We report the genotyping analysis of Toxoplasma gondii isolates in samples collected from 88 immunocompromised patients, along with clinical and epidemiological data. Most of these samples were collected in France during the current decade by the Toxoplasma Biological Resource Center. Lack of specific anti-Toxoplasma treatment, pulmonary toxoplasmosis, and involvement of multiple organs were the 3 main risk factors associated with death for this patient group. Genotyping results with 6 microsatellite markers showed that type II isolates were predominant among patients who acquired toxoplasmic infection in Europe. Non-type II isolates included 13 different genotypes and were mainly collected from patients who acquired toxoplasmosis outside Europe. Type III was the second most common genotype recovered from patients, whereas type I was rare in our population. Three nonarchetypal genotypes were repeatedly recovered from different patients who acquired the infection in sub-Saharan Africa (genotypes Africa 1 and Africa 2) and in the French West Indies (genotype Caribbean 1). The distribution of genotypes (type II vs. non-type II) was not significantly different when patients were stratified by underlying cause of immunosuppression, site of infection, or outcome. We conclude that in immunocompromised patients, host factors are much more involved than parasite factors in patients' resistance or susceptibility to toxoplasmosis.

We present a case of disseminated congenital toxoplasmosis in a newborn born to a mother who had been immunized against toxoplasmosis before conception. The mother was reinfected, likely by ingestion of imported raw horse meat during pregnancy. This clinical presentation is exceptional in France and raised the possibility of infection by a highly virulent Toxoplasma strain. The strain responsible was isolated from the peripheral blood of the newborn, and when genotyped with microsatellite markers, it exhibited an atypical genotype, one which is very uncommon in Europe but had been described in South America. We tested the hypothesis of a reinfection with a different genotype by using an experimental mouse model, which confirmed that acquired immunity against European Toxoplasma strains may not protect against reinfection by atypical strains acquired during travel outside Europe or by eating imported meat.

The isolation of Candida species from a single blood culture is considered sufficient evidence for the initiation of systemic antifungal therapy. However, blood cultures still lack sensitivity. Previous reports have suggested that the combined serological detection of mannanemia and anti-mannan antibodies may be useful for the diagnosis of systemic candidiasis caused by Candida albicans (specificity and sensitivity 93% and 80%, respectively). In this study, serological tests to detect Candida albicans mannan and Candida albicans antibodies (Platelia Candida Antigen and Antibody tests; Bio-Rad, France) were applied retrospectively to a series of patients with at least one Candida-positive blood culture and from whom at least one serum sample, taken before or on the day of blood culture, was available. Forty-five patients were selected, including 23 infected by Candida albicans, 4 by Candida glabrata, 9 by Candida tropicalis, 5 by Candida parapsilosis, and 4 by Candida krusei. Serological tests were positive in 73% of patients at least 2 days, and in some patients, up to 15 days before blood cultures became positive. These data suggest that serological surveillance of at-risk patients using the Platelia Candida tests could result in earlier initiation of antifungal therapy, especially when used in conjunction with blood cultures. In this way, more efficient management of nosocomial infections caused by Candida species can be achieved.

Ocular toxoplasmosis is a major cause of posterior uveitis worldwide. The diagnosis is based mainly on ophthalmological examination. Biological diagnosis is necessary in atypical cases, and this requires aqueous humor sampling by anterior chamber paracentesis. We evaluated real-time PCR targeting the Toxoplasma gondii 529-bp repeat element, the Goldmann-Witmer coefficient (GWC), and immunoblotting for the diagnosis of toxoplasmic retinochoroiditis in 54 patients with atypical uveitis. The results of these biological tests, applied to paired aqueous humor-serum samples, were compared to the clinical findings. Combining either PCR or the GWC with immunoblotting increased the sensitivity to 73% or 70%, respectively. Together, PCR and the GWC had 80% sensitivity. If feasible, sensitivity can be increased by combining the three methods (85% sensitivity). The interval between symptom onset and anterior chamber paracentesis strongly influenced the detection of specific intraocular antibody synthesis. The sensitivity of the GWC increased from 45% to 56% when sampling was performed 10 days after symptom onset, and that of immunoblotting increased from 53% to 72% when puncture was performed 30 days after symptom onset. PCR analysis of aqueous humor samples detected toxoplasmic DNA in 55% of patients. In contrast to the results of immunoblotting and the GWC, the results of PCR were not influenced by the interval between symptom onset and paracentesis. PCR was more informative than the GWC and immunoblotting for immunocompromised patients. Acute necrotizing retinal lesions were significantly larger in PCR-positive patients, with a mean of 3.5 optic disc diameters, than in PCR-negative patients, with a mean of 1.5 optic disc diameters.

