Molecular epidemiology of methicillin-resistant Staphylococcus aureus from bacteraemia in northern Italy.

BACKGROUND: Vancomycin is frequently used in the treatment of methicillin-resistant Staphylococcus aureus (MRSA) bacteraemia; reduced susceptibility to vancomycin is therefore disturbing. METHODS: molecular epidemiological analysis of 81 MRSA bacteraemia isolates collected during 2002-10 in the province of Bolzano, northern Italy was performed. MICs of a range of antimicrobials were determined by agar microdilution, screening for hGISA was by Macro-Etest and Etest GRD and confirmed by PAP-AUC with vancomycin and teicoplanin. All isolates were characterised by toxin gene profiling, agr, spa, and SCCmec-typing; MLST and PFGE were carried out on representative strains. RESULTS: The dominant clones identified were ST8-MRSA-IVc (55%) and ST228- and ST111-MRSA-I (25%); most of the latter two lineages (19/20; 95%) were GISA or PAP-AUC confirmed hGISA. One ST8-MRSA-IVc isolate harboured ccrA2B2 together with ccrA4B4. The remainder were diverse genotypically and belonged to MLST clonal complexes 1, 22, 45 and 398. CONCLUSIONS: Diverse lineages of MRSA were identified as causing bacteraemia in a province in northern Italy. The association of a specific genotype with the hGISA and GISA phenotypes among representatives of the second most common lineage identified is of clinical concern.

Investigation of bacterial and viral agents and immune status in Behcet's disease patients from Iran.

AIM: Behçet's disease (BD) is an autoimmune disorder associated with HLA-B51 positivity. Serologic/genomic findings have suggested microbes as possible causative agents and the geographical distribution suggests environmental influences. METHODS: We performed comparative analyses of 40 patients with BD or related symptoms not fulfilling BD criteria. Patients originating from different regions of Iran were tested by molecular/serological methods for human herpes viruses and parvovirus B19, two Chlamydiae species, as well as Coxiella, Listeria, Yersinia, Leptospira and Mycobacterium paratuberculosis. Human leukocyte antigen-typing was performed: testing of cytokine profiles and immune mediators representative for the cellular immune system, including neopterin/kynurenine production. RESULTS: No apparent differences in interleukin (IL)-4, 6, 8 and 10 were observed, whereas production of soluble IL-2-receptor and tumor necrosis factor (TNF)-alpha were more pronounced in the BD group. Neopterin/kynurenine production was comparable, although both groups showed twice the levels of healthy people. No significant differences of herpes simplex virus (HSV) antibody titres were observed but higher titres against Chlamydophila pneumoniae were found in the controls. In 20 BD patients and controls neither parvovirus B19 DNA was detected nor bacterial DNA. Viral DNA of Epstein-Barr virus (EBV), cytomegalovirus (CMV) and human herpes virus (HHV)8 was detected more frequently in the BD group, whereas HSV DNA was only found in the controls, indicating that stomatitis might be caused by HSV. CONCLUSION: Although no significant association of BD was detected with a single pathogen, our findings suggest that detection of HSV DNA or Chlamydiae would rather argue against classic BD. Whether there is a discriminative potential of the tested immune mediators/receptors has to be elucidated in further studies.

IFN-gamma mediated pathways in patients with fatigue and chronic active Epstein Barr virus-infection.

BACKGROUND: Chronic active Epstein Barr virus (EBV)-infection is characterized by mononucleosis like symptoms including fatigue, lymphadenopathy and/or hepatosplenomegaly and serologic evidence for ongoing EBV replication. Interferon-gamma (IFN-gamma) triggers several antiviral mechanisms in target cells including the induction of indoleamine-2,3-dioxygenase (IDO), which degrades the essential amino acid tryptophan to kynurenine. Because tryptophan is a precursor of the neurotransmitter 5-hydroxytryptamine (serotonin), tryptophan depletion by IDO can cause mood disturbances in patients with chronic immune activation. METHODS: This study investigated the tryptophan metabolism in 20 patients with chronic active EBV-infection, who were followed up for 4 to 8 months and in 10 healthy age-matched controls. The clinical suspicion of chronic active EBV infection was verified by the presence of circulating antibodies against EBV early antigen (EA) and virus capsid antigen (VCA). RESULTS: Patients with detectable EBV-DNA had higher serum neopterin (p<0.01) and lower tryptophan concentrations (p=0.01) than EBV-DNA negative patients. Serum concentrations of neopterin, indicating Th-1 mediated immune activation via IFN-gamma, were positively correlated to enhanced tryptophan degradation (rs=0.650, p<0.001) in patients, but not in healthy individuals. Patients suffering from more severe symptoms (as assessed by questionnaires) tended to have aggravated tryptophan degradation. CONCLUSION: Our data show that EBV viremia is associated with cell-mediated immune activation and increased tryptophan degradation, which may partly account for the symptoms found in this disorder.

