A Herpes Simplex Virus 1 DNA Polymerase Multidrug Resistance Mutation Identified in a Patient With Immunodeficiency and Confirmed by Gene Editing.

BACKGROUND: Herpes simplex virus 1 can cause severe infections in individuals who are immunocompromised. In these patients, emergence of drug resistance mutations causes difficulties in infection management. METHODS: Seventeen herpes simplex virus 1 isolates were obtained from orofacial/anogenital lesions in a patient with leaky severe combined immunodeficiency over 7 years, before and after stem cell transplantation. Spatial/temporal evolution of drug resistance was characterized genotypically-with Sanger and next-generation sequencing of viral thymidine kinase (TK) and DNA polymerase (DP)-and phenotypically. CRISPR/Cas9 was used to introduce the novel DP Q727R mutation, and dual infection-competition assays were performed to assess viral fitness. RESULTS: Isolates had identical genetic backgrounds, suggesting that orofacial/anogenital infections derived from the same virus lineage. Eleven isolates proved heterogeneous TK virus populations by next-generation sequencing, undetectable by Sanger sequencing. Thirteen isolates were acyclovir resistant due to TK mutations, and the Q727R isolate additionally exhibited foscarnet/adefovir resistance. Recombinant Q727R mutant virus showed multidrug resistance and increased fitness under antiviral pressure. CONCLUSIONS: Long-term follow-up of a patient with severe combined immunodeficiency revealed virus evolution and frequent reactivation of wild-type and TK mutant strains, mostly as heterogeneous populations. The DP Q727R resistance phenotype was confirmed with CRISPR/Cas9, a useful tool to validate novel drug resistance mutations.

Heterogeneity and evolution of thymidine kinase and DNA polymerase mutants of herpes simplex virus type 1: implications for antiviral therapy.

BACKGROUND: Infections caused by acyclovir-resistant isolates of herpes simplex virus (HSV) after hematopoietic stem cell transplantation (HSCT) are an emerging concern. An understanding of the evolutionary aspects of HSV infection is crucial to the design of effective therapeutic and control strategies. METHODS: Eight sequential HSV-1 isolates were recovered from an HSCT patient who suffered from recurrent herpetic gingivostomatitis and was treated alternatively with acyclovir, ganciclovir, and foscavir. The diverse spectra and temporal changes of HSV drug resistance were determined phenotypically (drug-resistance profiling) and genotypically (sequencing of the viral thymidine kinase and DNA polymerase genes). RESULTS: Analysis of 60 clones recovered from the different isolates demonstrated that most of these isolates were heterogeneous mixtures of variants, indicating the simultaneous infection with different drug-resistant viruses. The phenotype/genotype of several clones associated with resistance to acyclovir and/or foscavir were identified. Two novel mutations (E798K and I922T) in the viral DNA polymerase could be linked to drug resistance. CONCLUSIONS: The heterogeneity within the viral populations and the temporal changes of drug-resistant viruses found in this HSCT recipient were remarkable, showing a rapid evolution of HSV-1. Drug-resistance surveillance is highly recommended among immunocompromised patients to manage the clinical syndrome and to avoid the emergence of multidrug-resistant isolates.

A Rare Case of Hepatic Vanishing Bile Duct Syndrome Occurring after Combination Therapy with Nivolumab and Cabozantinib in a Patient with Renal Carcinoma.

Tyrosine kinase inhibitors (TKIs) and immune checkpoint inhibitors (ICIs) significantly improve the outcomes of patients with advanced clear cell renal cell carcinoma (ccRCC); however, high-grade toxicities can occur, particularly during combination therapy. Herein, we report a patient with advanced metastatic ccRCC, who developed grade 4 cholestasis during combined therapy with nivolumab and cabozantinib. After the exclusion of common disorders associated with cholestasis and a failure of corticosteroids (CS), a liver biopsy was performed that demonstrated severe ductopenia. Consequently, a diagnosis of vanishing bile duct syndrome related to TKI and ICI administration was made, resulting in CS discontinuation and ursodeoxycholic acid administration. After a 7-month follow-up, liver tests had returned to normal values. Immunological studies revealed that our patient had developed robust T-cells and macrophages infiltrates in his lung metastasis, as well as in skin and liver tissues at the onset of toxicities. At the same time, peripheral blood immunophenotyping revealed significant changes in T-cell subsets, suggesting their potential role in the pathophysiology of the disease.

