The rapid spread of SARS-COV-2 Omicron variant in Italy reflected early through wastewater surveillance

The SARS-CoV-2 Omicron variant emerged in South Africa in November 2021, and has later been identified worldwide, raising serious concerns. A real-time RT-PCR assay was designed for the rapid screening of the Omicron variant, targeting characteristic mutations of the spike gene. The assay was used to test 737 sewage samples collected throughout Italy (19/21 Regions) between 11 November and 25 December 2021, with the aim of assessing the spread of the Omicron variant in the country. Positive samples were also tested with a real-time RT-PCR developed by the European Commission, Joint Research Centre (JRC), and through nested RT-PCR followed by Sanger sequencing. Overall, 115 samples tested positive for Omicron SARS-CoV-2 variant. The first occurrence was detected on 7 December, in Veneto, North Italy. Later on, the variant spread extremely fast in three weeks, with prevalence of positive wastewater samples rising from 1.0% (1/104 samples) in the week 5-11 December, to 17.5% (25/143 samples) in the week 12-18, to 65.9% (89/135 samples) in the week 19-25, in line with the increase in cases of infection with the Omicron variant observed during December in Italy. Similarly, the number of Regions/Autonomous Provinces in which the variant was detected increased from one in the first week, to 11 in the second, and to 17 in the last one. The presence of the Omicron variant was confirmed by the JRC real-time RT-PCR in 79.1% (91/115) of the positive samples, and by Sanger sequencing in 66% (64/97) of PCR amplicons. In conclusion, we designed an RT-qPCR assay capable to detect the Omicron variant, which can be successfully used for the purpose of wastewater-based epidemiology. We also described the history of the introduction and diffusion of the Omicron variant in the Italian population and territory, confirming the effectiveness of sewage monitoring as a powerful surveillance tool

DNA metabarcoding uncovers fungal diversity of mixed airborne samples in Italy

<div><p>Fungal spores and mycelium fragments are particles which become and remain airborne and have been subjects of aerobiological studies. The presence and the abundance of taxa in aerobiological samples can be very variable and impaired by changeable climatic conditions. Because many fungi produce mycotoxins and both their mycelium fragments and spores are potential allergens, monitoring the presence of these taxa is of key importance. So far data on exposure and sensitization to fungal allergens are mainly based on the assessment of few, easily identifiable taxa and focused only on certain environments. The microscopic method used to analyze aerobiological samples and the inconspicuous fungal characters do not allow a in depth taxonomical identification. Here, we present a first assessment of fungal diversity from airborne samples using a DNA metabarcoding analysis. The nuclear ITS2 region was selected as barcode to catch fungal diversity in mixed airborne samples gathered during two weeks in four sites of North-Eastern and Central Italy. We assessed the taxonomic composition and diversity within and among the sampled sites and compared the molecular data with those obtained by traditional microscopy. The molecular analyses provide a tenfold more comprehensive determination of the taxa than the traditional morphological inspections. Our results prove that the metabarcoding analysis is a promising approach to increases quality and sensitivity of the aerobiological monitoring. The laboratory and bioinformatic workflow implemented here is now suitable for routine, high-throughput, regional analyses of airborne fungi.</p></div

Jackknifed principal coordinate analysis (PCoA) plot of Bray-Curtis distances between the samples of the four sites.

<p>Ellipsoids show the statistical confidence of the analysis.</p

Box plots of Chao1 and Shannon diversity indices estimated for each site.

<p>Significant differences among sites were calculated with ANOVA and Duncan’s new multiple range test and are indicated by different letters a-c (p-value < 0.05).</p

Doughnut charts showing fungal taxonomic composition at division (A) and class (B) level in the four sampling sites.

<p>Relative abundances of taxa are reported in percentage. “Unidentified fungi” comprehends Fungi sp. and fungi with no blast hit. Taxa accounting for <0.1% of reads are grouped as “other”.</p

DNA metabarcoding uncovers fungal diversity of mixed airborne samples in Italy - Fig 5

<p>A) Bar charts showing the taxonomic composition at genus level in the four sampling sites. Abundances of taxa are reported with the percentage values of reads. Taxa accounting for <0.1% of reads are grouped as “Other”. B) Venn diagram shows the number of unique and shared taxa identified at the genus level among sites (as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0194489#pone.0194489.s006" target="_blank">S2 Table</a>).</p

Geographical location of the sampling sites in the North-Eastern and Central Italy.

<p>Region and city names are reported (FVG: Friuli Venezia Giulia). The map has been retrieved and modified from <a href="http://www.d-maps.com" target="_blank">http://www.d-maps.com</a>.</p