Local administration of cidofovir for human papilloma virus associated skin lesions in transplant recipients.

Human papilloma virus (HPV)-associated diseases are increasingly diagnosed in solid organ recipients. Cidofovir (CDV) is a broad-spectrum antiviral agent with activity against all human herpes viruses and HPV. From 2000-2004, a total of 1303 solid organ transplants (SOT) were performed at our center. Six transplant recipients were treated with topical CDV for HPV-associated lesions. One cardiac recipient responded to a single injection of CDV into his recurrent anal condylomata. In a renal recipient with recurrent penile condylomata CDV was injected into the lesions four times (2 week interval) until lesions regressed. One renal recipient developed multiple vaginal and anal intradermal neoplasias, which relapsed after laser ablation. The lesions were repeatedly injected with CDV and completely disappeared. Two renal recipients with widespread verrucae vulgares were treated with CDV gel, which resulted in regression of the lesions. One patient developed donor derived verrucae vulgares on both transplanted hands, which responded to CDV gel. Four of the six patients were switched from calcineurin inhibitors (CNIs) to Sirolimus (SIR). CDV was found effective in the treatment of HPV-associated skin lesions in SOT recipients. It needs to be determined whether switch from CNIs to SIR might have contributed to the beneficial effect of CDV.

Functional T-cell anergy in a case of persistent polyclonal B-cell lymphocytosis.

The T-cell population of a patient with persistent polyclonal B-cell lymphocytosis (PPBL) presenting with an intermittent Epstein-Barr virus (EBV)-associated disease was studied. Unstimulated T-cells did not express CD40 ligand (CD40L), whereas activation with IL-2 led to expression of this costimulatory molecule. CD40L expression was inhibited upon incubation with the supernatant of an EBV-positive B-cell line (SM) which had been grown spontaneously from the patient's peripheral blood cells. The supernatant of SM cells effectively inhibited cytotoxic T-cells. Elevated levels of IL-10, TNF-alpha and soluble CD40 were found in the supernatant of SM cells. Additionally, enhanced levels of LMP-1 protein were detected.

Tolerability and efficacy of N-chlorotaurine in epidemic keratoconjunctivitis--a double-blind, randomized, phase-2 clinical trial.

The aim of this study was to assess the tolerability and efficacy of N-chlorotaurine (NCT), an endogenous antimicrobial agent, in epidemic keratoconjunctivitis. In a prospective double-blind, randomized phase 2b study, the infected eyes were treated for 7 days with eye drops containing 1% aqueous solution of N-chlorotaurine (33 subjects) or gentamicin (27 subjects, control group). Adenovirus types 3, 4, 8, 19, and 37 were detected in 39 subjects (65%), enteroviruses in 8 (13.3%), and staphylococci in 5 (8.3%). Subjective and objective symptoms were scaled and added to a subjective and objective score, respectively, on day 1 (baseline), day 4, and day 8. Analyzing the whole study population, the subjective score on day 8 was lower in the NCT group (P = 0.016), whereas there were no differences in the objective score. However, in severe infections caused by adenovirus type 8 (n = 20) both the subjective and objective score were lower in the NCT group on day 4 (P = 0.003 and 0.015, respectively), which was also true for the subjective score on day 8 (P = 0.004) in this subgroup. The frequency of subepithelial infiltrates was similar in both groups. N-chlorotaurine was well-tolerated, shortened the duration of illness, and seems to be a useful causative therapeutic approach in severe epidemic keratoconjunctivitis.

Functional analysis of the mutated Epstein-Barr virus oncoprotein LMP1(69del): implications for a new role of naturally occurring LMP1 variants.

