Oral HSV Infections: Molecular biology and photodynamic therapy as new diagnostic and therapeutic tools

Globally, it has been estimated that 3.7 billion people are Herpes simplex 1 (HSV) seropositive. In addition, numerous studies highlight the presence of HSV-1 and 2 (HSV-2) in the oral cavity of asymptomatic patients. The rate of asymptomatic shedding of both HSV-1 and HSV-2, the re-exacerbation following dental treatments and finally the professional risk, and new techniques to improve the diagnosis and treatment of this widespread pathology, will discussed. In particular the effectiveness of photodynamic therapy (PDT) in combination with molecular biology (Polymerase Chain reaction, PCR) tested on a cohort of patients will presented. PDT was performed by diode laser light at 660 nm and 100 mWatt of power (Helbo, Bredent Medical, Senden, Germany), combined with photosensitizer phenothiazine chloride at 1%. The results show that the main HSV spread mode is asymptomatic. Particularly, HSV-1 is statistically more detectable in HIV positive and oncological patients than in immunocompetent ones (p<0,01) but not in transplant patients (p>0,01). HSV-2 is more detectable in HIV positive patients than healthy ones (p<0,01) but not in cancer and transplant patients (p>0,01). Regarding the exacerbation of HVS-1 after dental treatments, studies in the literature are discordant. Finally, for the occupational risk, dentistry team appears to be more exposed than the general population. Extremely interesting data emerged from PDT applied in patients with herpetic lesion. The PCR in fact, besides being decisive in some doubtful clinical cases, has shown a statistically significant reduction of the viral load 100 and 150 times the first and the second cycle of PDT. Clinically PDT has proven effective in immediately reducing symptoms and healing, also increasing the recurrence interval. Therefore, it could be a powerful aid in patients with herpetic manifestations to be applied before carrying out any treatments in order to decrease the viral spread

Evaluation of “Caterina assay”: An Alternative Tool to the Commercialized Kits Used for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Identification

Abstract: Here we describe the first molecular test developed in the early stage of the pandemic to diagnose the first cases of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in Sardinian patients in February–March 2020, when diagnostic certified methodology had not yet been adopted by clinical microbiology laboratories. The “Caterina assay” is a SYBR®Green real-time reverse-transcription polymerase chain reaction (rRT-PCR), designed to detect the nucleocapsid phosphoprotein (N) gene that exhibits high discriminative variation RNA sequence among bat and human coronaviruses. The molecular method was applied to detect SARS-CoV-2 in nasal swabs collected from 2110 suspected cases. The study article describes the first molecular test developed in the early stage of the declared pandemic to identify the coronavirus disease 2019 (COVID-19) in Sardinian patients in February–March 2020, when a diagnostic certified methodology had not yet been adopted by clinical microbiology laboratories. The assay presented high specificity and sensitivity (with a detection limit ≥50 viral genomes/µL). No false-positives were detected, as confirmed by the comparison with two certified commercial kits. Although other validated molecular methods are currently in use, the Caterina assay still represents a valid and low-cost detection procedure that could be applied in countries with limited economic resource

