Features of cryptococcosis among 652 HIV-seronegative individuals in France: a cross-sectional observational study (2005-2020).

OBJECTIVES: We aimed to describe features and outcomes of cryptococcosis among HIV-seronegative individuals in a large surveillance network for cryptococcosis in France. METHODS: We included incident cases of cryptococcosis in HIV-seronegative individuals from 2005 to 2020. We compared patient characteristics, disease presentations, cryptococcal antigen results, and induction antifungal treatments according to underlying disease. We examined factors associated with 90-day mortality. Among patients with disseminated infections, we investigated whether receipt of flucytosine and polyene combination was associated with lower mortality. RESULTS: Among 652 individuals, 209 (32.1%) had malignancy, 130 (19.9%) were solid-organ transplant recipients, 204 (31.3%) had other immunocompromising conditions, and 109 (16.7%) had no reported underlying factor. The commonest presentations were disseminated infections (63.3%, 413/652) and isolated pulmonary infections (25.3%, 165/652). Solid-organ transplant patients were most likely to have disseminated infections and a positive serum cryptococcal antigen result. Patients with malignancy were older and less likely to receive a flucytosine-containing regimen for disseminated infections than others (58.7%, 78/133 vs. 73.2%, 194/265; p 0.029). The crude 90-day case-fatality ratio was 27.2% (95% CI, 23.5%-31.1%). Age ≥60 years (aOR: 2.75 [1.78-4.26]; p < 0.001), meningitis/fungaemia (aOR: 4.79 [1.80-12.7]; p 0.002), and malignancy (aOR: 2.4 [1.14-5.07]; p 0.02) were associated with higher 90-day mortality. Receipt of flucytosine and polyene combination was associated with lower 90-day mortality (aOR: 0.40 [0.23-0.71]; p 0.002) in multivariable analysis and inverse probability of treatment weighted analysis (aOR: 0.45 [0.25-0.80]; p 0.006). DISCUSSION: HIV-seronegative individuals with cryptococcosis comprise a wide range of underlying conditions with different presentations and outcomes, requiring a tailored approach to diagnosis and management.

Invasive fungal diseases in patients with autoimmune diseases: a case series from the French RESSIF network.

OBJECTIVES: We aimed to describe patients with autoimmune diseases (AID) developing invasive fungal disease (IFD) and identify factors associated with short-term mortality. METHODS: We analysed cases of IFD associated with AID from the surveillance network of invasive fungal diseases (Réseau de surveillance des infections fongiques invasives, RESSIF) registry of the French national reference centre for invasive mycoses. We studied association of AID-specific treatments with 30-day mortality. We analysed total lymphocyte and CD4-T cell counts in patients with Pneumocystis jirovecii pneumonia (PCP). RESULTS: From 2012 to 2018, 549 individuals with IFD and AID were included, mainly with PCP (n=227, 41.3%), fungemia (n=167, 30.4%) and invasive aspergillosis (n=84, 15.5%). Rheumatoid arthritis (RA) and anti-neutrophil cytoplasmic antibodies (ANCA)-associated vasculitides (AAV) were the most frequent AID in PCP (n=55 and 25, respectively) and invasive aspergillosis (n=15 and 10, respectively), inflammatory bowel diseases (IBDs) were predominant in fungemia (n=36). At IFD diagnosis, 365 (66.5%) patients received glucocorticoids (GCs), 285 (51.9%) immunosuppressants, 42 (7.7%) tumor necrosis factor (TNF)-α blockers, 75 (13.7%) other biologics. Mortality at 30 days was 28.1% (143/508). Fungemia and high-dose GCs were independently associated with higher 30-day mortality. In PCP patients, lymphopenia <1500/mm3 was frequent (132/179, 73.7%) even if CD4+T cell count exceeded 200/mm3 in 56/78 patients (71.8%) (median 472.5/mm3, IQR 160-858). CONCLUSION: IFD associated with AID occurs primarily in RA, AAV and IBD, especially when treated with GCs and immunosuppressants. Mortality is high, especially for patients on high-dose GCs. Lymphopenia may help identify risk of PCP, but normal CD4+T cell count does not rule out the risk. Further studies are needed to assess the individual risk factors for IFD.

Fecal dysbiosis associated with colonic hypersensitivity and behavioral alterations in chronically Blastocystis-infected rats.

