Despite substantial progress to combat tuberculosis (TB), the infection remains a major global health problem. The global prevalence of tuberculosis infection (TBI) is closer to one-fourth, with large regional differences (Cohen et al., 2019). China is one of the countries with a high burden of TB and TBI, with approximately 350 million persons living with the infection (Gao et al., 2015; Houben and Dodd, 2016). Preventing TBI and stopping progression from infection to disease are critical for reducing TB incidence to the levels envisaged by the End TB Strategy (Uplekar et al., 2015).

It is estimated that over 81.6 million people living at high altitude in the world (Tremblay and Ainslie, 2021). High-altitude regions encompass several countries with high TB burdens, including Peru, Bolivia, India, China, Nepal, and Ethiopia. These areas collectively contribute approximately 8-12% of global TB cases (Gelaw et al., 2019). The synergistic interaction between high-altitude environmental factors and socioeconomic disparities creates a unique epidemiological landscape for tuberculosis transmission. Chronic hypobaric hypoxia, a defining characteristic of these regions, induces evolutionary adaptations such as EPAS1/HIF-2α polymorphisms in Tibetan populations (Tashi et al., 2017). While these genetic adaptations enhance survival in oxygen-deprived environments, they may simultaneously attenuate critical hypoxia-induced inflammatory responses essential for MTB containment (Bucşan et al., 2022). Specifically, these adaptations compromise macrophage antimicrobial functions, potentially facilitating bacterial persistence (Li et al., 2018). These biological vulnerabilities are exacerbated by structural challenges including geographic isolation, socioeconomic barriers to care-seeking, and suboptimal housing conditions that promote airborne transmission (Wang et al., 2023). This multifaceted challenge demands integrated public health strategies that simultaneously address the distinct pathophysiology of high-altitude TB and the systemic healthcare access limitations faced by these populations. Although several epidemiological studies have investigated the association between altitude and tuberculosis incidence, these studies were conducted at relatively early stages, lacked studies in very high altitude (VHA; >3500 m) areas, and seldom focused on tuberculosis infection (Gardiner et al., 1923; Mansoer et al., 1999; Olender et al., 2003; Pérez-Padilla and Franco-Marina, 2004; Vargas et al., 2004; Vree et al., 2007).

Tibet is a provincial autonomous region of China that is located in the Qinghai-Tibet Plateau at an average altitude >4000 m above sea level and ~95% of its population is of Tibetan ethnicity (Wu, 2001). The unique hypobaric hypoxia environment and other related factors may have caused the suboptimal health status among the Tibetan population. Physiological, anthropological and genomic research over the last 50 years has characterized the phenotypic and genotypic differences between Han Chinese and Tibetans living at altitude (Weitz et al., 2016; He et al., 2017). For TB, it has greater burdens than other areas of China. In 2023, the notified incidence of TB in Tibet was 107/100,000 population, ranking third in China (Wang et al., 2024).

To address this evidence gap, the Tibet Autonomous Region CDC implemented a population-based, cross-sectional survey to quantify TBI prevalence across altitudinal gradients and evaluate potential altitude-dependent variations in key epidemiological risk factors.

Table 1 shows detailed information about the population assessed. Of 7320 eligible participants, 6533 actually participated including questionnaire and C-TST, with a response rate of 89.3% ( Supplementary Figure S1 ). Overall, roughly 55% of participants were female and 12% were 60 years or older in this survey which was higher than the general population of Tibet (Office of the Leading Group of the State Council for the Seventh National Population Census). About 95% are Tibetan. Nearly 60% of participants had a BCG scar. A tenth of participants reported having ever smoked and about a third had ever consumed alcohol. BMI distribution showed that almost a quarter of the participants were overweight or obese.

In total, 1016 (15.5%) of 6533 persons who underwent C-TST had an induration of 5 mm or more, 121 (1.9%) participants had blistering, necrosis, or lymphangitis. In multivariate analysis, the following factors demonstrated significant associations with positive C-TST results: engagement in heavy physical work (farmer and worker), male gender, increasing age, presence of BCG scars, elevated BMI (≥25 kg/m²), residence at high altitude, history of tobacco use, low physical exercise levels, reported insomnia, and previous pneumonia diagnosis ( Figure 1 ). The Boruta feature importance analysis identified altitude of residence as the most determinant predictor of C-TST positivity, with an inverse relationship observed - higher altitudes were associated with decreased risk of positive test results ( Figure 2 ).

