In December 2019 many cases of pneumonia without a defined causative factor were reported in Wuhan China, with some patients rapidly developing respiratory distress syndrome or acute respiratory failure [1]. On 7 January a novel coronavirus was identified by the Chinese Center for Disease Control and Prevention (CCDC) from the throat swab sample of a patient and was subsequently named ‘2019-nCoV’ by the WHO [2]. The 2019-nCoV infection caused clusters of severe respiratory illness similar to severe acute respiratory syndrome. Most patients were men and a sizable proportion suffered from underlying conditions [3]. The first COVID-19 case in Greece was diagnosed on 26 February of 2020 and on March 23rd with 695 confirmed cases and 17 deaths a nation-wide restriction on freedom of movement was enforced. On 8 March the first patient with confirmed pneumonia by 2019-nCoV was admitted in an Intensive Care Unit (ICU) due to pulmonary failure. Globally, up until 29 October 2021, there have been 245,373,039 confirmed cases of COVID-19, including 4,979,421 deaths, reported to WHO [4]. In Greece, for the same time period, a total number of 734,778 cases have been reported with 15,856 registered deaths [5]. The virus continued to devastate public health and economy worldwide enduring a second, a third and now a fourth wave of outbreaks, and only mass vaccination could protect the general public. A high vaccination rate is essential in order to approach herd immunity and to bring the pandemic under control (Mathieu et al., 2021). Up to 28 October 2021, a total of 6,838,727,352 vaccine doses have been administered worldwide [4] and 12,692,458 in Greece [6].

SARS-CoV-2, like other RNA viruses, is prone to genetic evolution with the development of mutations over time, resulting in the emergence of multiple variants that may have different characteristics compared to its ancestral strains. New variants may possess the ability to evade available detection methods, therapeutic regimens and reduce vaccination effectiveness. Even a single mutation can drastically affect a virus’ ability to evade the immune system and complicate the vaccine development progress against the virus. Adaptive mutations in the viral genome can alter the virus’s pathogenic potential and the continuous emergence of new virus variants with increased infectivity have obliged the scientific community to be vigilant for the virus evolution. Multiple variants of SARS-CoV-2 have been described, of which a few are considered variants of concern (VOCs), given their impact on public health [7]. Thus, to effectively control the pandemic, it is considered crucial to monitor the possible emergence of rare mutations especially in the spike-encoding region [8]. Tracking the SARS-CoV-2 variants and their mutational patterns in different areas not only can assist in the implementation of appropriate measures to protect the population (e.g., from super spreaders) but also provides valuable data to the vaccine development platform [7,9].

2. Discussion

Current study explored the SARS-CoV-2 mutational pattern in a representative cohort of samples collected from the center of Athens and several rural areas around Greece covering a period from early July till the end of August 2021. researchers chose to investigate certain key mutations which, according to current literature, have an important impact on public health or are carried by SARS-CoV-2 variants of concern. The prevalent mutations that researchers identified were the following spike mutations: N501Y (100%), D614G (96.4%) and P681R (90.1%) and the emerging variants were the Delta (90.1%), Alpha (28.2%) and Beta (2.7%).

The RT-qPCR technique was implemented for the identification of the mutations and variants since it is more affordable than other molecular techniques, such as sequencing. researchers suggest that this screening strategy could therefore be implemented more easily in most countries. Undoubtedly, if sequencing is also an option, it offers not only confirmation of the RT-PCR findings but also epidemiological advantages through the identification of both known and novel transcripts. No confirmation of current findings by sequencing and the relatively restricted sample of specimens used constitute the limitations of current study. As far as the former is concerned, although it does represent a weakness for analyzing clinical samples, it is not a strong disadvantage considering the main objective of these assays, which is to analyze samples more conveniently, in terms of money and time to keep molecular epidemiological surveillance in near-real-time. Therefore, researchers believe that these assays could provide information that helps prevent and control future outbreaks generated by the introduction of variants in new geographical locations [25].

Different designations have been given to the variants by different phylogenetic classification systems (e.g., Pango lineage, Nextstrain, GISAID). The WHO uses letters of the Greek alphabet to name distinct variants and has also categorizes the variants based on their properties and epidemiological impact [7]. Therefore, the Alpha, Beta, Gamma and Delta variants are considered variants of concern (VOC) according to WHO due to their important impact on transmissibility, severity and immunity affecting public health globally. Interestingly, the ECDC -with a similar classification system of the variants- classifies the Alpha not as a VOC but as a “De-escalated Variant”: either no longer circulating or with no impact and concerning properties [8]. The prevalence of the Alpha variant has declined worldwide after the emergence of the Delta variant [26].

Finding mutations, especially with known clinical impact, might have significant implications for public health policies, surveillance and immunization strategies; therefore, researchers consider that the world’s health authorities should promote tracking of mutations with important biological effects even through rapid screening techniques [17,27]. The fact that this pandemic has beset health, healthcare systems and the economy worldwide dictates the urgency for affordable and rapid techniques for SARS-CoV-2 mutations’ and variants’ immediate and effective global surveillance.

In conclusion, this manuscript describes the application of RT-qPCR assays to detect some mutations of interest in Greece at this pandemic period. researchers investigated three mutations of concern in the collected samples. SARS-CoV-2′s mutational pattern was explored through rapid molecular screening and is reported here for the first time in Greece, to current knowledge.