CYTOMEGALOVIRUS INFECTION IN THE FIRST DAYS OF LIFE: PREVALENCE IN THE SOUTH SARDINIA FROM 2016 TO 2019

Congenital cytomegalovirus (CMV) infection poses a significant health risk to women of childbearing age and pregnant women and their unborn babies. Screening for CMV represents a crucial objective for Public Health, since this virus is one of the leading causes of childhood disability. CMV belongs to the Herpesviridae family and, once contracted the infection, it remains latent inside the organism for life, but it could be reactivated in case of weakening of the immune system. CMV infections are generally asymptomatic, but in immunocompromised patients, they can cause serious complications. CMV can be transmitted vertically from mother to child, as well as a primary, recurrent, or chronic infection. Maternal-fetal transmission of CMV can occur in the uterus (congenital infection), during labor or delivery (perinatal infection), or by lactation (postnatal infection) and it can cause miscarriage, intrauterine growth retardation, congenital anomalies and neonatal or postnatal pathologies of different severity. The aim of this study was to investigate the prevalence of CMV infection in a court of newborns clinically evaluated in the South Sardinia University Hospital. METHODS/RESULTS The postnatal diagnosis of CMV infection was carried out within the first 2-3 weeks of life of the newborn through the detection of the virus from urine, blood, and saliva. From September 2016 to July 2019, a total of 297 urine samples from the Department of Neonatology of Policlinico of University of Cagliari were analyzed, and proviral DNA was obtained by GeneProof Phatogen Kit. Our results showed a notable decrease in CMV infection cases in newborns, from 24% to 4% DNA positives, estimated in the South Sardinia. CONCLUSIONS The National Institute of Health underlines that, in Italy, the prevalence of infection is among the lowest in the literature. The results shown are in line with epidemiological data on the frequency of CMV infections. Our results suggest a low prevalence in comparison with the last years in the same region. REFERENCES • Kohmer N, Nagel A, Berger A, Enders M, Hamprecht K, Korn K, Kortenbusch M, Überla K, Rabenau HF. Laboratory diagnosis of congenital CMV infection in newborns: Impact of pre-analytic factors. J Clin Virol. 2019;115:32-

Confirmation of the <i>DRB1-DQB1 loci</i> as the major component of <i>IDDM1</i> in the isolated founder population of Sardinia

There is considerable uncertainty and debate concerning the application of linkage disequilibrium (LD) mapping in common multifactorial diseases, including the choice of population and the density of the marker map. Previously, it has been shown that, in the large cosmopolitan population of the UK, the established type 1 diabetes IDDM1 locus in the HLA region could be mapped with high resolution by LD. The LD curve peaked at marker D6S2444, 85 kb from the HLA class II gene DQB1, which is known to be a major determinant of IDDM1. However, given the many unknown parameters underlying LD, a validation of the approach in a genetically distinct population is necessary. In the present report we have achieved this by the LD mapping of IDDM1 in the isolated founder population of Sardinia. Using a dense map of microsatellite markers, we determined the peak of LD to be located at marker D6S2447, which is only 6.5 kb from DQB1. Next, we typed a large number of SNPs defining allelic variation at functional candidate genes within the critical region. The association curve, with both classes of marker, peaked at the loci DRB1-DQB1. These results, while representing conclusive evidence that the class II loci DRB1-DQB1 dominate the association of the HLA region to type 1 diabetes, provide empirical support for LD mapping

Conditional linkage disequilibrium analysis of a complex disease superlocus, <i>IDDM1</i> in the HLA region, reveals the presence of independent modifying gene effects influencing the type 1 diabetes risk encoded by the major <i>HLA-DBQ1</i>, -<i>DRB1</i> disease <i>loci</i>

Type 1 diabetes mellitus is a common disease with a complex mode of inheritance. Its aetiology is underpinned by a major locus, insulin-dependent diabetes mellitus 1 (IDDM1) in the human leukocyte antigen (HLA) region of chromosome 6p21, and an unknown number of loci of lesser individual effect. In linkage analyses IDDM1 is a single peak, but it is evident that the linkage is caused by allelic variation of three adjacent genes in a 75 kb region, namely the class II genes, HLA-DRB1, -DQA1 and -DQB1. However, even these three genes may not explain all of the HLA association. We investigated, in the founder population of Sardinia, whether non-DQ/DR polymorphic markers within a 9.452 Mb region encompassing the whole HLA complex further influence the disease risk, after taking into account linkage disequilibrium with the disease loci HLA-DQB1, -DQA1 and -DRB1. We generalized the conditional association test, the haplotype method, to detect marker associations that are independent of the main DR/DQ disease associations. Three regions were identified as risk modifiers. These associations were not only independent of the polymorphic exon 2 sequences of HLA-DQB1, -DQA1 and -DRB1, but also independent of each other. The individual contributions of these risk modifiers were relatively modest but their combined impact was highly significant. Together, alleles of single nucleotide polymorphisms at the DMB and DOB genes, and the microsatellite locus TNFc, identified ∼40% of Sardinian DR3 haplotypes as non-predisposing. This conditional analysis approach can be applied to any chromosome region involved in the predisposition to complex traits

Major factors influencing linkage disequilibrium by analysis of different chromosome regions in distinct populations: demography, chromosome recombination frequency and selection

