Dogs and cats are susceptible to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). During the pandemic, several studies have been performed on owned cats and dogs, whereas limited data are available on the exposure to stray animals. The objective of this study was to investigate the exposure to SARS-CoV-2 of feral cats and kennel dogs in northeastern Italy, through serological and molecular methods. From May 2021 to September 2022, public health veterinary services collected serum, oropharyngeal, and rectal swab samples from 257 free-roaming dogs newly introduced to shelters, and from 389 feral cats examined during the routinely trap–neutered–return programs. The swabs were analyzed for viral RNA through a real-time reverse transcriptase PCR (rRT-PCR), and sera were tested for the presence of the specific antibody against SARS-CoV-2 (enzyme-linked immunosorbent assay). Serology was positive in nine dogs (9/257) and three cats (3/389), while two asymptomatic cats tested positive to rRT-PCR. One cat turned out to be positive both for serology and molecular analysis. In addition, this study described the case of a possible human-to-animal SARS-CoV-2 transmission in a cat that travelled in close contact to a COVID-19-positive refugee from Ukraine. This study shows that SARS-CoV-2 can infect, in natural conditions, stray cats and kennel dogs in northeastern Italy, although with a low prevalence.

Next-generation sequencing (NGS) from SARS-CoV-2-positive swabs collected during the last months of 2022 revealed a large deletion spanning ORF7b and ORF8 (426 nt) in six patients infected with the BA.5.1 Omicron variant. This extensive genome loss removed a large part of these two genes, maintaining in frame the first 22 aminoacids of ORF7b and the last three aminoacids of ORF8. Interestingly, the deleted region was flanked by two small repeats, which were likely involved in the formation of a hairpin structure. Similar rearrangements, comparable in size and location to the deletion, were also identified in 15 sequences in the NCBI database. In this group, seven out of 15 cases from the USA and Switzerland presented both the BA.5.1 variant and the same 426 nucleotides deletion. It is noteworthy that three out of six cases were detected in patients with immunodeficiency, and it is conceivable that this clinical condition could promote the replication and selection of these mutations.

Next generation sequencing (NGS) from SARS-CoV-2-positive swabs collected during the last months of 2022 revealed a large deletion between ORF7b and ORF8 (426 nt) in six patients infected with the BA.5.1 Omicron variant. This extensive genome loss removed a large part of these two genes, maintaining in frame the first 22 aminoacids of ORF7b and the last 3 aminoacids of ORF8. Interestingly, the deleted region was flanked by 2 small repeats, likely involved in the formation of a hairpin structure. Similar rearrangements, comparable in size and location to the deletion, were also identified in 15 sequences in the NCBI database. In this group, 7 out of 15 cases from the USA and Switzerland presented both the BA.5.1 variant and the same 426 nuclotides deletion. It is noteworthy that 3 out of 6 cases were detected in patients with immunodeficiency and is conceivable that this clinical condition could promote the replication and selection of these mutations.

The growing demand for precision diagnostics in cystic fibrosis and other genetic disorders, such as cancers, is driving the need for sequencing platforms that combine analytical robustness, scalability, and cost-efficiency. In this study, we performed a direct comparison between two leading Next-Generation Sequencing (NGS) platforms, MiSeq (Illumina, CA, USA) and DNBSEQ-G99RS (MGI Tech Co., Shenzhen, China), using a CE-IVD-certified CFTR panel (Devyser AB), selected for its complexity and variant spectrum, including SNVs, CNVs, and intronic polymorphisms. A total of 47 genomic DNA samples from routine clinical activity were analyzed on both platforms. Illumina sequencing covered all CFTR variants using standard workflows, while MGI data were generated from residual diagnostic DNA, with informed consent. Sequencing data were processed using Amplicon Suite v3.7.0 for variant calling, annotation, and ACMG classification. Quality control metrics and platform-specific parameters were also evaluated. Both platforms demonstrated complete concordance in variant detection, including SNVs, CNVs, and complex alleles (e.g., Poly-T/TG). Illumina exhibited slightly superior basecalling quality and allelic frequency uniformity, while MGI achieved higher sequencing depth (mean ~2793×) and demultiplexing efficiency. No false positives, false negatives, or discordant HGVS annotations were observed. The use of full-gene CFTR sequencing enabled granular and technically rigorous cross-platform validation. These findings confirm the analytical equivalence of Illumina and MGI for diagnostic genotyping. Moreover, MGI’s greater data output and flow cell capacity may offer tangible advantages in high-throughput settings, including somatic applications such as liquid biopsy and molecular oncology workflows.