## 4 Discussions  
In this pilot study, we tested HNSCC and skin tumor samples using a very sensitive assay for detecting very small amounts of DNA in a multiplex design. The method was optimized to detect 67 etiologic agents simultaneously from _Papillomavirusidae_, _Herpesviridae_ and _Polyomavirusidae_ families. The assay has the advantage of testing many viral etiologies in a short period of time, by a completely automatized method, which removes any source of possible errors. The results of the testing were validated by the quality controls used. To compare our experimental results with other research data, we reviewed the recent scientific literature regarding these viruses' involvement in the carcinogenesis of HNSCCs. Herpes viruses are known to have an oncogenic role [5]. The highest prevalence of the viruses we tested was for EBV1 (61.63%) and EBV2 was detected in fewer samples (3.8%). Therefore, we are mentioning [PERSON] et al., who detected using Real Time PCR a 48% EBV prevalence in Primary Laryngeal Squamous Cell Carcinoma, from Formalin-fixed paraffin-embedded (FFPE) tissue sections [23]. [PERSON] et al. analyzed the prevalence and prognostic values of EBV infections in Japanese patients with different types of HNSCC from fresh frozen samples, using PCR. The authors detected DNA/EBV in 69.9% HNC cases, from which 73.3% was EBV1 and 18.5% EBV2. Even though EBV was highly prevalent in cancers of the oropharynx and nasopharynx, EBV could not be used as a prognostic marker for the studied cohort [24]. Both EBV types were detected in another study which analyzed EBV in FFPE HNSCC tissue samples by PCR and confirmed by nucleotide sequencing. The authors detected a 12.8% EBV prevalence with 90% EBV1 and 10% EBV2 [25]. [PERSON] et al. used EBV markers to develop a risk score for nasopharyngeal carcinoma which opens new opportunities for personalized risk prediction and for an early diagnosis of nasopharyngeal carcinoma [26]. Another team of researchers used the viral load of DNA/EBV detected in 75% of their tested nasopharyngeal carcinoma samples as a prognosis marker for a better disease-free survival [27]. A very recent case (September 2021) was published by a team of researchers from Canada: the authors described a 78-year-old Chinese male diagnosedwith a primary EBV-positive, HPV-negative nasopharyngeal carcinoma, in the left tonsil, as being the first case of primary tonsil EBV-positive nasopharyngeal carcinoma described in the literature [28]. Our results for EBV detection are comparable with the data published by the above-mentioned studies. HHV8 was detected in all 11 head and neck Kaposi Sarcoma cases, analyzed by immunostaininging [29]. HHV8 was also detected, by other authors, in oropharyngeal cancers [30] by immunohistochemistry, and in rare cases of nasopharyngeal cancer, by histology and immunohistochemical markers [31]. Our two cases detected positive for HHV8 are in line with the above cited papers. HHV6 and HHV7 were tested by PCR, and an oral shedding of HHV6 and HHV7 was observed, during all the radiotherapy period for the tested patients [32]. HHV6 was found in 16.7% of adult-onset recurrent respiratory papillomavirus biopsies, by RT PCR [33]. Our results show that we detected these viruses in a higher proportion than the ones published by other authors: 46.15% HHV7, and 30.76% HHV6. [PERSON] et al. quantified the DNA/CMV in plasma of patients with HNSCC under radiotherapy and chemotherapy, and for their study patients they found that plasma positivity was associated with a higher risk of death [34]. [PERSON] et al. analyzed the CMV antigen in salivary glands tumors (70.6%), and they found that CMV might play an important role in the development of these kinds of tumors [35]. [PERSON] et al. recently published a case of a HIV-negative patient with oropharyngeal cancer in remission, which developed a buccal ulcer positive for CMV. The authors conclude that opportunistic infections with CMV can affect the quality of life and mortality [36].