A randomized trial of hyperimmune globulin to prevent congenital cytomegalovirus.

BACKGROUND: Congenital infection with human cytomegalovirus (CMV) is a major cause of morbidity and mortality. In an uncontrolled study published in 2005, administration of CMV-specific hyperimmune globulin to pregnant women with primary CMV infection significantly reduced the rate of intrauterine transmission, from 40% to 16%. METHODS: We evaluated the efficacy of hyperimmune globulin in a phase 2, randomized, placebo-controlled, double-blind study. A total of 124 pregnant women with primary CMV infection at 5 to 26 weeks of gestation were randomly assigned within 6 weeks after the presumed onset of infection to receive hyperimmune globulin or placebo every 4 weeks until 36 weeks of gestation or until detection of CMV in amniotic fluid. The primary end point was congenital infection diagnosed at birth or by means of amniocentesis. RESULTS: A total of 123 women could be evaluated in the efficacy analysis (1 woman in the placebo group withdrew). The rate of congenital infection was 30% (18 fetuses or infants of 61 women) in the hyperimmune globulin group and 44% (27 fetuses or infants of 62 women) in the placebo group (a difference of 14 percentage points; 95% confidence interval, -3 to 31; P=0.13). There was no significant difference between the two groups or, within each group, between the women who transmitted the virus and those who did not, with respect to levels of virus-specific antibodies, T-cell-mediated immune response, or viral DNA in the blood. The clinical outcome of congenital infection at birth was similar in the two groups. The number of obstetrical adverse events was higher in the hyperimmune globulin group than in the placebo group (13% vs. 2%). CONCLUSIONS: In this study involving 123 women who could be evaluated, treatment with hyperimmune globulin did not significantly modify the course of primary CMV infection during pregnancy. (Funded by Agenzia Italiana del Farmaco; CHIP ClinicalTrials.gov number, NCT00881517; EudraCT no. 2008-006560-11.).

Maternal, fetal, and neonatal parameters for prognosis and counseling of HCMV congenital infection.

To investigate retrospectively the prognostic significance of maternal, fetal, and neonatal parameters and clinical outcome in 150 HCMV congenital infections during the period 1995-2009. HCMV fetal infection was investigated in amniotic fluid and fetal blood samples. HCMV congenital infection was confirmed in newborn urine and blood samples. Symptomatic infection was defined in HCMV-infected fetuses and in infected newborns on the basis of physical and instrumental findings. Follow-up at 3, 6, 12 months, and then annually up to school age, included clinical evaluation, funduscopic, audiologic, neurologic, and cognitive assessment. Overall, 122/150 (81.3%) newborns were asymptomatic and 28/150 (18.7%) were symptomatic at birth. The best prognostic maternal parameter of symptomatic infection at birth was gestational age at infection (P = 0.037). The best fetal virological markers were HCMV DNA levels in amniotic fluid (P < 0.001), antigenaemia levels (P = 0.007), HCMV DNA levels in blood (P = 0.004), and HCMV-specific IgM index values (P = 0.002). The only significant neonatal parameter was HCMV DNA level in blood [P = 0.006; OR, 3.62 (95% CI, 1.46-8.97)]. Symptoms at birth correlated significantly with long-term sequelae (P = 0.021). A trend towards a risk of sequelae in early (n = 15/58 examined) versus late (n = 6/57 examined) maternal infection was documented. The risk of symptomatic congenital infection at birth increased linearly with the number of significant maternal, fetal, and neonatal parameters.

Slow increase in IgG avidity correlates with prevention of human cytomegalovirus transmission to the fetus.

Following primary human cytomegalovirus (HCMV) infection, virus-specific IgG antibody shift from low to high avidity with individual variations in the rate of avidity maturation. The kinetics of the avidity maturation of IgG specific for HCMV nuclear antigen in pregnant women with primary infection was investigated. Absorbance values used for avidity index calculation of 286 sequential sera collected from 69 pregnant women with primary HCMV infection were retrieved. Percent difference in absorbance values of IgG antibody bound to the solid phase after urea treatment (IgG avidity) between early (T1, 0-90, median 31 days) and late (T2, 91-180, median 136 days) serum samples was calculated for each woman. Three groups of women were identified: 24/69 (34.8%) women showed high (>100%) avidity increase between T1 and T2 (pattern H), 29/69 (42%) low (<50%) increase (pattern L), and 16/69 (23.2%) intermediate increase (pattern I). Avidity values in T1 samples were significantly higher in women with pattern L compared to women with pattern H (P=0.01). Altogether, 28/69 (40.6%) women transmitted HCMV infection to their fetuses. Fetal infection preferentially occurred (P<0.01) in women with pattern H (15/24, 62.5%) compared with women with pattern L (7/29, 24.1%). In conclusion, different patterns of IgG avidity maturation can be detected following primary HCMV infection. Pregnant women with pattern H (rapid IgG avidity increase) appear to be at higher risk for fetal infection, whereas, pregnant women developing early antibody with high avidity appear to be at a lower risk of vertical transmission.

