Abstract:
Introduction. Since the COVID-19 outbreak in late December 2019, more than 290 043
cases and 6713 deaths have been reported in the Republic of Moldova as of 27th of September 2021. The paucity of in-depth knowledge of the risk factors for severe COVID-19,
insufficient diagnostic tools for the detection of SARS-CoV-2, absence of specific and effective drug treatments as well as the absence of a best measure to monitor emerging
pathogens like SARS-CoV-2 led to a major burden on health care and economic systems
across the world. As SARS-CoV-2 continues to acquire genetic changes over time during
this pandemic, the full genome sequencing and identification of viral variants must continue.
This study aimed to perform whole-genome sequencing of SARS-VoV-2 isolates from Moldova and to provide a ‘real-time’ overview of the viral genotypes circulating in this geographical area. Suspected cases were screened for SARS-CoV-2 as per the advisory of the
Ministry of Health.
Material and methods. 4 nasopharyngeal swabs were collected in June 2020. Samples
were obtained from patients aged between 20 and 60 years old, residing in the Republic
of Moldova. All samples from COVID-19 patients were collected from individuals with
mild and severe COVID-19 symptoms and confirmed by real-time reverse transcriptionpolymerase chain reaction targeting the envelop (E) and nucleocapsid (N). The RNA samples were sequenced using an Illumina MiSeq sequencer according to standard procedure
developed previously. The metadata was created and the sequences were deposited on
GISAID international repository.
Results. Four patients from different geographic regions of the Republic of Moldova were
confirmed positive for SARS-CoV-2. Complete (29,900 nucleotides, 29,886 nucleotides,
27,179 nucleotides, 29,867 nucleotides) genomes were obtained. Phylogenetic analysis
showed that the Moldovan sequences belonged to different clusters. EPI_ISL_516935
(GISAID accesion ID) belongs to clade G and EPI_ISL_516934, EPI_ISL_516936,
EPI_ISL_516938 belong to clade GR. These clades are prevalent in EU. All isolates showed
typical mutation (Spike D614G) for G clade. Three genomes showed mutations on nucleocapsid protein, the G204R which is specific for GR clade. We also observed other aminoacids substitutions (NS3 V90F, NSP2 G339S, NSP3 D218E, NSP3 V1243A, NSP12 P323L,
Spike E1202Z, Spike L763V, NS7a E95Z, NSP3 N1785D, Spike G769V, N P151L, N R203K,
NSP12 R457C, NSP13 A379V, NSP15 N4S NS3 S165F, NSP2 L410F), mutations that are
less frequent.
Conclusions. The four SARS-CoV-2 sequences obtained from Republic of Moldova represent two different introductions into the country. The genetic heterogeneity is as noted
globally. The Spike D614G and N G204R has a rapid spread in the pandemic. Continuous
monitoring and analysis of the sequences of new cases from Republic of Moldova and the
other affected countries would be vital to understand the genetic evolution and rates of
substitution of the SARS-CoV-2.