Predicting febrile neutropenic patients at low risk using the MASCC score: does bacteremia matter?

BACKGROUND: Febrile neutropenic cancer patients represent a heterogeneous population with a limited proportion at risk of serious medical complications. The Multinational Association for Supportive Care in Cancer (MASCC) score has been developed and validated for identifying low-risk patients at the onset of febrile neutropenia. Since bacteremia, although not documented at baseline, is a predictor of pejorative outcome, the purpose of this study was to investigate the possible interaction between the MASCC score and bacteremic status and to assess whether, assuming that bacteremic status could be predicted at onset of febrile neutropenia, adding bacteremia as a covariate in a risk model would improve the accuracy of low-risk patients identification. METHODS: Two consecutive multicentric observational studies were carried out from 1994 till 2005 involving 2,142 febrile neutropenic patients. The study data bases were retrospectively used for the present analysis. RESULTS: A predictive value was found for the MASCC score in all strata obtained by stratification for the bacteremic status with odds ratios for successful outcome being, in patients with a score ≥21, respectively, 6.06 (95%CI: 4.51-8.15), 3.42 (95%CI: 1.95-5.98), and 6.04 (95%CI: 3.01-12.09) in patients without bacteremia, gram-positive bacteremia, and gram-negative bacteremia. No interaction between the MASCC score and the bacteremic status was present. A clinical prediction rule integrating the MASCC score and the bacteremic status was not helpful in improving the identification of low-risk patients. This rule may then be used in a general population of patients with febrile neutropenia without having concerns for a lower predictive value in bacteremic patients. CONCLUSIONS: Our results suggest that the knowledge, provided we could find a model to predict it at fever onset, of a bacteremic etiology of the fever would be of little additional value to the MASCC score when attempting to identify low-risk patients.

Performance of Xpert MTB/RIF Ultra for diagnosis of pulmonary and extra-pulmonary tuberculosis, one year of use in a multi-centric hospital laboratory in Brussels, Belgium.

Among the challenges in controlling tuberculosis, a rapid and accurate diagnostic test for the detection of Mycobacterium tuberculosis complex (MTBc) and its resistance to first line therapies is crucial. We evaluated the performance of the Xpert MTB/RIF Ultra assay (Xpert Ultra) for the rapid detection of MTBc and rifampicin resistance (RR) in 1120 pulmonary and 461 extra-pulmonary clinical specimens and compared it with conventional phenotypic techniques. The Xpert Ultra assay detected MTBc in 223 (14.1%) samples with an overall sensitivity and specificity, using culture as the "gold standard", of 91.1% (95% CI, 85.6-95.1) and 94.5% (95% CI, 93.1-95.6), respectively. The sensitivity of the Xpert Ultra test for smear-negative extra-pulmonary specimens was high (87.1%), even higher than with smear-negative pulmonary specimens (81.8%). But this enhanced sensitivity came with a low overall specificity of smear-negative extra-pulmonary specimens (66.7%). For 73 patients, 79/1423 (3.4%) negative mycobacterial culture samples were found to be positive with Xpert Ultra. Clinical data was necessary to correctly interpret potential false-positive results, especially trace-positive results. Sensitivity of the Xpert Ultra to detect RR compared to drug susceptibility testing was 100% (95% CI, 29.2-100) and specificity was 99.2% (95% CI, 95.8-100). We concluded that the Xpert Ultra test is able to provide a reliable TB diagnosis within a significantly shorter turnaround time than culture. This is especially true for paucibacillary samples such as smear-negative pulmonary specimens and extra-pulmonary specimens.

