As a common chronic infectious disease caused by Mycobacterium tuberculosis (MTB), TB is a major public health problem in developing countries and a great threat to human health (Glaziou et al., 2018). According to the latest World Health Organization report, there is an estimated 9.9 million new cases of TB worldwide in 2021, and China is one of the countries with high TB burden (World Health Organization, 2021). Studies had shown that although a quarter of the world’s population were infected with MTB, while only 10% of those infected population eventually develop active TB (Moonan et al., 2020; Luo et al., 2022). This suggested that a large number of the infected population might be naturally resistant to MTB infection, and host genetic susceptibility played non-negligible roles in TB development. In recent years, many studies had provided important clues to explore the roles of host genetic polymorphism in susceptibility to pulmonary TB (PTB) (Zhang et al., 2021; Chen et al., 2021). However, these results could only explain part of the heritability of PTB, and further exploration of the relationship between gene variations and PTB susceptibility would help to comprehensively understand the pathogenesis of MTB infection.

N6-methyladenosine (m6A) was the most common and enriched mRNA post-transcriptional modification, and widely distributed in prokaryotes and eukaryotes (Roundtree et al., 2017; Chen and Wong, 2020). M6A modification related enzymes mainly composed of m6A methyltransferase (METTL3, METTL14), m6A demethylase (FTO, ALKBH5) and m6A-binding protein (IGF2BP1, YTHDF1) (Hua et al., 2020). The m6A modification played important roles in a variety of biological processes, including mRNA translation, cell differentiation, immune response (Zhao et al., 2017) and dysregulated m6A modification was closely linked to various diseases, including autoimmune diseases, cancer (Han et al., 2019; Wang et al., 2021). The genetic variation in m6A-related genes might affect m6A modification level through altering the RNA sequence of target sites or key flanking nucleotides, and was considered to be a novel, important perspective for studying the disease susceptibility (Hua et al., 2020; Lin et al., 2020; Ruan et al., 2021).

FTO and ALKBH5 were the main m6A demethylases, which could reverse RNA methylation by oxidizing the N-methyl group at m6A site to a hydroxymethyl group, and were involved in the development of immune diseases and cancer (Wu et al., 2021; Deng et al., 2022). In addition, several typical single nucleotide polymorphisms (SNP) in FTO (rs9939609, rs1121980), ALKBH5 (rs9913266, rs12936694) genes had been reported to affect the risk of gastric cancer, type 2 diabetes, autoimmune thyroid disease (Song et al., 2021; Binh et al., 2022; Li et al., 2022). PTB was also considered an immune-related infectious disease because the susceptibility of host to MTB was affected by the immune function (Feng et al., 2014; Liu et al., 2017). We speculated that FTO, ALKBH5 gene variations might be involved in the immune response to MTB, and a few studies also suggested a key role of FTO gene variation in the risk of PTB (Feng et al., 2014; Naderi et al., 2017). It was possible that ALKBH5 gene SNP also had a similar role in PTB, but relevant study was limited. Therefore, we performed this case–control study to assessed the association of ALKBH5, FTO genes polymorphisms, as well as their levels, with PTB susceptibility in a Chinese population.

Except for environmental factors, socioeconomic factors, host gene variation was also an important risk factor for PTB development. In recent years, many studies had proposed the potential contribution roles of m6A-related genes SNPs in the risk of multiple diseases, especially cancer (Lin et al., 2020; Ruan et al., 2021; Chen et al., 2021). Herein, we selected two main m6A demethylases (ALKBH5, FTO), and attempted to investigate whether multiple SNPs in ALKBH5, FTO genes were linked to the risk of PTB. According to our results, no SNP was found to be associated with the susceptibility to PTB, while ALKBH5, FTO genes variations might play certain influence on some clinical manifestations of PTB patients. It should be noted that this was the first study for revealing the abnormally reduced levels of ALKBH5, FTO in patients with PTB.

At present, the studies regarding ALKBH5, FTO genes were mainly focused on cancer, and ALKBH5, FTO had been identified as important malignant regulators of tumor cell phenotypes and therapeutic responses (Lan et al., 2020; Zhang et al., 2021). Some previous studies speculated that SNPs located at the key regulatory sites of ALKBH5, FTO might affect the development of disease by influencing host gene expression (Ren et al., 2021; Li et al., 2022). On the other hand, ALKBH5, FTO genes exerted an important role in the immune response to MTB infection, their gene variations might also affect PTB risk. The FTO gene rs9939609 polymorphisms was significantly associated with obesity in different populations, including Chinese population (Tan et al., 2008). Similar to other malnutrition states, obesity was also known to impair immune function, affect leukocyte counts and alter cell-mediated immune responses (de Heredia et al., 2012). Moreover, the impaired immune function could result in to an increased susceptibility of the host to pathogens such as MTB, coxsackie virus (Karlsson and Beck, 2010). Hence, it had been speculated that the FTO gene polymorphism might play important roles in the risk of immune-related human infectious diseases such as PTB, and the roles on PTB occurs through obesity-related immunocompetence. Given that the roles in FTO-SNPs in immune-related infectious, we have reason to believe that ALKBH5 gene SNPs exert a similar role. Feng et al. found that compared with the common genotype TT, FTO rs9939609 AA genotype was strongly associated with a significantly increased risk of PTB in Chinese population (Feng et al., 2014). However, rs9939609 variant showed no significant difference in genotype, allele frequencies between PTB patients and controls in Iranian population in another study (Naderi et al., 2017). These two studies differ in the number of participants, race, experimental methods, which might lead to inconsistent results. In this study, we similarly did not find any significant association between rs9939609 variant and PTB susceptibility, and the same results were also observed for FTO rs6499640, rs8047395, rs1121980. Compared with single SNP, haplotypes tend to had a stronger power of predicting disease-related genes (Chen et al., 2018), and we detected a significantly lower frequency of FTO GAAA haplotype in PTB patients. This partially supported the hypothesis that FTO genetic polymorphisms might affect the host’s susceptibility to PTB, while the specific mechanisms need to be further explored by well-designed studies. Our study was the first to assessed the association of ALKBH5 gene SNPs (rs8400, rs9913266, rs12936694, rs4925144) with PTB susceptibility, however no significant association was found. This provided some clues for future study on the role of ALKBH5 gene variation in PTB development.