Differential outcome of neurological HCMV infection in two hematopoietic stem cell transplant recipients.

BACKGROUND: Human cytomegalovirus (HCMV) infection of the central nervous system (CNS) is a rare but life threatening condition which may follow hematopoietic stem cell transplantation. Diagnosis, monitoring and treatment approaches rely on anecdotal reports. CASE PRESENTATIONS: The different outcomes of HCMV CNS disease in an adult and a pediatric T-cell depleted hematopoietic stem cell transplant (HSCT) recipient are reported. In the first case, HCMV encephalitis emerged in the context of simultaneous impairment of the T- and B-cell immunity. Antiviral treatment only reduced viral load in peripheral blood and the patient died. In the second case, an HCMV radiculopathy was observed and antiviral treatment was adjusted on the basis of intrathecal drug level. In addition, donor HCMV-specific cytotoxic T lymphocytes (CTLs) were infused. Viral load in the CNS decreased and the patient recovered from the acute event. In neither case were drug-resistant HCMV variants observed in blood or CNS samples. CONCLUSIONS: T-cell depleted HSCT appears a predisposing condition for CNS HCMV infection since never observed in other HSCT recipients at our center in the last 15 years. Intensive diagnostic approaches and timely aggressive combination treatments might improve clinical outcome in these patients.

The epidemiology of central venous catheter-related bloodstream infection in our renal units is changing.

Recent reports have shown an increase in the rate of Gram-negative bacteremia in several settings, including catheter-related bloodstream infections (CRBSI). To analyze if the epidemiology of CRBSI is also changing in hemodialysis patients, we revisited the etiology of CRBSIs in our renal unit over 8 years. During the observed periods, 149 episodes of CRBSIs were reported and the CRBSI incidence rate, ranged between 0.67 and 0.82 episodes/1000 tCVC days. Of these 149 episodes, 84 (56.3%) were due to Gram-positive bacteria, 62 (41.6%) to Gram-negative bacteria, and 3 (2.1%) to polymicrobial flora, no episodes of fungi were found. There was a trend, but not statistically significative, increase over time in the number of Gram-negative CRBSIs among the total CRBSIs, rising from 37.8% in the first period to 41.2% in the second period and to 44.3% in the last period, with a parallel decrease in the percentage of Gram-positive CRBSIs (from 59.5% to 56.9% and subsequently to 54.1%). Between Gram-negative, we reported an intensification of CRBSI due to Enterobacterales, particularly Escherichia coli. Among the Gram-negative, we have isolated germs rarely reported in the literature, such as Burkholderia cepacia, Pantoea agglomerans, and Rhizobium radiobacter. Regarding Gram-positive bacteria, a triplicate incidence of Staphylococcus aureus was reported with MRSA accounting for 42% in the third period. Among the Gram-positive bacteria, we reported two episodes of Kocuria kristinae and two of Bacillus spp.Our data demonstrated that the epidemiology of CRBSI in the same center, will change over time and Gram-negative strains are an increasing cause of CRBSI. The limitation of the present report is that statistical significance has not been reached, probably due to the limited number of CRBSI. New bacteria, both Gram-negative and Gram-positive, are emerging. Collaboration with the Microbiology Department appears essential to an appropriate diagnosis.

Post-transplant lymphoproliferative disorders and Epstein-Barr virus DNAemia in a cohort of lung transplant recipients.

