. jirovecii) dihydropteroate synthase (DHPS) mutants in France are still limited. In this study, mutant prevalence in the Brest region (western France) was determined. Archival pulmonary specimens from 85 patients infected with P. jirovecii and admitted to our institution (University Hospital, Brest) from October 2007 to February 2010 were retrospectively typed at the DHPS locus using a polymerase chain reaction-restriction fragment length polymorphism assay. Type identification was successful in 66 of 85 patients. Sixty-four patients were infected with a wild type, whereas mutants were found in 2 patients (2/66, 3%). Medical chart analysis revealed that these 2 patients usually lived in Paris. Another patient usually lived on the French Riviera, whereas 63 patients were from the city of Brest. Thus, the corrected prevalence of mutants in patients who effectively lived in our geographic area was 0% (0/63). Taking into account that i) Paris is characterized by a high prevalence of mutants from 18.5% to 40%, ii) infection diagnoses were performed in the 2 Parisians during their vacation <30 days, iii) infection incubation is assumed to last about 2 months, the results provide evidence of mutant circulation from Paris to Brest through infected vacationers. The study shows that the usual city of patient residence, rather than the city of infection diagnosis, is a predictor of mutants and that P. jirovecii infections involving mutants do not represent a public health issue in western France.