Yu et al. BMC Microbiology (2020) 20:75 https://doi.org/10.1186/s12866-020-01733-7 RESEARCH ARTICLE Open Access Dysbiosis of saliva microbiome in patients with oral lichen planus Fei Yan Yu1†, Qian Qian Wang2†, Miao Li2, Ya-Hsin Cheng3, Yi-Shing Lisa Cheng4, Yu Zhou1, Xi Yang2, Fang Zhang1, Xuejun Ge2, Bin Zhao5 and Xiu Yun Ren2* Abstract Background: Oral microbiota is not only important for maintaining oral health but also plays a role in various oral diseases. However, studies regarding microbiome changes in oral lichen planus (OLP) are very limited. To the best of our knowledge, there has been only two studies investigating salivary microbiome changes in OLP. Therefore, the purpose of this study was to identify the characteristic microbial profile in the saliva of OLP patients, with or without erosive lesions, and compare that with recurrent aphthous ulcer (RAU), a common oral immunological disorder that also shows multiple erosive/ulcerative lesions. Whole saliva samples were collected from 20 patients with OLP (erosive E, n = 10 and non-erosive NE, n = 10), 10 patients with RAU (U) and 10 healthy controls (C). DNA was extracted from the saliva samples, and the 16S rDNA gene V4 hypervariable region was analyzed using Illumina sequencing. Results: We obtained 4949 operational taxonomic units (OTUs) from the V4 region in all saliva samples. Community composition analysis showed a clear decreased relative abundance of genera Streptococcus and Sphingomonas in saliva from RAU patients when compared to the other three groups. Relative abundance of Lautropia and Gemella were higher in E group, whereas relative abundance of Haemophilus and Neisseria were higher in NE group when compared to C group. Abiotrophia and Oribacterium were higher in OLP (combining E and NE groups), while Eikenella and Aggregatibacter were lower when compared to C group. There was statistically significance in α-diversity between E and RAU groups(p < 0.05). Significant differences in β-diversity were detected in bacteria between E and C; NE and C; as well as E and NE groups. The LDA effect size algorithm identified the g_Haemophilus might be the potential biomarker in NE group. Conclusions: We found that salivary microbiome in erosive OLP was significantly different from that found in RAU; and these changes may be related to the underlying disease process rather than presence of ulcerative/erosive lesions clinically. In addition, our findings in bacterial relative abundance in OLP were significantly different from the previously reported findings, which points to the need for further research in salivary microbiome of OLP. Keywords: Oral lichen planus, Salivary microbiome, 16S rDNA, High-throughput sequencing Background Oral lichen planus (OLP) is a common oral mucosal dis- ease with or without accompanying lesions in skin, nails, * Correspondence: [email protected] eyes, or urogenital tissue [1]. OLP is regarded as a †Fei Yan Yu and Qian Qian Wang contributed equally to this work. chronic T-cell-mediated inflammatory disease, exhibiting 2Department of Periodontology, Shanxi Medical University School and a higher prevalence in women than men, especially those Hospital of Stomatology, No. 63, New South Road, Yingze District, Taiyuan, Shanxi 030001, People’s Republic of China over fifty years of age [2, 3]. The lesions typically spread Full list of author information is available at the end of the article throughout the oral mucosa and usually involve the © The Author(s). 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Yu et al. BMC Microbiology (2020) 20:75 Page 2 of 12 posterior buccal mucosa bilaterally [3]. There are essen- distribution already, without an underlying mucocutane- tially two forms of oral lesions in OLP: erosive, and re- ous disease. Therefore, a group of recurrent aphthous ul- ticular or non-erosive [4], with the latter being the most ceration (RAU) patients was also included as a common. However, erosive lesions are more significant comparison to further test if the microbiota changes oc- to the patients because they usually cause sore mouth curred in OLP would be unique to OLP. RAU is a com- and may affect the ability to eat and maintain good oral mon oral mucosal disease also characterized by multiple hygiene [5]. Notably, it is common for patients to have oral ulcers, a feature similar to erosive OLP. Both OLP both reticular and erosive lesions. Although the etiology and RAU are T cell-mediated immunological disorders; of OLP is still unknown, various factors have been sug- however, the pattern of the