BACKGROUND: Infectious gastroenteritis is a risk factor for the development of post-infectious Irritable Bowel Syndrome (PI-IBS). Recent clinical studies reported a higher prevalence of the intestinal parasite Blastocystis in IBS patients. Using a rat model, we investigated the possible association between Blastocystis infection, colonic hypersensitivity (CHS), behavioral disturbances and gut microbiota changes. METHODS: Rats were orally infected with Blastocystis subtype 4 (ST4) cysts, isolated from human stool samples. Colonic sensitivity was assessed by colorectal distension and animal behavior with an automatic behavior recognition system (PhenoTyper), the Elevated Plus Maze test and the Forced Swimming tests. Feces were collected at different time points after infection to study microbiota composition by 16 S rRNA amplicon sequencing and for short-chain fatty acid (SFCA) analysis. RESULTS: Blastocystis-infected animals had non-inflammatory CHS with increased serine protease activity. Infection was also associated with anxiety- and depressive-like behaviors. Analysis of fecal microbiota composition showed an increase in bacterial richness associated with altered microbiota composition. These changes included an increase in the relative abundance of Oscillospira and a decrease in Clostridium, which seem to be associated with lower levels of SCFAs in the feces from infected rats. CONCLUSIONS: Our findings suggest that experimental infection of rats with Blastocystis mimics IBS symptoms with the establishment of CHS related to microbiota and metabolic shifts.

Genotyping Approach for Potential Common Source of Enterocytozoon bieneusi Infection in Hematology Unit.

Microsporidiosis is a fungal infection that generally causes digestive disorders, especially in immunocompromised hosts. Over a 4-day period in January 2018, 3 patients with hematologic malignancies who were admitted to the hematology unit of a hospital in France received diagnoses of Enterocytozoon bieneusi microsporidiosis. This unusually high incidence was investigated by sequence analysis at the internal transcribed spacer rDNA locus and then by 3 microsatellites and 1 minisatellite for multilocus genotyping. The 3 isolates had many sequence similarities and belonged to a new genotype closely related to genotype C. In addition, multilocus genotyping showed high genetic distances with all the other strains collected from epidemiologically unrelated persons; none of these strains belonged to the new genotype. These data confirm the epidemiologic link among the 3 patients and support a common source of infection.

Efficient and reproducible experimental infections of rats with Blastocystis spp.

Although Blastocystis spp. infect probably more than 1 billion people worldwide, their clinical significance is still controversial and their pathophysiology remains poorly understood. In this study, we describe a protocol for an efficient and reproducible model of chronic infection in rats, laying the groundwork for future work to evaluate the pathogenic potential of this parasite. In our experimental conditions, we were unable to infect rats using vacuolar forms of an axenically cultivated ST4 isolate, but we successfully established chronic infections of 4 week-old rats after oral administration of both ST3 and ST4 purified cysts isolated from human stool samples. The infection protocol was also applied to 4 week-old C57BL/9, BALB/C and C3H mice, but any mouse was found to be infected by Blastocystis. Minimal cyst inoculum required for rat infection was higher with ST3 (105) than with ST4 (102). These results were confirmed by co-housing experiments highlighting a higher contagious potential of ST4 in rats compared to ST3. Finally, experiments mimicking fecal microbiota transfer from infected to healthy animals showed that Blastocystis spp. could easily infect a new host, even though its intestinal microbiota is not disturbed. In conclusion, our results provide a well-documented and robust rat model of Blastocystis chronic infection, reproducing "natural" infection. This model will be of great interest to study host parasite interactions and to better evaluate clinical significance of Blastocystis.

Invasive Infections Caused by Nannizziopsis spp. Molds in Immunocompromised Patients.

We report 2 new cases of invasive infections caused by Nannizziopsis spp. molds in France. Both patients had cerebral abscesses and were immunocompromised. Both patients had recently spent time in Africa.

Encephalitis caused by an unusual human herpes virus type 6 and Toxoplasma gondii co-infection in a cord blood transplant recipient.

BACKGROUND: The case of a central nervous system human herpes virus type 6 (HHV-6) and Toxoplasma gondii co-infection after an umbilical cord blood transplantation in a chronic myelomonocytic leukaemia patient is reported. CASE REPORT: A 65-year-old Caucasian man underwent an umbilical cord blood transplantation within the context of chronic myelomonocytic leukaemia. On day 37 post-graft, he presented with a severe headache; PCRs of cerebrospinal fluid and blood were positive for T. gondii and HHV-6. The patient was treated with pyrimethamine and sulfadiazine associated with ganciclovir. CONCLUSION: HHV-6 reactivation can trigger a reactivation of T. gondii. This case suggests that patients who are seropositive for T. gondii and who present with HHV-6 reactivation should be monitored closely for toxoplasmosis.

Posttraumatic axillary false aneurysm after luxatio erecta of the shoulder: case report and literature review.

