Monitoring of cytomegalovirus (CMV) infection in solid organ transplant recipients: quantitation of CMV DNAemia by two real-time polymerase chain reaction assays

Background and aim: Quantification of cytomegalovirus (CMV) DNAemia is essential in clinical management of post-transplant infection. We evaluated the performances of two quantitative real-time polymerase chain reaction (PCR) assays. Materials and Methods: 114 serial whole blood samples collected from 14 actively infected transplant recipients were processed by Abbott RealTime CMV PCR kit (Abbott Molecular) and CMV ELITe MGB™ kit (ELITech Group). The Quality Control for Molecular Diagnostics human CMV panels was also tested. Results: Sixteen (14%) samples resulted negative and 59 (51.7%) positive with a quantitative result for both assays. In the 59 samples, the coefficient of correlation was 0.856. Bland-Altman analysis showed a mean difference of <0.11 log10 copies/mL (standard deviation=0.38 log10 copies/mL). The assays gave CMV-DNA loads differing by 1 log10 DNA copies/mL in 57 samples (96.6%) and by <0.5 log10 DNA copies/mL in 48 samples (81.3%). Eleven (9.6%) samples were positive with a quantitative result with Abbott and negative with ELITech. Sixteen (14%) positive samples with a quantitative result for Abbott resulted positive but below the lower limit of quantification (LLQ) for ELITech. Twelve (10.5%) samples resulted negative with ELITech and positive but below the LLQ with Abbott. No samples were positive with ELITech and negative with Abbott. Conclusions: The assays showed a good correlation between CMVDNA levels detected and variation in CMV-DNA <0.5 log10 was observed in the majority of the samples. The viral load kinetic profiles of the assays were overlapping in all patients, but Abbott showed higher sensitivity in samples containing lower amount of DNA. The clinical value of this greater sensitivity requires further investigation

Kinetics of cytomegalovirus and Epstein-Barr virus DNA in whole blood and plasma of kidney transplant recipients: Implications on management strategies

This retrospective multicenter cohort study investigated the kinetics (ascending and descending phases) of cytomegalovirus (CMV) and Epstein-Barr virus (EBV)-DNA in whole blood (WB) and plasma samples collected from adult kidney transplant (KT) recipients. CMV-DNA kinetics according to antiviral therapy were investigated. Three hundred twenty-eight paired samples from 42 episodes of CMV infection and 157 paired samples from 26 episodes of EBV infection were analyzed by a single commercial molecular method approved by regulatory agencies for both matrices. CMV-DNAemia followed different kinetics in WB and plasma. In the descending phase of infection, a slower decay of viral load and a higher percentage of CMV-DNA positive samples were observed in plasma versus WB. In the 72.4% of patients receiving antiviral therapy, monitoring with plasma CMV-DNAemia versus WB CMV-DNAemia could delay treatment interruption by 7-14 days. Discontinuation of therapy based on WB monitoring did not result in relapsed infection in any patients. Highly different EBV-DNA kinetics in WB and plasma were observed due to lower positivity in plasma; EBV positive samples with a quantitative result in both blood compartments were observed in only 11.5% of cases. Our results emphasize the potential role of WB as specimen type for post-KT surveillance of both infections for disease prevention and management

Measles outbreaks in the Emilia-Romagna Region, Italy, during 2016

Background and aim. Despite the availability of a vaccine,measles continues to be endemic in Italy, where an increase of cases was reported during 2016. This study describes the measles outbreaks in Emilia-Romagna Region (ERR), one of the Italian regions mostly affected. Materials and Methods. A total of 101 suspected cases were reported in ERR during 2016. Laboratory diagnosis by serological and/or molecular methods was performed on 142 specimens (78 urine, 19 oral fluid and 45 sera) related to 97 suspected cases. For positive cases, measles virus (MV) strains involved were identified. Results. Among 101 suspected cases, 72 (71.3%) were confirmed. Vaccination status was known for 61 (84.7%) cases, of which 56 (91.8%) were unvaccinated. The highest incidence was found in the age group 15-39 years. In addition, for the 34.7% (25/72) of confirmed cases, the transmission occurred in nosocomial settings, where healthcare workers were involved (60% of cases). Roma/Sinti population were also involved in 12.5% (9/72)or confirmed cases. Both groups are considered hard-to-reach for immunization. The phylogenetic analysis showed circulation of MV strains belonging to genotype B3 and D8 in 45 (80.4%) and 11 cases (19.6%), respectively. In 94.7% of cases, the measles endemic transmission was demonstrated. Conclusions. This data obtained through active surveillance showed the endemic transmission of MV within a population with immunity gaps including healthcare workers (20.8% of confirmed cases), among which the spread of two endemic MV strains was observed

