No abstract

BackgroundThe taxonomic status of the brown dog tick (Rhipicephalus sanguineus sensu stricto), which has long been regarded as the most widespread tick worldwide and a vector of many pathogens to dogs and humans, is currently under dispute.MethodsWe conducted a comprehensive morphological and genetic study of 278 representative specimens, which belonged to different species (i.e., Rhipicephalus bursa, R. guilhoni, R. microplus, R. muhsamae, R. pusillus, R. sanguineus sensu lato, and R. turanicus) collected from Europe, Asia, Americas, and Oceania. After detailed morphological examination, ticks were molecularly processed for the analysis of partial mitochondrial (16S rDNA, 12S rDNA, and cox1) gene sequences.ResultsIn addition to R. sanguineus s.l. and R. turanicus, three different operational taxonomic units (namely, R. sp. I, R. sp. II, and R. sp. III) were found on dogs. These operational taxonomical units were morphologically and genetically different from R. sanguineus s.l. and R. turanicus. Ticks identified as R. sanguineus s.l., which corresponds to the so-called “tropical species” (=northern lineage), were found in all continents and genetically it represents a sister group of R. guilhoni. R. turanicus was found on a wide range of hosts in Italy and also on dogs in Greece.ConclusionsThe tropical species and the temperate species (=southern lineage) are paraphyletic groups. The occurrence of R. turanicus in the Mediterranean region is confirmed. A consensual re-description of R. sanguineus s.s. and R. turanicus will be necessary to solve the taxonomic problems within the so-called R. sanguineus group.

No abstract

Objectives: To detect possible severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) RNA contamination of inanimate surfaces in areas at high risk of aerosol formation by patients with coronavirus disease 2019 (COVID-19). Methods: Sampling was performed in the emergency unit and the sub-intensive care ward. SARS-CoV-2 RNA was extracted from swabbed surfaces and objects and subjected to real-time RT-PCR targeting RNAdependent RNA polymerase and E genes. Virus isolation from positive samples was attempted in vitro on Vero E6 cells. Results: Twenty-six samples were collected and only two were positive for low-level SARS-CoV-2 RNA, both collected on the external surface of continuous positive airway pressure helmets. All transport media were inoculated onto susceptible cells, but none induced a cytopathic effect on day 7 of culture. Conclusions: Even though daily contact with inanimate surfaces and patient fomites in contaminated areas may be a medium of infection, our data obtained in real-life conditions suggest that it might be less extensive than hitherto recognized.

Various next generation sequencing (NGS) based strategies have been successfully used in the recent past for tracing origins and understanding the evolution of infectious agents, investigating the spread and transmission chains of outbreaks, as well as facilitating the development of effective and rapid molecular diagnostic tests and contributing to the hunt for treatments and vaccines. The ongoing COVID-19 pandemic poses one of the greatest global threats in modern history and has already caused severe social and economic costs. The development of efficient and rapid sequencing methods to reconstruct the genomic sequence of SARS-CoV-2, the etiological agent of COVID-19, has been fundamental for the design of diagnostic molecular tests and to devise effective measures and strategies to mitigate the diffusion of the pandemic. Diverse approaches and sequencing methods can, as testified by the number of available sequences, be applied to SARS-CoV-2 genomes. However, each technology and sequencing approach has its own advantages and limitations. In the current review, we will provide a brief, but hopefully comprehensive, account of currently available platforms and methodological approaches for the sequencing of SARS-CoV-2 genomes. We also present an outline of current repositories and databases that provide access to SARS-CoV-2 genomic data and associated metadata. Finally, we offer general advice and guidelines for the appropriate sharing and deposition of SARS-CoV-2 data and metadata, and suggest that more efficient and standardized integration of current and future SARS-CoV-2-related data would greatly facilitate the struggle against this new pathogen. We hope that our ‘ vademecum ’ for the production and handling of SARS-CoV-2-related sequencing data, will contribute to this objective.

The sequence of the full-length gene encoding for the main capsid protein VP2 of 58 canine parvovirus (CPV) type 2c strains, along with recent CPV-2a/2b strains, was determined and analysed in comparison with reference CPV isolates. The CPV-2c strains displayed a low genetic variability and shared amino acid changes already detected in recent CPV-2a/2b isolates, with a phylogenetic clustering accounting for their geographical distribution. Analysis of the selection pressure driving CPV evolution confirmed that the VP2 gene is under purifying selection. The emergence and global spread of the new CPV variant provides an interesting model to better understand virus evolution.

No abstract

New SARS-CoV-2 mutations are constantly emerging, raising concerns of increased transmissibility, virulence or escape from host immune response. We describe a nested RT-PCR assay (~1500 bps) to detect multiple nucleotide changes resulting in key spike protein mutations distinctive of the major known circulating SARS-CoV-2 variants, including the three Variants of Concern (VOCs) 20I/501Y.V1 (United Kingdom), 20H/501Y.V2 (South Africa), and 20 J/501Y.V3 (Brazil), as well as the 20E.EU1 variant (Spain), the CAL.20C recently identified in California, and the mink-associated variant (GR, lineage B.1.1.298). Prior to application to field samples, the discriminatory potential of this PCR assay was explored using GISAID and Nextclade. To extend variant detection to challenging matrices such as sewage, where the amplification of long fragments is problematic, two short nested RT-PCR assays (~300 bps) were also designed, targeting portions of the region spanned by the long nested assay. The three newly-designed assays were then tested on field samples, including 31 clinical samples (7 fully-sequenced swab samples, and 24 uncharacterized ones) and 34 urban wastewater samples, some of which collected in areas where circulation of VOCs had been reported. The long assay successfully amplified 29 of the 31 swabs (93%), allowing the correct identification of variants 20I/501Y.V1 and 20E.EU1 present in the panel of previously characterized samples. The Spanish variant was detected in 14/24 of the uncharacterized samples as well. The sequences obtained using the short assays were consistent with those obtained with the long assay. Mutations characteristic of VOCs (UK and Brazilian variant) and of other variant (Spanish) were detected in sewage samples. To our knowledge, this is the first evidence of the presence of sequences harboring key mutations of 20I/501Y.V1 and 20 J/501Y.V3 in urban wastewaters, highlighting the potential contribution of wastewater surveillance to explore SARS-CoV-2 diversity. The developed nested RT-PCR assays can be used as an initial rapid screening test to select clinical samples containing mutations of interest. This can speed up diagnosis and optimize resources since it allows full genome sequencing to be done only on clinically relevant specimens. The assays can be also employed for a rapid and cost-effective detection of VOCs or other variants in sewage for the purposes of wastewater-based epidemiology. The approach proposed here can be used to better understand SARS-CoV-2 variant diversity, geographic distribution and impact worldwide.