Vascular complications after dislocation of the shoulder are rare. We report a case of glenohumeral inferior dislocation (luxatio erecta) responsible for an acute ischemia of the upper limb. Endovascular treatment with a covered stent associated with the evacuation of the compressive hematoma was privileged. In the second stage, an axillary bypass was carried out because of an intrastent thrombosis responsible for an acute ischemia of the right upper limb. The stabilization of the glenohumeral articulation was obtained later with an anterior coracoid bone block. The conventional surgical treatment remains the standard treatment. Hybrid techniques with endovascular clamping can be useful in the presence of proximal arterial lesions. Endovascular treatment is an interesting therapeutic alternative in the urgency and in selected cases but its mid- and long-term results should still be evaluated.

Blastocystis, an unrecognized parasite: an overview of pathogenesis and diagnosis.

Blastocystis sp. is among the few enteric parasites with a prevalence that often exceeds 5% in the general population of industrialized countries and can reach 30-60% in developing countries. This parasite is frequently found in people who are immunocompromised (patients with human immunodeficiency virus/acquired immunodeficiency syndrome or cancer) and a higher risk of Blastocystis sp. infection has been found in people with close animal contact. Such prevalence in the human population and the zoonotic potential naturally raise questions about the impact of these parasites on public health and has increased interest in this area. Recent in vitro and in vivo studies have shed new light on the pathogenic power of this parasite, suggesting that Blastocystis sp. infection is associated with a variety of gastrointestinal disorders, may play a significant role in irritable bowel syndrome, and may be linked with cutaneous lesions (urticaria). Despite recent significant advances in the knowledge of the extensive genetic diversity of this species, the identification of extracellular proteases as virulence factors and the publication of one isolate genome, many aspects of the biology of Blastocystis sp. remain poorly investigated. In this review, we investigate several biological aspects of Blastocystis sp. (diversity and epidemiology, diagnosis tools and pathophysiology). These data pave the way for the following challenges concerning Blastocystis sp. research: deciphering key biological mechanisms and pathways of this parasite and clarification of its clinical impact in humans.

Characterization of two cysteine proteases secreted by Blastocystis ST7, a human intestinal parasite.

Blastocystis spp. are unicellular anaerobic intestinal parasites of both humans and animals and the most prevalent ones found in human stool samples. Their association with various gastrointestinal disorders raises the questions of its pathogenicity and of the molecular mechanisms involved. Since secreted proteases are well-known to be implicated in intestinal parasite virulence, we intended to determine whether Blastocystis spp. possess such pathogenic factors. In silico analysis of the Blastocystis subtype 7 (ST7) genome sequence highlighted 22 genes coding proteases which were predicted to be secreted. We characterized the proteolytic activities in the secretory products of Blastocystis ST7 using specific protease inhibitors. Two cysteine proteases, a cathepsin B and a legumain, were identified in the parasite culture supernatant by gelatin zymographic SDS-PAGE gel and MS/MS analysis. These proteases might act on intestinal cells and disturb gut function. This work provides serious molecular candidates to link Blastocystis spp. and intestinal disorders.

Development and evaluation of a real-time PCR assay for detection and quantification of blastocystis parasites in human stool samples: prospective study of patients with hematological malignancies.

Blastocystis anaerobic parasites are widespread worldwide in the digestive tract of many animal species, including humans. Epidemiological Blastocystis studies are often limited by the poor sensitivity of standard parasitological assays for its detection. This report presents a highly sensitive real-time quantitative PCR (qPCR) assay developed to detect Blastocystis parasites in stool samples. The assay targets a partial sequence of the Blastocystis small ribosomal subunit (SSU) rRNA gene, allowing subtyping (ST) of Blastocystis isolates by direct sequencing of qPCR products. This qPCR method was assessed in a prospective study of 186 patients belonging to two cohorts--a group of 94 immunocompromised patients presenting hematological malignancies and a control group of 92 nonimmunocompromised patients. Direct-light microscopy and xenic in vitro stool culture analysis showed only 29% and 52% sensitivity, respectively, compared to our qPCR assay. Of the 27 (14.5%) Blastocystis-positive patients, 8 (4%) experienced digestive symptoms. No correlation was found between symptomatic patients and immune status, parasite load, or parasite subtypes, although subtyping of all isolates revealed a high (63.0%) prevalence of ST4. Two unexpected avian subtypes were found, i.e., ST6 and ST7, which are frequently isolated in Asia but rarely present in Western countries. In conclusion, this qPCR proved by far the most sensitive of the tested methods and allowed subtype determination by direct sequencing of qPCR products. New diagnostic tools such as the qPCR are essential for evaluating the clinical relevance of Blastocystis subtypes and their role in acute or chronic digestive disorders.