Universal Newborn Screening for Congenital Cytomegalovirus Infection - From Infant to Maternal Infection: A Prospective Multicenter Study

Introduction: Most infants at risk for cytomegalovirus (CMV)-associated sensorineural hearing loss (SNHL) are unrecognized because of the absence of a universal neonatal CMV screening. The search of CMV-DNA by molecular methods in salivary swabs was demonstrated to be a reliable approach. This study describes the results obtained by carrying out a universal screening for congenital CMV (cCMV) infection including all live-born newborns in three Italian sites, as well as the therapeutic interventions and clinical outcome of the CMV-infected neonates. Moreover, CMV maternal infection's characteristics were evaluated. Methods: To confirm or exclude cCMV infection, a CMV-DNA-positive result on a first salivary swab was followed by repeated saliva and urine samples collected within 21 days of age. Breast milk samples were also collected. The search of CMV-DNA was performed with a single automated quantitative commercial real-time PCR assay, regardless of the type of samples used. Results: A total of 3,151 newborns were enrolled; 21 (0.66%) of them were congenitally infected (median saliva viral load at screening, 6.65 [range, 5.03-7.17] log10 IU/ml). Very low/low viral load in screening saliva samples (median value, 1.87 [range, 1.14-2.59] log10 IU/ml) was associated with false-positive results (n = 54; 1.7%). CMV-DNA was detected in almost half of the breast milk samples of mother-infant pairs with a false-positive result, suggesting that contamination from breast milk may not be the only explanation in the study population. cCMV infection confirmation with the search of CMV-DNA in a urine sample proved to be the gold standard strategy, since false-positive results were observed in 4/54 (7.5%) of the repeated saliva samples. Symptomatic cCMV infection was observed in 3/21 (14.3%) infants; notably, one (4.7%) developed moderate unilateral SNHL at 5 months after birth. Finally, two symptomatic cCMV infections were associated with primary maternal infection acquired in the first trimester of gestation; one newborn with severe cCMV symptoms was born to a mother with no CMV checkups in pregnancy. Conclusion: Without universal neonatal CMV screening, some infected infants who develop late neurological sequelae may not be recognized and, consequently, they are not able to benefit early from instrumental and therapeutic interventions to limit and/or treat CMV disease

Pseudozyma aphidis bloodstream infection in a patient with aggressive lymphoma and a history of intravenous drug use: Case report and review of the literature

Pseudozyma aphidis is an environmental fungus which causes opportunistic infections in immunocompromised patients. Here we report the case of a 54-year-old, intravenous drug user woman, newly diagnosed to have an aggressive lymphoma, who developed a bloodstream infection caused by P. aphidis treated successfully with amphotericin-B therapy. The precise identification was assessed by sequencing. We propose to consider intravenous drug use as a risk factor for invasive infections due to this environmental yeast

Fetal Brain Damage in Human Fetuses with Congenital Cytomegalovirus Infection: Histological Features and Viral Tropism

Human cytomegalovirus (HCMV) causes congenital neurological lifelong disabilities. To date, the neuropathogenesis of brain injury related to congenital HCMV (cCMV) infection is poorly understood. This study evaluates the characteristics and pathogenetic mechanisms of encephalic damage in cCMV infection. Ten HCMV-infected human fetuses at 21 weeks of gestation were examined. Specifically, tissues from different brain areas were analyzed by: (i) immunohistochemistry (IHC) to detect HCMV-infected cell distribution, (ii) hematoxylin-eosin staining to evaluate histological damage and (iii) real-time PCR to quantify tissue viral load (HCMV-DNA). The differentiation stage of HCMV-infected neural/neuronal cells was assessed by double IHC to detect simultaneously HCMV-antigens and neural/neuronal markers: nestin (a marker of neural stem/progenitor cells), doublecortin (DCX, marker of cells committed to the neuronal lineage) and neuronal nuclei (NeuN, identifying mature neurons). HCMV-positive cells and viral DNA were found in the brain of 8/10 (80%) fetuses. For these cases, brain damage was classified as mild (n = 4, 50%), moderate (n = 3, 37.5%) and severe (n = 1, 12.5%) based on presence and frequency of pathological findings (necrosis, microglial nodules, microglial activation, astrocytosis, and vascular changes). The highest median HCMV-DNA level was found in the hippocampus (212 copies/5 ng of human DNA [hDNA], range: 10-7,505) as well as the highest mean HCMV-infected cell value (2.9 cells, range: 0-23), followed by that detected in subventricular zone (1.7 cells, range: 0-19). These findings suggested a preferential viral tropism for both neural stem/progenitor cells and neuronal committed cells, residing in these regions, confirmed by the expression of DCX and nestin in 94% and 63.3% of HCMV-positive cells, respectively. NeuN was not found among HCMV-positive cells and was nearly absent in the brain with severe damage, suggesting HCMV does not infect mature neurons and immature neural/neuronal cells do not differentiate into neurons. This could lead to known structural and functional brain defects from cCMV infection

