**1.2 Diagnostic capability**


#### *1.2.1 Nuclear acid based testing of SARS-CoV-2 variants*

#### *1.2.1.1 Detection of mutated variants with standard RT-PCR*

As it multiplies, the virus's genome is continually changing. New SARS-CoV-2 pandemic events could be triggered by new variants containing genetic mutations. Because most PCR primers were built using solitary virions in their early stages [30], notably the standard genetic material (SARS-CoV-2, NC 045512.2) [31], just a genetic variation during a first evolution sequence could result in reduced RT-PCR test amplification efficiency and false-negative detection results [32]. Studies of genotyping samples reported to GenBank and GISAID indicated that variations in the ORF1ab region were most common in Germany and China [33].

Another analysis relies on 31,421 SARS-CoV-2 genomic specimens and discovered that the majority of alterations were with the objective of several N genome primers and probes [34], which could alter the accuracy of PCR amplification in RT-PCR tests that probe the N genome. Variations in the N genome have been observed to interfere with detection in some cases [35]. All objectives of the US CDC-recommended COVID-19 diagnosis primers had mutations, while the targets of N genome primers and probes used throughout Japan, Thailand, and China had various mutations in distinct clusters, suggesting that the N gene may not have been a reliable target for RT-PCR kits and that these N gene-based kits should be reported periodically for a rising alpha, beta, gamma, delta variants [35].

#### *1.2.1.2 Sequencing for diagnosis of SARA-CoV-2 variants*

When compared with real time-PCR, virus genotype had the limitations of being more expensive, requiring more analysis of given information, and having lesser medical efficiency, making it inappropriate for massive population detection. However, utilizing metagenomics RNA sequencing techniques [31], the first genetic arrangement of SARS-CoV-2 was obtained. A study by the WHO and China found that during the beginning of December 2019 to the middle of February 2020, Illumina and Nanopore technologies were used to identify and sequence 104 SARS-CoV-2 variants [36]. More than 1000 comparable variants have since been published in the GISAID and GenBank databases, and the genomic and proteomics of SARS-CoV-2 have also been found [37]. The benefit of homologous recombination identification is that it allows for the tracking of viral changes by gathering data on recent variants. The viral genome is sequenced for the detection and classification of novel coronavirus variants throughout time [36]. Random changes in the genetic coding accrue with the speed of about 2/month when the virus multiplies and expands, according to data from closely watching viral development [38]. The latest mutant (changed) coronaviruses had been discovered, such as alpha (B.1.1.7), beta (B.1.351), gamma (P.1), and delta (B.1.617.2), which could result in the virus spreading considerably faster [39].

High-throughput approaches or portable fast sequence arrangement technologies had already been designed as distinguishing equipment for COVID-19 due to increased demand. Nanopore target sequencing (NTS) is appealing for clinical testing since it is fast, accessible, and efficient. In 1 h of sequencing [40], an NTS technique sequencing viral areas can identify very few as 10 viral copies/mL. These recently developed portable or quantifiable technologies, in comparison to classic sequencing techniques, which are typically expensive, may give accurate elevated diagnostics during epidemics. In the United Kingdom, NTS is being used in a projected genetic monitoring effort to create real-time genetic monitoring of SARS-CoV-2 [41], allowing sample-to-report in less than 24 hours. The use of genetic and epidemiologic analyses together speeds up the detection of possible transmission events and aids in the implementation of prompt control and prevention measures. When NTS is utilized to examine for reductions and different mutations in the SARS-CoV-2 gene in patients who have been infected with the virus, a putative pathogenic mechanism may be uncovered [42]. Furthermore, a novel molecular testing method relying on Sanger sequence techniques was capable to identify SARS-CoV-2 Genetic code (RNA) from viruses suspended components in the transmissible channel, RNA extraction could be skipped altogether without sacrificing performance at a testing flow rate of more than 1,000,000 tests per day [43], meaning that RNA extraction may be omitted fully without sacrificing performance. Natural variations in large populations could be tracked at general genomic ranges or specific regions over time or within a geographic location with this capability, allowing the locations and provenance of mutations to be identified once the quantitative capability is in place (**Figure 3**).

#### *1.2.2 Protein-based testing of SARS-CoV-2 variants*


#### *1.2.2.1 Antibody testing*

As a growing number of people around the world prefer to keep a maximum distance from every person and remain at their houses, the concentration of pandemic protection and management has moved to comprehensive serological antibody