Linkage disequilibrium (LD) mapping of disease genes is complicated by population- and chromosome-region-specific factors. We have analysed demographic factors by contrasting intermarker LD results obtained in a large cosmopolitan population (UK), a large genetic isolate (Sardinia) and a subisolate (village of Gavoi) for two regions of the X chromosome. A dramatic increase of LD was found in the subisolate. Demographic history of populations therefore influences LD. Chromosome-region-specific effects, namely the pattern and frequency of homologous recombination, were next delineated by the analysis of chromosome 6p21, including the HLA region. Patterns of global LD in this region were very similar in the UK and Sardinian populations despite their entirely distinct demographies, and correlate well with the pattern of recombinations. Nevertheless, haplotypes extend across recombination hot spots indicative of selection of certain haplotypes. Subisolate aside, chromosome-region-specific differences in LD patterns appear to be more important than the differences in intermarker LD between distinct populations

The Inter-regional distribution of HLA class II haplotypes indicates the suitability of the Sardinian population for case–control association studies in complex diseases

We have analysed HLA class II gene-based substructure of the Sardinian population in order to evaluate the possible influence of this parameter in the mapping of common disease loci using association methods. We first examined the distribution of the HLA-DRB1-DQA1-DQB1 haplotypes in 631 newborns from seven different regions of the island, and found that the most frequent haplotypes were uniformly distributed in all regions, but at frequencies unique to Sardinia. Other haplotypes, common in other white European populations, are consistently rare or absent across the whole island. Analysis of molecular variance (AMOVA) showed a very low degree of genetic differentiation between the coastal regions, which have suffered repeated invasions over many years, and the most internal and isolated part of the island. This suggests that there has been little genetic flow from the various populations that have invaded the island during the last 3000 years and that Sardinia is a relatively homogeneous population. The validity of these unrelated control HLA haplotype frequencies and our claim of homogeneity were established by demonstrating the near identity of the affected family-based control (AFBAC) HLA haplotype frequencies in 243 type 1 diabetes and 495 multiple sclerosis families from Sardinia and those of the unrelated controls. These results indicate that robust case–control studies can be carried out in Sardinia offering cost efficiency over certain family-based designs

The <i>HLA-DPB1</i>-associated component of the <i>IDDM1</i> and its relationship to the major loci <i>HLA-DQB1, -DQA1</i>, and-<i>DRB1</i>

The major histocompatibility complex (MHC) HLA region on chromosome 6p21 contains the major locus of type I diabetes (IDDM1). Common allelic variants at the class II HLA-DRB1, -DQA1, and -DQB1 loci account for the major part of IDDM1. Previous studies suggested that other MHC loci are likely to contribute to IDDM1, but determination of their relative contributions and identities is difficult because of strong linkage disequilibrium between MHC loci. One prime candidate is the polymorphic HLA-DPB1 locus, which (with the DPA1 locus) encodes the third class II antigen-presenting molecule. However, the results obtained in previous studies appear to be contradictory. Therefore, we have analyzed 408 white European families (200 from Sardinia and 208 from the U.K.) using a combination of association tests designed to directly compare the effect of DPB1 variation on the relative predisposition of DR-DQ haplotypes, taking into account linkage disequilibrium between DPB1 and the DRB1, DQA1, and DQB1 loci. In these populations, the overall contribution of DPB1 to IDDM1 is small. The main component of the DPB1 contribution to IDDM1 in these populations appears to be the protection associated with DPBI*0402 on DR4-negative haplotypes. We suggest that the HLADP molecule itself contributes to IDDM1

Strategic Research & Innovation Agenda - Italia - Volume 1

La Strategic Research and Innovation Agenda - Italia (SRIA It) redatta dalla piattaforma tecnologica italiana per l’aeronautica (ACARE Italia) considerando peculiarità, esigenze e priorità nazionali - illustra obiettivi e roadmap tecnologica condivisi dai vari attori nazionali fornendo le linee di orientamento per le attività R-ST nonché un utile riferimento nella definizione delle strategie di investimento nazionale per il settore dell’Aeronautica e piu’ in generale del Trasporto Aereo, includendo anche i sistemi per missioni speciali civili e gli aspetti duali