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

Periodontal microbiota of Sardinian children: comparing 200-year-old samples to present-day ones

Introduction: The microrganisms of the human oral cavity include more than 700 species or phenotypes of bacteria. Some “diseases of civilization” are strictly correlated to changes in the microbiome following the food revolution that occurred after WWII. For that reason, a precise recognition of the microbiome profile before and after this period should be useful to determine the health-compatible model of icrobiome. The aim of this study was to compare the microbiome profiles (number of total cells, and pathogen types) of dental samples obtained from two distinct groups of children, a 200-year-old retrieved one and a present one. Methods: Two different groups of samples have been studied. The first group was a set of 50 recent subgingival plaque samples obtained from children of age 2-8 years, 14 males and 36 females. They were enrolled by the Department of Dental Disease Prevention (University of Cagliari, in Sardinia, Italy) during standard dental care procedures. None reported periodontal disease and none had been under antibiotic therapy during the previous 6 months. The second group was an old retrieved group that included 24 teeth from 6 different 6- to 8-year-old crania fragments; they were obtained from a 200-year-old charnel-house located in Villaputzu, a city close to Cagliari. Representative periodontal bacteria have been identified by a previously published real-time PCR procedure (Sokransky et al., 1998) in which P. gingivalis and T. forsythia (red complex), A. Original article 2/5 www.jpnim.com Open Access Journal of Pediatric and Neonatal Individualized Medicine • vol. 6 • n. 1 • 2017 Orrù • Contu • Casula • Demontis • Blus • Szmukler-Moncler • Serreli • Maserati • Steri • Fanos • Coghe • Denotti actinomycetemcomitans (green complex) and F. nucleatum (orange complex) were detected. In addition, the title of each pathogen was expressed as a percentage of the total bacteria (biofilm) in the sample. Results and discussion: The profile of periodontal microbiomes, between recent/ancient samples showed a significant difference relative to Sokransky’s red complex bacteria (p < 0.05). In all analyzed periodontal strains, the pathogenic bacteria P. gingivalis and T. forsythia showed the highest title in the recent group. Conclusions: Our hypothesis is that the transfer of “commensal-pathogen” as an absolute number on the oral biofilm might be linked to the distinct alimentary habits of the two populations. Some diet rich in reducing agents, such as processed meat-based foods, might be able to increase the average number of pathogen anaerobic bacteria in the oral microbiota. The outcome would be an increase of the oral systemic diseases reported with these pathogens. Our data suggest that the ancient Sardinian population was able to control the pathogen oral anaerobic biofilm by some diet rich in antioxidant compounds. Further investigations are required to focus on the genetic profile and the health status of this ancient population but it appears that molecular microbiology might be considered as the “time machine” in oral biology

Changes in the oral status and periodontal pathogens in a Sardinian rural community from pre-industrial to modern time

The oral microbial profile in humans has evolved in response to lifestyle changes over the course of different eras. Here, we investigated tooth lesions and the microbial profile of periodontal bacteria (PB) in dental calculus of a Sardinian pre-industrial rural community. In total, 51 teeth belonging to 12 historical individuals buried in an ossuary in the early 1800s and 26 modern teeth extracted from 26 individuals from the same geographical area were compared to determine the oral health status, bacterial load and amount of most relevant PB. Total caries and bacterial genomes count appeared to be sex-related in historical samples. Historical females presented a higher incidence of caries, PB pathogens and a higher bacterial load than historical males. Furthermore, we compared the PB profile of the historical individuals with the modern ones, revealing a notable increase in modern individuals of PB belonging to “Red complex bacteria” often associated with periodontitis and other chronic diseases of modern life. Our findings could be explained through an analysis of environmental factors such as socioeconomic, hygienic and healthy conditions that can have a great impact on oral health and bacterial composition among individuals of the same and different era

Periodontal microbiota of Sardinian children: comparing 200-year-old samples to present-day ones

Introduction: The microrganisms of the human oral cavity include more than 700 species or phenotypes of bacteria. Some “diseases of civilization” are strictly correlated to changes in the microbiome following the food revolution that occurred after WWII. For that reason, a precise recognition of the microbiome profile before and after this period should be useful to determine the health-compatible model of microbiome. The aim of this study was to compare the microbiome profiles (number of total cells, and pathogen types) of dental samples obtained from two distinct groups of children, a 200-year-old retrieved one and a present one. Methods: Two different groups of samples have been studied. The first group was a set of 50 recent subgingival plaque samples obtained from children of age 2-8 years, 14 males and 36 females. They were enrolled by the Department of Dental Disease Prevention (University of Cagliari, in Sardinia, Italy) during standard dental care procedures. None reported periodontal disease and none had been under antibiotic therapy during the previous 6 months. The second group was an old retrieved group that included 24 teeth from 6 different 6- to 8-year-old crania fragments; they were obtained from a 200-year-old charnel-house located in Villaputzu, a city close to Cagliari. Representative periodontal bacteria have been identified by a previously published real-time PCR procedure (Sokransky et al., 1998) in which P. gingivalis and T. forsythia (red complex), A. actinomycetemcomitans (green complex) and F. nucleatum (orange complex) were detected. In addition, the title of each pathogen was expressed as a percentage of the total bacteria (biofilm) in the sample. Results and discussion: The profile of periodontal microbiomes, between recent/ancient samples showed a significant difference relative to Sokransky’s red complex bacteria (p < 0.05). In all analyzed periodontal strains, the pathogenic bacteria P. gingivalis and T. forsythia showed the highest title in the recent group. Conclusions: Our hypothesis is that the transfer of “commensal-pathogen” as an absolute number on the oral biofilm might be linked to the distinct alimentary habits of the two populations. Some diet rich in reducing agents, such as processed meat-based foods, might be able to increase the average number of pathogen anaerobic bacteria in the oral microbiota. The outcome would be an increase of the oral systemic diseases reported with these pathogens. Our data suggest that the ancient Sardinian population was able to control the pathogen oral anaerobic biofilm by some diet rich in antioxidant compounds. Further investigations are required to focus on the genetic profile and the health status of this ancient population but it appears that molecular microbiology might be considered as the “time machine” in oral biology

Genome-wide association analyses based on whole-genome sequencing in Sardinia provide insights into regulation of hemoglobin levels

We report genome-wide association study results for the levels of A1, A2 and fetal hemoglobins, analyzed for the first time concurrently. Integrating high-density array genotyping and whole-genome sequencing in a large general population cohort from Sardinia, we detected 23 associations at 10 loci. Five signals are due to variants at previously undetected loci: MPHOSPH9, PLTP-PCIF1, ZFPM1 (FOG1), NFIX and CCND3. Among the signals at known loci, ten are new lead variants and four are new independent signals. Half of all variants also showed pleiotropic associations with different hemoglobins, which further corroborated some of the detected associations and identified features of coordinated hemoglobin species production

Genome sequencing elucidates Sardinian genetic architecture and augments association analyses for lipid and blood inflammatory markers

We report similar to 17.6 million genetic variants from whole-genome sequencing of 2,120 Sardinians; 22% are absent from previous sequencing-based compilations and are enriched for predicted functional consequences. Furthermore, similar to 76,000 variants common in our sample (frequency >5%) are rare elsewhere (<0.5% in the 1000 Genomes Project). We assessed the impact of these variants on circulating lipid levels and five inflammatory biomarkers. We observe 14 signals, including 2 major new loci, for lipid levels and 19 signals, including 2 new loci, for inflammatory markers. The new associations would have been missed in analyses based on 1000 Genomes Project data, underlining the advantages of large-scale sequencing in this founder population