Small cell carcinoma of the endometrium: A clinicopathological study and management of three cases.

Endometrial small cell carcinoma (ESCC) is an extremely rare and aggressive tumor with poor prognosis. It is characterized by early regional and systemic spread leading to rapid development of lymph nodes, pelvic and extrapelvic metastasis and compromising the outcome. In this paper, we reported three cases of ESCC confirmed by pathological and immunohistochemistry studies. In one case, ESCC was associated with endometrioid carcinoma and carcinosarcoma, while the other two cases were pure ESCC. Two cases were diagnosed at early stage IA of the International Federation of Gynecology and Obstetrics (FIGO) cancer staging system. They were treated by surgery followed by pelvic external radiation and brachytherapy with favorable outcome (no recurrence was confirmed and a survival was 1 and 5years, respectively). The third case was diagnosed with visceral metastasis and was treated with 6 cycles of cisplatin plus etoposide. She died 8months after diagnosis. Due to its rarity, there is no standard guideline for the management of ESCC. Its treatment is extrapolated from that of both, the conventional endometrial carcinoma and the small cell carcinoma of the lungs, which share similarities with ESCC. Thus, multimodal therapeutic including surgery, radiation therapy and chemotherapy, seems to be the best therapeutic approach. Randomized clinical trials with multiples cases of ESCC are encouraged to clearly define the optimal therapeutic approach to this rare tumor.

Zygomycosis in the immunocompromised patient: a case report.

Zygomycosis is a highly aggressive infection observed in immunocompromised patients, such as those with haematological malignancies. The sites most frequently involved are the sinuses and the lungs. New diagnostic tools and new antifungal treatments are essential in order to diagnose early and treat efficiently infections due to moulds. We report a case of sinusitis due to Absidia corymbifera occurring during chemotherapy-induced bone marrow aplasia in a patient with acute leukaemia. The sinusitis was successfully treated with AmBisome, and surgical debridement.

Intravesical instillation of cidofovir in the treatment of hemorrhagic cystitis caused by adenovirus type 11 in a bone marrow transplant recipient.

Hemorrhagic cystitis that occurs late after bone marrow transplantation (BMT) in BMT recipients is often associated with adenovirus or polyomavirus BK infections. Intravesical instillation of cidofovir in a BMT recipient with intractable hemorrhagic cystitis resulted in clinical improvement. Local cidofovir therapy for viral hemorrhagic cystitis could be an alternative to intravenous administration of cidofovir.

Prophylaxis of febrile neutropenia in adults receiving chemotherapy needs to be adapted to the risk.

A major advance in the management of febrile neutropenia (FN) has been the stratification of the population of adult patients with FN for the risk of complications and death. Using validated reliable predictive instruments, such as the Multinational Association for Supportive Care in Cancer score, it is possible to identify a population of 'low-risk' patients, who can benefit from simplified and less expensive therapeutic approaches (e.g., orally administered antimicrobial therapy and early home return). Prevention of FN by the use of granulopoietic colony-stimulating factor (G-CSF) has been successfully applied to patients at 'high risk' of developing FN. In addition to the aggressiveness of chemotherapy, which usually defines the 'high-risk' status, the role of a series of factors that increase both the risk of FN and the complications rate has been recognized and should probably be taken into consideration when selecting patients for G-CSF prophylaxis. The cost of the G-CSF is the major limiting factor for their broad use; further efforts should be made to match the cost issue with the need of protecting from the development of FN most patients treated with chemotherapy for cancer.

Early detection and identification of commonly encountered Candida species from simulated blood cultures by using a real-time PCR-based assay.