In addition to disease susceptibility, several host gene variations had been shown to affect the occurrence of the clinical manifestations of PTB patients. Li et al. revealed that lncRNA NEAT1 rs3825071 variant was significantly related to the sputum smear-positive among PTB patients (Li et al., 2022). Similarly, our results provided strong evidence of significant association between rs8400 AA genotype, A allele, rs9913266 A allele frequencies and sputum smear-positive. This suggested that these SNP might contribute to the hierarchical management of PTB patients to develop different and appropriate therapeutic schedule. In addition, the results of another study suggested that CYP27A1 rs17470271, rs933994 variants might affect the development of leukopenia, drug resistance in PTB patients, respectively (Zhang et al., 2021). In the present study, we found that rs8400, rs4925144 variants respectively affected the risk of hypoproteinemia, leukopenia. Furthermore, ALKBH5 rs9913266 A allele was closely related to the occurrence of DILI, and FTO rs8047395 AA genotype, A allele frequencies were significantly decreased in PTB patients with drug resistance. Based on these results, it was reasonable to speculate that ALKBH5, FTO genes variations were closely related to the development process of PTB, and these SNPs could be used as potential indicators for some specific clinical manifestations of PTB patients.

It was of a great necessity to investigate the m6A demethylase expression level, and some studies found that the abnormally expressed level of ALKBH5, FTO were related to the pathogenesis of many diseases (Zhang et al., 2017; Guimarães-Teixeira et al., 2021; Deng et al., 2022). The study by Deng et al. showed that ALKBH5 level was downregulated in both PBMCs and T cells in systemic lupus erythematosus patients, and the overexpression of ALKBH5 could promoted apoptosis and inhibited the proliferation of T cells (Deng et al., 2022). The decreased FTO mRNA levels had been proved to be associated with a poor prognosis of renal cell carcinoma (36). Hence, this case-control study was conducted to detect the ALKBH5, FTO mRNA levels in PBMC from PTB patients, and revealed that the ALKBH5, FTO expression levels were significantly decreased in PTB patients in comparison with controls. This provided key evidence to confirm that ALKBH5, FTO were involved in the occurrence of PTB, and their levels could be used as a potential, auxiliary indicator for PTB diagnosis, which should be explored and verified by further studies. Our study also showed that ALKBH5 level was significantly associated with drug resistance, and FTO level was significantly associated with sputum smear-positive, ESR in PTB patients. These significant associations between ALKBH5, FTO levels and the clinical manifestations were contributed to understand the role of ALKBH5, FTO in the development of PTB.

The strengths of this study included the good design and novel topic. We simultaneously examined ALKBH5, FTO gene variations, expression levels, and analyzed their association with PTB patients. However, some limitations should not be neglected. First, the sample size could be increased to get more reliable results, although our study appeared to be adequate, and the statistical power of every SNP exceeded 85%. Second, we could not analyze the interaction between genetic variations and environmental factors due to no environment information. Third, other SNPs with important functions might not be selected. The studies with multiple centers, large samples were needed to accurately assess the mechanism of ALKBH5, FTO genes in the pathogenesis of PTB in the future.

In summary, a significantly decreased frequency of GAAA haplotype in PTB demonstrated that FTO gene polymorphisms might associated with PTB susceptibility, whereas ALKBH5 SNPs had no predictive value in evaluating PTB susceptibility. Several SNPs in ALKBH5, FTO genes were associated with the clinical features of PTB patients such as drug resistance, DILI, hypoproteinemia, and ALKBH5, FTO levels were significantly lower in PTB patients. This study will help to further understand the pathogenesis of PTB, which could contribute to formulated more appropriate clinical treatment measures.

The original contributions presented in the study are included in the article/ Supplementary Materials . Further inquiries can be directed to the corresponding author.

This study was approved by the Ethics Committee of Anhui Medical University (20200250). The patients/participants provided their written informed consent to participate in this study.

T-PZ and H-ML designed the study. H-ML conducted the experiment. RL performed the statistical analyses. L-JW participated in sample collection. T-PZ and H-ML drafted the manuscript. All authors contributed to the article and approved the submitted version.