The interaction analyses revealed significant modification effects of both smoking status (interaction p=0.0065) and BCG vaccination (interaction p=0.028) on the altitude-C-TST positivity association ( Figure 3 ). Notably, while each 500-meter elevation increase was associated with a 31.9% reduced C-TST positivity risk among non-smokers (β=-6.37×10⁻⁴, p<0.001), no significant altitude effect was observed in smokers (β=-9.56×10⁻⁵, p=0.6). Similarly, BCG-vaccinated individuals demonstrated a stronger inverse altitude association (34.7% risk reduction per 500m, β=-6.93×10⁻⁴, p<0.001) compared to unvaccinated individuals (17.7% reduction, β=-3.54×10⁻⁴, p=0.03), representing a 50% attenuation of the altitude protective effect. These findings suggest that both smoking and lack of BCG vaccination may compromise the beneficial physiological adaptations to high-altitude environments.

In subgroup analyses stratified by altitude, distinct epidemiological patterns were observed. Among HA residents, C-TST positivity was significantly associated with occupational exposures, whereas in VHA populations, positivity was predominantly linked to individual physiological factors, including age, sex, BMI, BCG vaccination status and smoking history. Comorbidity profiles also exhibited altitude-dependent variations: in HA regions, pneumonia and type 2 diabetes were significant risk factors for C-TST positivity, whereas in VHA areas, pneumonia, chronic pharyngitis, and hypertension demonstrated stronger associations with positive test results ( Supplementary Figure S2 ).

After excluding active TB cases, the crude prevalence of TBI in this survey population was 15.7% (95% CI: 14.7%-16.4%). Geospatial analysis revealed the highest crude prevalence rates in central counties of the province ( Figure 4 ). Mediation analysis indicated that the observed inverse relationship between study site altitude and crude TBI prevalence was mediated by per capita land space, accounting for a significant proportion of this association (IE= -6.29e-05, p=0.04, Supplementary Figure S3 ). Using best-practice analytical methods with statistical adjustment, the estimated TBI prevalence among individuals aged 5 years and older in Tibet Autonomous Region was 20.7% (95% CI: 14.3%-33.0%) ( Table 2 ). The estimated prevalence of TBI was approximately 1·3 times higher in men than in women, and was highest in people aged 60 years or older. Prevalence was lower in stratum 3 than in strata 2 and 1. The estimated prevalence of TBI in the HA region was nearly 3 times higher than that in the VHA region, but the reported incidence of TB in the HA region was only 1.5 times that in the VHA region ( Table 2 ).

High-altitude regions face unique public health challenges characterized by extreme environmental conditions, pervasive poverty, limited healthcare access, and low health awareness, all of which exacerbate disease burdens. These areas often experience heightened vulnerability to infectious diseases like TB. Studies are encouraged in countries with considerable high-altitude populations, to understand how the environmental stress (physiological and social) of high altitude impacts physiology, adaptation, health, and disease (Tremblay and Ainslie, 2021). In response, we launched a prospective cohort study in Tibet to investigate MTB infection and disease progression, with this work representing the baseline survey. The results reveal that both the TBI prevalence and TB incidence in the VHA region are lower than those in the HA region, and distinct risk factors exist between the two regions.