In a recent study, Candida species in clinical blood samples were detected using a real-time PCR-based method (Maaroufi et al, J Clin Microbiol 2003, 41:3293-3298). For the present study, we evaluated the efficiency of this method as an adjunct to the BACTEC blood culture system to early detection of positivity and negativity of simulated low candidemias. We first established an in vitro correlation between the inoculum of the most frequently encountered Candida species and the time to positivity of these microorganisms. Then, aliquots from blood culture bottles infected with a final average candidal inoculum of 3.18 colony-forming units (CFU)/culture bottle (range, 1 to 6 CFU) were collected at increasing incubation times, and DNA was extracted and submitted to the TaqMan-based PCR assay. To optimize this assay, we evaluated the effect of adding 0.5% bovine serum albumin (BSA) to DNA extracts and found that it decreased the effects of inhibitors. Using specific probes for the tested Candida species, the PCR assay was positive on blood culture aliquots collected from the BACTEC system after a minimum culture turnaround time (TAT) of 3.11 +/- 1.24 hours. Addition of BSA to PCR reaction mixtures improves the TAT (1.84 +/- 0.41 hours). Hence, the combination of DNA "amplification" in the culture bottles by normal growth with an additional DNA amplification by PCR might be a reliable tool facilitating the early diagnosis of low candidemias.

Strategies for the empirical management of infection in cancer patients with emphasis on the emergence of resistant gram-negative bacteria.

Combinations of antibiotics (namely penicillins and aminoglycosides) have been advocated in the 1970s for the empirical therapy of FN in cancer patients in order to take advantage of the possible synergism between these agents and to extend the potential antimicrobial spectrum of empirical therapy. Later, with the development of potent broad spectrum antibiotics, the need for combinations became less obvious as monotherapy with these new agents appeared as effective and less toxic than previously used combinations. However, today we are facing a major challenge through the emergence of multi-resistant microrganisms. With such bacteria, we might be coming back to the pre-antibiotic era when no active agents were available. This situation is due, in part, by the excessive use of antibiotics, namely as a prophylaxis for infection, and is complicated by the fact that very few new effective antibiotics are being developed by the pharmaceutical industry. Under these circumstances, it is likely that we will have to resort to "old timers" such as the polymyxins. It is also possible that combination therapy will come back in favor to take advantage of the synergism and extend the spectrum of coverage, just as it has been the case for the management of resistant tuberculosis. At the same time, the development of multidisciplinary antimicrobial stewardship is mandatory for efficient infection control and minimizing emergence of antimicrobial resistance.

Outpatient oral antibiotics for febrile neutropenic cancer patients using a score predictive for complications.

PURPOSE: Since febrile neutropenic patients were recognized to constitute a heterogeneous population, several models have been developed for predicting the risk of serious medical complications. The Multinational Association for Supportive Care in Cancer score and its derived clinical prediction rules have been validated, but thus far there were no data about its use for simplifying therapy in predicted low-risk patients. PATIENTS AND METHODS: In a single institution, we followed all episodes of febrile neutropenia between January 1999 and November 2003. Those patients predicted at low risk for complications, who were not receiving antibacterials at fever onset and were eligible for treatment with oral antibiotics, were treated with ciprofloxacin and amoxicillin-clavulanate and were discharged if they were clinically stable or improving after an initial observation period. The primary end point of the study was the rate of resolution of the febrile neutropenic episode without complications, among these early discharged patients. RESULTS: Of 383 first febrile neutropenic episodes predicted at low risk of complication, 178 patients (33 men and 145 women, mainly with solid tumors) were treated orally; they constituted the basis of our analysis. Seventy-nine patients (44%) were discharged early (with a median time to discharge of 26 hours); no complications occurred among them but three patients had to be readmitted, resulting in a success rate of 96% (95% CI, 92% to 100%). CONCLUSION: Our study shows that oral therapy followed by early discharge was feasible in a small but significant proportion of patients selected by a strategy combining predicted low risk and medical and nonmedical criteria.