## 4 Molecular Analyses
### Viruses in Parchments
The detection of JSRV in these manuscripts, through the application of the shotgun DNA analyses, suggests that what we have identified is the homologous endogenous sequences of the virus that are inserted in the genome of the sheep, which perfectly supports that the investigated folios were made from sheepskin. We could not detect this virus in the virobiota of the _C. Assemanianus_ sample R, which was also identified as being from a sheep, but it can be explained by the lower retrieval of the genetic material from this sample due to the non-invasive sampling that was applied in this folio using erasers [3]. This virus was not detected in the samples of _L. Sinaiticum_ S6 [3] and _Kiev Folia_[8], both of which were shown to be manufactured, fully and partially, respectively, from calfskin (see Table 1). The discovery of ovine retroviruses in the samples that were identified a priori as sheepskins opens an interesting perspective that could be further used to investigate the population genetics in the parchment materials by the performance of further bioinformatic analysis [94]. Another interesting case was the detection of the Dasheen mosaic virus (DsMV) in the microbiomes of the two samples that were investigated from the _L. Sinaiticum_ (see Figure 9) [3]. This is a single-stranded RNA virus, belonging to a group of plant pathogenic viruses, specifically to the genus _Potyvirus_, which naturally infects the Taro plant (_Colocasia esculenta_) [95]. The detection of this ssRNA virus in a DNA sequencing analysis was explained by its integration into the genome of the host plant, Taro [3]. All types of viruses can become endogenous by integrating the viral copy sequences into the genomes of various host organisms. These sequences are generally referred to as endogenous viral elements (EVEs) [96]. The integration of potyvirus sequences into host plant genomes is well known [97], and it may be the only plausible explanation for this finding. However, the detection of DsMV in this codex led to an intriguing hypothesis through a literature search. Taro is known to have been cultivated in ancient Egypt, where it was an important crop  
Figure 9: Shotgun metagenomics showing the relative abundance (%) of viruses in the microbiomes of ancient Slavonic codices [3,8] (and in _Glagolita Clozianus_, this study). The overall relative abundances of the viruses in the microbiomes were: _L. Sinaiticum_ S5: 3%; _L. Sinaiticum_ S6: \(<\)1%; _C. Assemanianus_ P: 15%; _C. Assemanianus_ R: \(<\)1%; Kiev Folia: 53%; _Glagolita Clozianus_: 11% (This study). plant [97]. At this time, this plant was mainly used for the consumption of edible leaves and corms, but also for its high concentration of anthocyanin [98]. [PERSON] et al. [99] previously described that some inks that were used for parchment illustration were composed of anthocyanins which were extracted from different plants, through a process in which the organic dye was precipitated with an inert binder, usually a metallic salt. Thus, could this be an indication that some of the inks that were used in this codex were composed of anthocyanin extracted from Taro? Well, one hypothesis to explain the detection of the DsMV in the _Liturgarium Sinaiticum_ could be that the Taro plant, which was very common in ancient Egypt, was used to extract the anthocyanins for the local production of the inks. In fact, previous studies on this codex, using X-ray fluorescence analysis (XRF) in a non-destructive and non-invasive manner, led to the conclusion that an organic dye had been applied for the yellow and a blue pigment [100]. Although only a hypothesis, the discovery of the Dasheen mosaic virus could indirectly support the previous theory, which some philologists suggest, that this codex was composed and written in the same place where it was found, namely in the monastery of _St. Catherine_ in Sinai, Egypt [101], by using the materials that were available in this area.