Beyond appearance: An unusual manifestation of isolated oral secondary syphilis

Syphilis is a sexually acquired chronic infection caused by Treponema pallidum and is characterized by a variety of clinical manifestations. The secondary stage of the disease results from the hematogenous and lymphatic dissemination of treponemes after a few weeks or months, and it is characterized by recurrent activity of the disease, with muco-cutaneous as well as systemic manifestations. Mucosal lesions range from small, superficial ulcers that resemble painless aphthae to large gray plaques, and they are generally associated with systemic manifestations of the disease. The exclusive asymptomatic oral localization not associated with general manifestations is uncommon but may actually be unrecognized and under-reported. We report a case of isolated oral manifestation as the unique presentation of secondary syphilis

Merkel cell polyomavirus (MCPyV) in the context of Immunosuppression. Genetic analysis of noncoding control region (NCCR) variability among a HIV-1-positive population

Background: Since limited data are available about the prevalence of Merkel cell polyomavirus (MCPyV) and the genetic variability of its noncoding control region (NCCR) in the context of immunosuppression, this study aimed to investigate the distribution of MCPyV in anatomical sites other than the skin and the behavior of NCCR among an HIV-1-positive population. Methods: Urine, plasma, and rectal swabs specimens from a cohort of 66 HIV-1-positive patients were collected and subjected to quantitative real-time polymerase chain reaction (qPCR) for MCPyV DNA detection. MCPyV-positive samples were amplified by nested PCR targeting the NCCR, and NCCRs alignment was carried out to evaluate the occurrence of mutations and to identify putative binding sites for cellular factors. Results: MCPyV DNA was detected in 10/66 urine, in 7/66 plasma, and in 23/66 rectal samples, with a median value of 5 × 102 copies/mL, 1.5 × 102 copies/mL, and 2.3 × 103 copies/mL, respectively. NCCR sequence analysis revealed a high degree of homology with the MCC350 reference strain in urine, whereas transitions, transversions, and single or double deletions were observed in plasma and rectal swabs. In these latter samples, representative GTT and GTTGA insertions were also observed. Search for putative binding sites of cellular transcription factors showed that in several strains, deletions, insertions, or single base substitutions altered the NCCR canonical configuration. Conclusions: Sequencing analysis revealed the presence of numerous mutations in the NCCR, including insertions and deletions. Whether these mutations may have an impact on the pathogenic features of the virus remains to be determined. qPCR measured on average a low viral load in the specimens analyzed, with the exception of those with the GTTGA insertion

COS-7-based model: methodological approach to study John Cunningham virus replication cycle

John Cunningham virus (JCV) is a human neurotropic polyomavirus whose replication in the Central Nervous System (SNC) induces the fatal demyelinating disease, progressive multifocal leukoencephalopathy (PML). JCV propagation and PML investigation have been severely hampered by the lack of an animal model and cell culture systems to propagate JCV have been very limited in their availability and robustness. We previously confirmed that JCV CY strain efficiently replicated in COS-7 cells as demonstrated by the progressive increase of viral load by quantitative PCR (Q-PCR) during the time of transfection and that archetypal regulatory structure was maintained, although two characteristic point mutations were detected during the viral cycle. This short report is an important extension of our previous efforts in defining our reliable model culture system able to support a productive JCV infection. Supernatants collected from transfected cells have been used to infect freshly seeded COS-7 cell line. An infectious viral progeny was obtained as confirmed by Western blot and immunofluorescence assay. During infection, the archetype regulatory region was conserved. Importantly, in this study we developed an improved culture system to obtain a large scale production of JC virus in order to study the genetic features, the biology and the pathogenic mechanisms of JC virus that induce PML

COS-7-based model: methodological approach to study John Cunningham virus replication cycle

John Cunningham virus (JCV) is a human neurotropic polyomavirus whose replication in the Central Nervous System (SNC) induces the fatal demyelinating disease, progressive multifocal leukoencephalopathy (PML). JCV propagation and PML investigation have been severely hampered by the lack of an animal model and cell culture systems to propagate JCV have been very limited in their availability and robustness. We previously confirmed that JCV CY strain efficiently replicated in COS-7 cells as demonstrated by the progressive increase of viral load by quantitative PCR (Q-PCR) during the time of transfection and that archetypal regulatory structure was maintained, although two characteristic point mutations were detected during the viral cycle. This short report is an important extension of our previous efforts in defining our reliable model culture system able to support a productive JCV infection. Supernatants collected from transfected cells have been used to infect freshly seeded COS-7 cell line. An infectious viral progeny was obtained as confirmed by Western blot and immunofluorescence assay. During infection, the archetype regulatory region was conserved. Importantly, in this study we developed an improved culture system to obtain a large scale production of JC virus in order to study the genetic features, the biology and the pathogenic mechanisms of JC virus that induce PML

Which is the best PML risk stratification strategy in natalizumab-treated patients affected by multiple sclerosis?