clinical lesions and the gested to contribute to the pathogenesis, including sys- pathogenesis mechanism of RAU are different from OLP temic medications [6, 7], dental restorative materials such except the common clinical feature of multiple ulcera- as amalgam [8] and composite or resin-based materials tions [14]. Using the Illumina MiSeq sequencing, a high- [9], cinnamon-containing products, and microorganisms. throughput sequencing technology, we examined the Whether oral microbial changes in OLP disease state relative abundances in different bacteria, the diversity of is not clear, but some studies have confirmed the correl- bacteria, and the dominant bacteria species in different ation between OLP and oral microorganism [10]. Born- study groups. The results reveal a distinctive bacterial stein et al. [5] investigated microbial in OLP patients signature for OLP, which is different from RAU, and with non-erosive/asymptomatic lesions and found that shed a light to the different underlying immunological bacterial counts for Capnocytophaga sputigena, Eikenella mechanism of these two diseases. corrodens, Lactobacillus crispatus, Mobiluncus curtisii, Neisseria mucosa, Prevotella bivia, Prevotella intermedia, Results and S. agalactiae at the sites of OLP lesions are signifi- Characterization of study subjects and microbial analysis cantly higher when compared to the same sites in con- The demographic and clinical characteristics of the four trol subjects. In addition, bacterial counts for study groups: OLP patients with erosive lesions (E), OLP Bacteroides ureolyticus, Dialister species, Staphylococcus patients without erosive lesions (NE), patients with oral haemolyticus,andStreptococcus agalactiae were also sig- aphthous ulcer (U) and healthy controls (C) were shown nificantly higher in OLP gingival lesions when compared in Table 1. All subjects were free of hypertension, dia- to those found in the contralateral unaffected sites betes, smoking cigarette habit and alcohol-drinking. By within the same patient. Since these OLP patients were t-test, there was no significant differences in age, gender, asymptomatic, the differences in microbial species and and saliva pH, respectively, among groups (p < 0.05). quantity found in OLP lesions are most likely due to A total of 1,173,674 clean reads were obtained from OLP disease process but not patients’ inadequate hy- the analysis of saliva collected from all subjects. An aver- giene maintenance caused by soreness/discomfort from age of 9236 high quality sequences between 200 and these OLP lesions. Although the clinical types (erosive 400 bp in length were clustered into 4949 operational or reticular) and presence or absence of symptoms were taxonomic units (OTUs) with a 97% identity of coverage not specified, Ertugrul et al. [11] also found that the pro- for each group. portion of Aggregatibacter actinomycetemcomitans, Por- phyromonas gingivalis, Prevotella intermedia, Tannerella Alpha diversity of microbiome in each group forsythia, and Treponema denticola in total bacteria were The salivary bacterial community in E, NE, U and C higher in subgingival plaque samples taken from subjects groups was first analyzed quantitatively by Shannon and having periodontitis and OLP than those having peri- Simpson diversity indices, respectively. No significant odontitis without OLP. The results of these studies sug- differences were observed in the comparison between gest that there may be correlation between the microbe NE and C or E and C (Fig. 1a and b). The index value of and OLP. To the best of our knowledge, there has been the U group was significantly higher than those found in only two studies investigating salivary microbiome in any of the other three groups. OLP in the literature so far [12, 13], and the changes of Samples from each group were further analyzed by oral microbiome in OLP remains unclear. rarefaction curves, which showed that the number of Therefore, the purpose of this study was to identify OTUs was also higher in the saliva samples from U the characteristic microbial profile in the saliva of OLP group than those found in NE, E or C groups, respect- patients with or without erosive lesion. The difference ively (Fig. 1c). between erosive OLP and non-erosive OLP is the pres- The rarefaction curves indicated that almost entire bac- ence of multiple ulcerative lesions in the clinical presen- terial population in our samples of OLP saliva was repre- tation of OLP. It is likely that the presence of oral sented in our results based on 97% good’s coverage.