BACKGROUND AND OBJECTIVES: The role of carboxyterminal deletions of the latent membrane protein-1 (LMP1) in Epstein-Barr virus (EBV) infection and oncogenesis is unclear. Here we describe functional properties of a rare 69-bp LMP1 deletion mutant (LMP1(69del)) isolated from a patient with polyclonal B-cell lymphocytosis. DESIGN AND METHODS: Colony focus assay was used to evaluate the transforming capacity of LMP1(69del) in comparison to that of wild-type LMP1 from EBV strain B95/8. Transient transfectants of B-, T-, epithelial and 3T3 cells, and stable transfectants with ecdysone-inducible LMP1 expression were produced. The signaling capacity of both LMP1s on nuclear transcription factors NFkappaB and AP-1 were studied. Secretion of matrix metalloproteinase MMP-9, apoptosis, and EBV lytic and latent gene expression were also investigated. RESULTS: LMP(69del) showed transforming properties comparable to those of the wild-type oncoprotein. Induction of NFkappaB but a markedly reduced influence on AP-1 were observed. Both oncoproteins induced secretion of MMP-9, and enhanced pre-apoptotic effects in Jurkat-T cells leading to increased Fas/Apo-1 and doxorubicin-mediated apoptosis. Furthermore, LMP1(69del) showed a more effective down-regulation of the EBV lytic cycle master gene BZLF1(Zebra) than did wild-type LMP1. INTERPRETATION AND CONCLUSIONS: (i) LMP1(69del) possesses oncogenic properties, (ii) the observed impaired activity on AP-1 does not interfere with MMP-9 induction, (iii) the enhanced inhibition of BZLF1 could compensate for previously described mutations of our isolate leading to a more lytic phenotype and may be responsible for counteracting permanent virus replication in the chronic active EBV syndrome observed in this patient.

Risk for cytomegalovirus infection following reduced intensity allogeneic stem cell transplantation.

Preliminary data suggest a faster immune recovery following non-myeloablative stem cell transplantation because of the persistence of recipient T cells, but the real impact on post-transplant infectious complications remains unknown. We retrospectively analysed the incidence of cytomegalovirus (CMV) infection in twenty patients following reduced intensity conditioning with busulfan/fludarabine+/-thiotepa and post-transplant immunosuppression with cyclosporine A/mycophenolate mofetil. Results were compared with 20 patients receiving myeloablative transplants during the same time period and who were matched for CMV risk group and for donor origin. The cumulative incidence of CMV infection following reduced intensity vs. myeloablative transplants was 60.4% vs. 40.0%, respectively (p value 0.1, log rank test). The risk for CMV infection in both cohorts was increased after in vivo T cell depletion with antithymocyte globulin (75% and 60%, respectively). Acute GVHD preceded the diagnosis of CMV infection by a median of 25 (range, 9-61) days following reduced intensity transplants and a median of 14 (range, 10-34) days in myeloablative transplants. Recurrent CMV infections were observed only in patients receiving reduced intensity transplants. Using multivariate analysis only reduced intensity transplantation and in vivo T cell depletion had a significant impact on the risk of CMV infection. In our series the incidence for CMV infection following reduced intensity transplants seems to be increased as compared with risk-matched myeloablative transplants. When adding anti-T cell antibodies to the conditioning regimen, the risk for CMV infection increases by up to 75%. Thorough studies of the risk of post-transplant viral infection are necessary to optimize surveillance as well as pre-emptive and/or prophylactic treatment strategies in the non-myeloablative transplantation setting.

Human parvovirus Bb19 detection in asymptomatic blood donors: association with increased neopterin concentrations.

Serum neopterin concentrations were determined in 20,000 blood donations. For the 400 donations with neopterin concentrations above the 98 th percentile and another 1200 donations with neopterin concentrations in the lower range, results of human parvovirus (HPV) B19 tests were compared. Infectious specimens were identified by dot blot hybridization assay and polymerase chain reaction (PCR) that used the outer primers and detected 1 pg of HPV B19 DNA, corresponding to approximately 10(5) copies of the genome, in the specimens and by a nested PCR that detected 1-10 fg of DNA, corresponding to 10(2)-10(3) copies of the genome. Of 400 specimens with neopterin concentrations > or =12 nmol/L (98th percentile, current cutoff), 10 tested positive by dot blot hybridization assay (9 of these were confirmed by nested PCR). Among 1200 specimens with low neopterin concentrations (<12 nmol/L), no specimen containing HPV B19 DNA was detected. These findings suggest an association between elevated neopterin concentrations and HPV B19 infectivity.

Fatal infection of a pet monkey with Human herpesvirus.

Concerns have been raised about pet monkeys as a potential threat to humans. We report the opposite situation, a danger to pets that arises from humans. Similar to herpesvirus B (Cercopithecine herpesvirus 1), which endangers humans but not its host species, Human herpesvirus 1 can act as a "killer virus" when crossing the species barrier to New World monkeys.