BACKGROUND: Post-transplant lymphoproliferative disorders (PTLD) are serious complications in lung transplant recipients. No consensus on EBV DNAemia levels predictive of PTLD has been reached. In addition, in many instances EBV DNAemia is determined in patients with suggestive symptoms only. METHODS: The characteristics of five patients with PTLD as well as the prevalence of EBV DNAmia in a cohort of 137 consecutive patients receiving lung transplantation are described. RESULTS: Twenty-six out of 137 patients (18.9%) were excluded from the analysis because lost at follow-up or dead from PTLD-independent reasons within three months of transplantation. EBV DNA in peripheral blood mononuclear cells (PBMC) was determined in 83/111 patients (74.8%) because of potential PTLD-related symptoms, while 28 patients (25.2%) showed no symptoms and were not examined. EBV DNAemia was positive in 53/83 patients (63.8%), and negative in 30/83 patients (36.2%). PTLD was diagnosed in five (4.5%) patients at a median time of 270 (range 120-870) days following transplantation. All five PTLD (three large B-cell lymphomas, one Hodgkin lymphoma and one possible pre-neoplastic lesion) were potentially associated with EBV infection. However, only 3/5 patients with PTLD had detectable EBV DNAemia: < 1,000 copies EBV DNA/1 × 105 PBMC in one patient and > 1,000 copies EBV DNA/1 × 105 PBMC in two patients. CONCLUSION: A systematic multidisciplinary (clinical, radiologic, virologic and histologic) approach is mandatory for the diagnosis and management of PTLD in lung transplant recipients, while monitoring of symptomatic patients only may provide an incomplete or late picture of the clinical problem. In addition, staining for EBV antigens and quantification of EBV DNA in biopsy specimens should always be performed to understand the role of EBV infection in the pathogenesis of PTLD.

Human cytomegalovirus-specific CD4(+) and CD8(+) T-cell response determination: comparison of short-term (24h) assays vs long-term (7-day) infected dendritic cell assay in the immunocompetent and the immunocompromised host.

Human cytomegalovirus (HCMV)-specific CD4(+) and CD8(+) T-cells were measured in the immunocompetent host as well as in 13 solid-organ transplant recipients (SOTR), and 12 young hematopoietic stem cell transplant recipients (HSCTR) by using a long-term (7-day) assay based on PBMC stimulation by HCMV-infected dendritic cells (iDC), and two short-term (24h) assays, one for CD4(+) stimulation by infected cell lysate (iCL), and the other for CD8(+) stimulation by a pool of 34 epitopic peptides (pep-pool). In the immunocompetent, the number of T-cells activated by either iCL or the pep-pool was significantly reduced with respect to iDC. In both SOTR and HSCTR, the number of T-cells activated by iDC was comparable to that activated by iCL or the pep-pool. A significant correlation between iDC-activated T-cells and T-cells activated by either iCL or the pep-pool was observed. In conclusion, whenever a rapid result is needed, short-term assays may efficiently replace the iDC assay.

High success rate in salvage of catheter-related bloodstream infections due to Staphylococcus aureus, on behalf of project group of Italian society of nephrology.

BACKGROUND: Catheter-related bloodstream infections caused by Staphylococcus aureus represent one of the most fearful infections in chronic haemodialysis patients with tunnelled central venous catheters. Current guidelines suggest prompt catheter removal in patients with positive blood cultures for S. aureus. This manoeuvre requires inserting a new catheter into the same vein or another one and is not without its risks. METHODS: A protocol based on early, prompt diagnosis and treatment has been utilized in our renal unit since 2012 in an attempt to salvage infected tunnelled central venous catheters. We prospectively observed 247 tunnelled central venous catheters in 173 haemodialysis patients involving 167,511 catheter days. RESULTS: We identified 113 catheter-related bloodstream infections (0.67 episodes per 1000 days/tunnelled central venous catheter). Forty were caused by S. aureus, including 19 by methicillin-resistant S. aureus (79% saved) and 21 by methicillin-sensitive S. aureus (90% saved), of which 34 (85%) were treated successfully. Eight recurrences occurred and six (75%) were successfully treated. A greater than 12 h time to blood culture positivity for S. aureus was a good prognostic index for successful therapy and tunnelled central venous catheter rescue. CONCLUSION: Our data lead us to believe that it is possible to successfully treat catheter-related bloodstream infection caused by S. aureus and to avoid removing the tunnelled central venous catheter in many more cases than what has been reported in the literature. On the third day, it is mandatory to decide whether to replace the tunnelled central venous catheter or to carry on with antibiotic therapy. Apyrexia and amelioration of laboratory parameters suggest continuing systemic and antibiotic lock therapy for no less than 4 weeks, otherwise, tunnelled central venous catheter removal is recommended.

Complete Genome and Plasmids Sequences of a Clinical Proteus mirabilis Isolate Producing Plasmid Mediated NDM-1 from Italy.