Our findings demonstrate a significant inverse association between altitude and TBI risk, with prevalence decreasing as elevation increases. This observation aligns with several previous studies conducted in high-altitude regions. For instance, research in the Peruvian Andes reported a lower TBI prevalence at elevations above 3,000 meters compared to lowland areas (Tashi et al., 2017). Similarly, a study in Turkey found reduced TB infection rates in high-altitude zones (Tanrikulu et al., 2008). However, our findings demonstrate that smoking status and BCG vaccination significantly modify the protective association between altitude and C-TST positivity. The complete attenuation of altitude’s protective effect among smokers suggests that tobacco-induced pulmonary alterations may override the natural protective mechanisms conferred by high-altitude adaptation (Liu et al., 2025). Furthermore, the 50% weaker protective effect observed in BCG-unvaccinated individuals implies that vaccine-induced immunity may synergize with physiological responses to high-altitude hypoxia, potentially enhancing host defense against MTB infection (Zhu et al., 2021). Our mediation and mixed-effects analyses demonstrate that the observed altitude-dependent reduction in TBI risk is likely mediated through two primary pathways: decreased population density at higher elevations (β = -0.22, p=0.02), which reduces transmission opportunities, and altitude-specific physiological adaptations that enhance host defenses. Notably, we found BCG vaccine effectiveness showed significant altitude-dependent enhancement (15% increase per 500m elevation, p=0.015), suggesting high-altitude residents may develop more robust vaccine-induced immunity, potentially through hypoxia-driven immune modulation (HIF-1α upregulation) and vitamin D-mediated pathways (Fabri et al., 2011; Burtscher et al., 2020; Calvo, 2020). These findings provide mechanistic insights into how both environmental and biological factors collectively contribute to the unique TBI epidemiology in high-altitude populations.

Subgroup analysis revealed distinct TBI risk patterns between HA and VHA regions. Occupational exposure was the predominant risk factor in HA areas, with manual laborers demonstrating a 3.5-fold increased risk. In contrast, VHA populations exhibited physiology-driven susceptibility, where advanced age (aOR=1.71), elevated BMI (aOR=1.35), and smoking (aOR=1.39) emerged as primary determinants. These epidemiological distinctions call for targeted interventions: workplace-focused measures, including ventilation standards and pre-employment screening in HA regions, versus prioritized screening of high-risk populations in VHA areas. The comorbidity patterns associated with TBI risk also differed significantly between HA and VHA regions. In HA areas, diabetes mellitus remained a significant risk factor for TBI (Lee et al., 2017), while VHA populations exhibited a distinct association between hypertension and TBI. Previous a systematic review indicated that there was a significant correlation between altitude and the prevalence of hypertension among inhabitants of Tibet, corresponding to a 2% increase in prevalence for every 100 m increase in altitude (Mingji et al., 2015). Moreover, in VHA regions, the respiratory system becomes particularly vulnerable (Talaminos-Barroso et al., 2021), where smoking demonstrates an altitude-dependent increase in TBI risk, while exercise may paradoxically diminish its protective immune benefits, likely due to compounded respiratory stress under hypoxic conditions. Therefore, these findings advocate for altitude-adapted comorbidity screening protocols, particularly emphasizing rigorous cardiopulmonary monitoring in VHA regions.

This study also has some limitations. First, because many men and young individuals were not at home during the survey period, there was a gap between our research population and the general population in Tibet. Although we adjusted the population structure based on the model, the estimated prevalence may still be biased (Floyd et al., 2013). After statistical adjustments, best-practice analytical methods estimated the prevalence of TBI among people aged 5 years or older in China to be 20.5% (95% CI 14.2%- 32.8%), which was at least 30% higher than the models without adjustment. The gap may be due to the difference between the sampled population structure and the general population structure and sampling. The proportion of female participants (55%) was higher than that in the general population (49%). In addition, simple random sampling increases the probability of communities or villages with low population density being selected, which may reduce the crude rate. Second, the WHO recommended the EC skin test, a newer MTB antigen-based skin test as an alternative to diagnose TBI, in 2022. However, it has not been extensively validated in people living in plateau areas. Although randomized diagnostic trials have shown that this positive determination criterion demonstrated high specificity and sensitivity of cutoff of ≥5 mm redness or induration, it might not be the most appropriate cutoff for this study population.

Our findings demonstrate that while the overall prevalence of TBI in Tibet (20.5%) is comparable to China’s national average, the region exhibits critical altitude-specific epidemiological characteristics that demand focused attention. These findings highlight the need to increase investment in marginalized populations, particularly high-altitude communities that face complex environmental and physiological challenges, to identify unique barriers to tuberculosis elimination and develop targeted tuberculosis control strategies.