Although screening for maternal toxoplasmic seroconversion during pregnancy is based on immunodiagnostic assays, the diagnosis of clinically relevant toxoplasmosis greatly relies upon molecular methods. A problem is that this molecular diagnosis is subject to variation of performances, mainly due to a large diversity of PCR methods and primers and the lack of standardization. The present multicentric prospective study, involving eight laboratories proficient in the molecular prenatal diagnosis of toxoplasmosis, was a first step toward the harmonization of this diagnosis among university hospitals in France. Its aim was to compare the analytical performances of different PCR protocols used for Toxoplasma detection. Each center extracted the same concentrated Toxoplasma gondii suspension and tested serial dilutions of the DNA using its own assays. Differences in analytical sensitivities were observed between assays, particularly at low parasite concentrations (<2 T. gondii genomes per reaction tube), with "performance scores" differing by a 20-fold factor among laboratories. Our data stress the fact that differences do exist in the performances of molecular assays in spite of expertise in the matter; we propose that laboratories work toward a detection threshold defined for a best sensitivity of this diagnosis. Moreover, on the one hand, intralaboratory comparisons confirmed previous studies showing that rep529 is a more adequate DNA target for this diagnosis than the widely used B1 gene. But, on the other hand, interlaboratory comparisons showed differences that appear independent of the target, primers, or technology and that hence rely essentially on proficiency and care in the optimization of PCR conditions.

T oxoplasmosis is a widespread parasitic infection that is frequently asymptomatic in immunocompetent patients. However, this obligate intracellular protozoan parasite can evade the immune system (1, 2) and persist for the life of its host in cyst form, predominantly in the brain, retina, and muscles. Reactivation of latent cysts may occur when the immune system fails to maintain cytokine pressure, which mainly relies on gamma interferon (IFN-␥) (3). Cyst reactivation can lead to ocular toxoplasmosis, cerebral toxoplasmosis (CT), or disseminated toxoplasmosis, which involves most frequently the lungs but potentially all organs. Failure of an efficient Th1 immune response mainly results from acquired immunosuppression, through HIV infection or immunosuppressive therapy. Both primary acquired and reactivated infections are life-threatening in immunocompromised patients (ICPs). Definitive diagnosis can be obtained by the detection of parasites in blood, cerebrospinal fluid (CSF), bronchoalveolar lavage (BAL) fluid, or virtually any tissue by using PCR, which is the most sensitive method (4).Prevention of CT in patients with HIV is an object of consensus, and guidelines recommend co-trimoxazole (sulfamethoxazole-trimethoprim) chemoprophylaxis in Toxoplasma-seropositive patients when CD4 ϩ cell counts fall below 200 cells/l (4), a prophylactic regimen which also protects patients from Pneumocystis jirovecii pneumonia. Nevertheless, toxoplasmosis remains Citation Robert-Gangneux F, Sterkers Y, Yera H, Accoceberry I, Menotti J, Cassaing S, Brenier-Pinchart M-P, Hennequin C, Delhaes L, Bonhomme J, Villena I, Scherer E, Dalle F, Touafek F, Filisetti D, Varlet-Marie E, Pelloux H, Bastien P. 2015. Molecular diagnosis of toxoplasmosis in immunocompromised patients: a 3-year multicenter retrospective study.

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