Background: The spread of carbapenemase genes, such as blaNDM-1, in Proteus mirabilis poses a public health threat. The aim of the study was to characterize the genome and plasmids sequences of an NDM-1-positive strain (IBCRE14), which was isolated in 2019 from a catheterized patient hospitalized in Italy. Methods: Whole genome sequencing (WGS) of IBCRE14 was performed on extracted genomic DNA using Sequel I platform. Genome assembly was performed using "Microbial Assembly". Genomic analysis was conducted by uploading the contigs to ResFinder and PlasmidFinder databases from the Center for Genomic Epidemiology. Results: IBCRE14 had a genome size of 4,018,329 bp and harboured genes coding for resistance to aminoglycosides (aadA1), phenicol (cat), tetracycline (tetJ), and trimethoprim (dfrA1). A large plasmid (pIB_NDM_1) harboured antibiotic resistance genes against sulphonamide (sul1), trimethoprim (dfrA14), tetracycline (tetB), rifampicin (arr-2), aminoglycosides (aadA1, aph3-VI), and beta-lactams (blaOXA-10, blaNDM-1). Furthermore, a small plasmid (pIB_COL3M) harboured a qnrD1 gene coding for quinolone resistance. Conclusion: The ability to conjugate and the presence of a composite antibiotic resistance island suggests that pIB_NDM_1 could both acquire more resistance genes and easily disseminate. To our knowledge, this is the first report on an untypable plasmid harbouring blaNDM-1 in P. mirabilis, in Italy.

Surveillance of influenza virus B circulation in Northern Italy: summer-2008 isolation of three strains and phylogenetic analysis.

Influenza virus type B strains were unexpectedly detected and isolated in Italy during summer-fall 2008 from three patients travelling to Italy from Lebanon, Senegal and Uzbekistan. The three summer-fall strains matched to a high degree the hemagglutinin (HA1) of influenza virus type B strains circulating in Italy in the second part (January through April) of the 2007/2008 season, and HA1 of the type B strains included in the 2008/2009 vaccine (B/Yamagata/16/88 lineage). Surveillance of influenza virus circulation in Western countries also during the summer-fall season may help to trace and anticipate the appearance of new influenza virus variants.

Rapid typing, subtyping and RNA quantification of influenza virus type A strains in respiratory secretions.

During the winter-spring season 2006-2007, 38 influenza virus strains were identified in patients admitted to hospital with an acute respiratory tract infection. Infections were diagnosed in parallel by direct fluorescent antibody (DFA) staining using type-specific monoclonal antibodies and real-time reverse transcription (RT)-PCR targeting the gene M (nt 25-124). In addition, virus strains were isolated in MDCK cells. Overall, 37 influenza virus strains were type A, and one type B. Of these, 35 (80.4%) were detected and typed by real-time RT-PCR, 34 (80.1%) by DFA, and 27 (71.0%) by virus isolation. Subtyping of 37 influenza virus A strains by RT-PCR and DFA gave the following results: 4/6 H1 strains were correctly subtyped by both methods, while of the 29 H3 strains subtyped by RT-PCR 7 were missed by DFA. Thus, the overall concordance of the two subtyping methods was 28/37 (75.7%). Viral RNA quantification by real-time PCR showed that when respiratory secretion collection was done within 5 days after the onset of symptoms, viral load was greater than 1 x 10(6) RNA copies/ml. In conclusion, typing and subtyping of influenza virus type A strains may benefit from both MAbs and RT-PCR, while viral RNA quantification may provide an indication of symptom onset.

Cytomegalovirus DNAemia in pregnant women.

BACKGROUND: Cytomegalovirus (CMV) transmission from mother to fetus occurs at a much greater rate following primary rather than reactivated infections and CMV dissemination in the mother is considered a key step in the pathogenesis of fetal infection. However, knowledge of CMV DNAemia in CMV-seropositive pregnant women is very limited. OBJECTIVE: Major objective of this study was to assess the prevalence and diagnostic value of CMV DNAemia in a large population of seropositive pregnant women. STUDY DESIGN: Serologic and DNAemia results obtained from 2211 blood samples of 1371 consecutive pregnant women referred to our Institution for suspected CMV infection in the period 2001-2010 were reviewed. RESULTS: DNAemia was detected in 452/597 (75.7%) women with serologic evidence of primary CMV infection and in 4/774 (0.5%) women without evidence of primary infection. CONCLUSION: In pregnant women, CMV DNAemia is detected primarily during primary infection. CMV DNAemia determination may be helpful in the diagnosis of primary infection.

Kinetics of human cytomegalovirus (HCMV) DNAemia in transplanted patients expressed in international units as determined with the Abbott RealTime CMV assay and an in-house assay.