Background: The risk of progressive multifocal leukoencephalopathy (PML), a brain infection caused by John Cunningham virus (JCPyV), is the main limitation to the use of natalizumab, highly effective in the treatment of relapsing remitting multiple sclerosis (RRMS) patients. Establishing the PML risk against expected benefits represents an obligatory requirement of MS treatment algorithm. In order to achieve this goal, the aims of this study were to establish if JCPyV-DNA detection and non-coding control region (NCCR) arrangements could play a role of biomarkers, supporting anti-JCPyV antibodies measurement, actually the only parameter for PML risk stratification. Methods: Thirty RRMS patients in treatment with natalizumab were enrolled. Urine and blood samples were collected according to this calendar: baseline (T0), 4 (T1), 8 (T2), 12 (T3), 16 (T4), 20 months (T5) after beginning of natalizumab therapy. After JCPyV DNA extraction, a specific quantitative-PCR (Q-PCR) and arrangements’ analysis of NCCR and Viral Capsid Protein 1 (VP1) were carried out. Results: Q-PCR detected JCPyV DNA in urine and blood from baseline (T0) to 20 natalizumab infusions (T5), although JC viral load in urine was significantly higher compared to viremia, at all selected time points. A contextual analysis of the anti-JCPyV-antibodies versus JCPyV-DNA detection revealed that viral DNA preceded the antibodies’ presence in the serum. During the first year of natalizumab treatment, sequences isolated from blood displayed an archetype JCPyV NCCR structure with the occurrence of point mutations, whereas after one year NCCR re-organizations were observed in plasma and PBMC with duplication of NF-1 binding site in box F, duplication of box C and partial or total deletion of box D. VP1 analysis showed the amino acid change mutation S269F in plasma and S267L in PBMC, involving the receptor-binding region of VP1. Phylogenetic analysis suggested a stability and a similarity across different isolates of the JCPyV VP1. Conclusions: We highly recommend considering JCPyV-DNA detection and NCCR re-organizations as viral biomarkers in order to accurately identify JCPyV-infected patients with a specific humoral response not yet detectable and to identify NCCR arrangements correlated with the onset of neurovirulent variants

In vitro activity of fosfomycin against mucoid and non-mucoid Pseudomonas aeruginosa strains.

OBJECTIVES: Pseudomonas aeruginosa is the most frequent infectious agent in cystic fibrosis patients. P. aeruginosa resistance to first line antibiotics limits therapeutic options, but the therapeutic potential of older generation antibiotics, such as fosfomycin is under investigation. Fosfomycin does not belong to any other antibiotic class and acts by inhibiting the biosynthesis of the bacterial cell wall during the initial phases. A major problem for the use of fosfomycin against P. aeruginosa is the absence of a clinical breakpoint, the last one of 32 μg/mL was proposed in 2013 by the CA-SFM (Comité de l'Antibiogramme de la Société Française de Microbiologie). METHODS: Sixty-one strains of P. aeruginosa (thirty mucoid and thirty-one non mucoid) were collected from respiratory samples of cystic fibrosis patients. All isolates were identified by MALDI-TOF (Bruker, Bremen, Germany). Fosfomycin MICs against P. aeruginosa were measured using an automated system and confirmed by the gold standard method. RESULTS: There was no significant difference between mucoid and non-mucoid strains. MIC distribution and susceptibility rates were obtained by agar dilution method and from this data we measured MIC50 and MIC90 which were equal to 32 μg/mL and 64 μg/mL, respectively. From automated method results we measured a very major error (VME), major error (ME) and categorical agreement (CA) which were equal to 0%, 11% and 89%, respectively. Comparing automated and agar dilution methods, a Cohen's kappa equal to 73% (0.726) was measured. CONCLUSIONS: Our data suggest that fosfomycin has good effect against mucoid and non-mucoid strains of P. aeruginosa and automated systems can be implemented in clinical microbiology laboratories to assess fosfomycin with rapid and reproducible results

Merkel cell polyomavirus DNA detection in respiratory samples: study of a cohort of patients affected by cystic fibrosis.

BACKGROUND: The role of Merkel cell polyomavirus (MCPyV) as a respiratory pathogen is controversial, and it is still unclear in patients with cystic fibrosis (CF). The aim of this study was to define the MCPyV prevalence and epidemiology in CF patients in order to gain new insights into the association between MCPyV infection and respiratory diseases. METHODS: A one-year study was conducted testing oropharyngeal aspirate samples from 249 and 124 CF and non-CF patients, respectively. Detection of MCPyV was carried out by nested polymerase chain reaction (PCR). Moreover, a sequence alignment to examine viral capsid protein 1 (VP1) and a phylogenetic analysis were performed. RESULTS: MCPyV DNA was detected in 65 out of 249 samples analyzed CF (26%), a percentage that was higher than that recorded in non-CF patients (0.8%). There were no statistically significant differences in MCPyV prevalence according to gender, while there was a correlation between MCPyV detection and age. Interestingly, an association between the presence of MCPyV and the concurrent isolation of Staphylococcus aureus was found. Sequence analysis of MCPyV VP1 and phylogenetic analysis revealed a 99% homology with the published sequences of these viruses in GenBank. CONCLUSIONS: Detection of MCPyV in CF patient specimens pointed out a possible interaction between the virus and CF. Further studies are necessary to fully understand the involvement of MCPyV in the pathogenesis of respiratory disorders