Authors' response: COVID-19: how accurate are seroprevalence studies?

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Digging deep into intramitochondrial symbiosis: dual transcriptomics of the hard tick Ixodes ricinus and its bacterial symbiont Midichloria mitochondrii

INTRODUCTION. Ixodes ricinus is a hard tick, widespread in Europe and in the Mediterranean basin, that can act as vector of multiple diseases of human and veterinary importance. Midichloria mitochondrii is a bacterial symbiont of this tick, capable of living in the intermembrane space of mitochondria (Beninati et al., 2004, Appl. Environ. Microbiol., 70:2596\u20132602). The bacterium was found to be present in most tick individuals (Lo et al., 2006, Environ. Microbiol., 8:1280-1287) and to be vertically transmitted to the progeny (Sassera et al., 2008, Appl. Environ. Microbiol., 74:6138-6140). To investigate the relationship between host and symbiont we designed an ad-hoc protocol of dual RNA-Seq to sequence the transcriptomes of both organisms in different phases of tick engorgement. MATERIALS AND METHODS. Ticks were collected on roe deer, cut in half and stored in RNALater on site, in order to freeze the transcription patterns in each stage. Ticks were dissected in RNALater and total RNA was extracted from ovaries and salivary glands. A custom library construction kit was designed and used in order to preserve transcripts from both organisms and to limit the prevalence of rRNA sequences in the sample (normally over 85%). The kit protocol included the use of custom designed probes for rRNA depletion. Libraries were sequenced on Illumina machines and the resulting reads were assembled to obtain a reference transcriptome to be used for downstream analyses. RESULTS AND CONCLUSIONS. Ticks were subjected to a custom protocol for RNA sequencing which allowed to obtain an average of 40 million reads per sample and over 50% of the reads were transcripts of host or symbiont. After assembly, over 18,000 tick transcripts and over 1,200 bacterial transcripts were obtained. Differential expression analysis will be performed in the upcoming months; so far, the main result of this work is the development of a sound protocol for dual RNA-seq in this system

Combining Genome Surveillance and Metadata To Characterize the Diversity of Staphylococcus aureus Circulating in an Italian Hospital over a 9-Year Period

ABSTRACT Staphylococcus aureus is an opportunistic pathogen and a leading cause of morbidity and mortality worldwide. Genomic-based surveillance has greatly improved our ability to track the emergence and spread of high-risk clones, but the full potential of genomic data is only reached when used in conjunction with detailed metadata. Here, we demonstrate the utility of an integrated approach by leveraging a curated collection of clinical and epidemiological metadata of S. aureus in the San Matteo Hospital (Italy) through a semisupervised clustering strategy. We sequenced 226 sepsis S. aureus samples, recovered over a period of 9 years. By using existing antibiotic profiling data, we selected strains that capture the full diversity of the population. Genome analysis revealed 49 sequence types, 16 of which are novel. Comparative genomic analyses of hospital- and community-acquired infection ruled out the existence of genomic features differentiating them, while evolutionary analyses of genes and traits of interest highlighted different dynamics of acquisition and loss between antibiotic resistance and virulence genes. Finally, highly resistant clones belonging to clonal complexes (CC) 8 and 22 were found to be responsible for abundant infections and deaths, while the highly virulent CC30 was responsible for rare but deadly episodes of infections. IMPORTANCE Genome sequencing is an important tool in clinical microbiology, as it allows in-depth characterization of isolates of interest and can propel genome-based surveillance studies. Such studies can benefit from ad hoc methods of sample selection to capture the genomic diversity present in a data set. Here, we present an approach based on clustering of antibiotic resistance profiles that allows optimal sample selection for bacterial genomic surveillance. We apply the method to a 9-year collection of Staphylococcus aureus from a large hospital in northern Italy. Our method allows us to sequence the genomes of a large variety of strains of this important pathogen, which we then leverage to characterize the epidemiology in the hospital and to perform evolutionary analyses on genes and traits of interest. These analyses highlight different dynamics of acquisition and loss between antibiotic resistance and virulence genes

Detection of the SARS-CoV-2 in different biologic specimens from positive patients with COVID-19, in Northern Italy

Coronavirus disease 2019 (COVID-19) diagnosis is based on molecular detection of SARS-CoV-2 in respiratory samples such as nasal swab (NS). However, the evidence that NS in patients with pneumonia was sometimes negative raises the attention to collect other clinical specimens. SARS-CoV-2 was shown in 10.3% rectal swabs (RS), 7.7% plasma, 1% urine, and 0% feces from 143 NS-positive patients. Potential infection by fluids different from respiratory secretion is possible but unlikely