BACKGROUND: Consensus human cytomegalovirus (HCMV) DNA cut-off values for preemptive therapy in transplant recipients have not yet been defined, mainly due to the lack of real-time PCR standardization. OBJECTIVES: (i) To compare the kinetics of HCMV DNA in transplanted patients using an in-house real-time PCR assay (Pavia assay) and the new Abbott RealTime CMV assay; (ii) to verify assay concordance in the identification of patients eligible for preemptive treatment and (iii) standardize results with international units (IUs) using the WHO International HCMV DNA Standard as a reference. STUDY DESIGN: The kinetics of HCMV disseminated infection was retrospectively evaluated in 513 stored whole blood samples from 37 transplanted patients enrolled in randomized prospective studies designed for the clinical validation of HCMV DNA cut-off values. Conversion factors of HCMV DNA copy number to WHO international units (IUs) were determined. RESULTS: Among the 513 samples, 352 (68.6%) were concordant positive, 42 (8.1%) concordant negative and 119 (23.1%) discordant. All discordant samples resulted positive by the Abbott RealTime CMV assay and negative by the Pavia assay, showing higher sensitivity for the Abbott RealTime CMV assay. A significant correlation was observed between concordant positive samples (r=0.89). HCMV DNAemia determined by each assay showed overlapping kinetics. Expression of results as IU/ml provided the best results in preemptive treatment simulation. CONCLUSION: HCMV DNAemia cut-off values determined using our in-house assay and expressed as IU/ml appear valid for use in commercial assays as well as other potential in-house assays.

Monitoring of human cytomegalovirus and virus-specific T-cell response in young patients receiving allogeneic hematopoietic stem cell transplantation.

In allogeneic hematopoietic stem-cell transplantation (HSCT) recipients, outcome of human cytomegalovirus (HCMV) infection results from balance between viral load/replication and pathogen-specific T-cell response. Using a cut-off of 30,000 HCMV DNA copies/ml blood for pre-emptive therapy and cut-offs of 1 and 3 virus-specific CD4(+) and CD8(+) T cells/µl blood for T-cell protection, we conducted in 131 young patients a prospective 3-year study aimed at verifying whether achievement of such immunological cut-offs protects from HCMV disease. In the first three months after transplantation, 55/89 (62%) HCMV-seropositive patients had infection and 36/55 (65%) were treated pre-emptively, whereas only 7/42 (17%) HCMV-seronegative patients developed infection and 3/7 (43%) were treated. After 12 months, 76 HCMV-seropositive and 9 HCMV-seronegative patients (cumulative incidence: 90% and 21%, respectively) displayed protective HCMV-specific immunity. Eighty of these 85 (95%) patients showed spontaneous control of HCMV infection without additional treatment. Five patients after reaching protective T-cell levels needed pre-emptive therapy, because they developed graft-versus-host disease (GvHD). HSCT recipients reconstituting protective levels of HCMV-specific T-cells in the absence of GvHD are no longer at risk for HCMV disease, at least within 3 years after transplantation. The decision to treat HCMV infection in young HSCT recipients may be taken by combining virological and immunological findings.

Role of prenatal diagnosis and counseling in the management of 735 pregnancies complicated by primary human cytomegalovirus infection: a 20-year experience.

BACKGROUND: The burden of congenital human cytomegalovirus (HCMV) infection is well recognized. However, screening for maternal infection remains controversial in view of diagnostic challenges, counseling difficulties, and absence of medical treatment. OBJECTIVE: To assess the role of prenatal diagnosis and counseling in the management of pregnancy complicated by primary HCMV infection. STUDY DESIGN: Retrospective study aimed at investigating diagnostic features, options, and pregnancy outcome in 735 women with primary HCMV infection over a period of 20 years (1990-2009). RESULTS: Overall, 25.6% women were found to be seronegative before the actual pregnancy. However, none were informed about HCMV infection and potential prevention strategies. Diagnosis of primary HCMV infection was achieved by seroconversion in 44.4% cases and by different combinations of virus-specific IgM, low IgG avidity, and DNAemia in 43.9% cases. Non-specific symptoms and/or haematological/biochemical alterations were recalled by 73.5% women. The onset of infection could be established, and counseling adjusted accordingly in >90% cases. The overall rate of vertical transmission was 37.1%, ranging from 5.6% for preconceptional infections to 64.1% for third trimester infections. Amniocentesis was chosen by 43.1% women, whereas pregnancy termination was requested by 15.6%. CONCLUSIONS: Reference virology centers and ad hoc trained and experienced physicians are required for accurate diagnosis of primary infection in pregnancy and ensuing counseling. Prenatal diagnosis has a central role in the management of pregnancies complicated by primary HCMV infection. HCMV-seronegative women should receive adequate information.