Potential use of artificial intelligence for vaginal swab analysis in the assessment of common genital disorders: a pilot study

Genital disorders, such as vulvo-vaginal candidiasis (VVC), bacterial vaginosis (BV), and aerobic vaginitis (AV), are very common among fertile women and negatively impact their reproductive and relational life. Vaginal culture can help in the diagnostic workflow of these conditions. Recently, culture-based techniques have taken advantages of up-front specimen processing units, which also include a digital imaging system to record images of plates at programmable time points. In this proof-of-concept study, we assessed the characteristics of digital plate images of vaginal swabs plated by WASPLab system into different media, in order to detect microbial growth morphotypes specific for each genital disorder. A total of 104 vaginal specimens were included: 62 cases of normal lactobacilli-dominated flora, 12 of BV, 16 of VVC, and 14 of AV were analysed. Vaginal specimens were plated by WASPLab system into different chromogenic media and blood agar plates. Plate images were taken automatically by the digital imager at 38 h post-inoculation. We found that each genital condition was characterized by specific morphotypes in terms of microbial growth and colony colour, thus allowing the potential use of artificial intelligence not only to assess the presence of specific microbial genera/species but also to 'categorize' peculiar clinical conditions

Multidrug-Resistant Cytomegalovirus Infection in a Patient With Granulomatosis With Polyangiitis During Immunosuppressive Treatment

Cytomegalovirus (CMV) infection is a major complica- tion in immunocompromised patients, including those with autoimmune diseases. Here, we describe the first case of granulomatosis with polyangiitis treated with steroids and cyclophosphamide, complicated by a multi- drug-resistant (MDR) CMV infection in presence of weak antiviral cellular immunity. Since reports regarding CMVinfection in rheumatological patients are rarely described and no guidelines on its management exist, the described case contributes to identify potential strategies to pre- dict the risk of CMV disease and developing of MDR-CMV in these patients, through virological and immunological surveillance

Monitoring of CMV-specific cell-mediated immunity in heart transplant recipients: clinical utility of the QuantiFERON®-CMV assay for the management of post-transplant CMV infection

Background: The clinical utility of QuantiFERON®-CMV (QFN®-CMV) assay in heart transplant recipients was assessed.Methods:Forty-four CMV-seropositive patients were enrolled: 17 received antiviral prophylaxis and 27 were managed pre-emptively. CMV-DNAemia monitoring was performed by a quantitative real-time PCR assay. QFN®-CMV assay was retrospectively performed on blood samples collected at five post-transplant time-points.Results:A higher proportion of patients with indeterminate QFN®-CMV result after the suspension of prophylaxis developed CMV infection compared to patients who showed a global T-cell responsiveness (P=0.036). Patients (42.9%) who reconstituted CMV-specific response following the first CMV-DNAemia showed median CMV-DNAemia peak 1 log of magnitude lower than patients with indeterminate results and all controlled viral replication spontaneously. The 25% of patients with an indeterminate result developed CMV disease.In the pre-emptive strategy group, no difference in the development of subsequent infection, magnitude of viral load and viral control were observed on the basis of QFN®-CMV measurements performed before and after the first CMV-DNAemia, respectively.Considering both CMV prevention strategies, the viral relapse was associated with the failure to reconstitute CMV-specific CMI after the resolution of the first episode of CMV infection (P=0.032).Conclusions:QFN®-CMV measurements can be a useful tool for identifying patients:i) at higher risk of developing infection after discontinuing antiviral prophylaxis;ii) with late CMV infection who would benefit from appropriate antiviral interventions andiii) at higher risk of viral relapses. QFN®-CMV measurements within 1 month post-transplant (early period) are not revealing