A Role for Mycobacterium in Alzheimer’s Disease?

26 October 2020

We looked forward to Lowry’s “Alzheimer's disease: protective effects of Mycobacterium vaccae, a soil-derived mycobacterium with anti-inflammatory and anti-tubercular properties, on the proteomic profiles of plasma and cerebrospinal fluid in rats” publication in the Journal of Alzheimer’s Disease [1] as an often-overlooked possibility towards Alzheimer’s disease (AD) genesis.

The thought that AD might be of a mycobacterial etiology has recently gained traction, but has been too little expressed. Mawanda and Wallace simply cite that “amyloidopathy—a condition characterized by elevated levels of serum amyloid and by amyloid deposition and aggregation in tissues—is a frequent occurrence in several acute and chronic systemic inflammatory conditions, especially chronic infections like tuberculosis.” [2]. Several studies have already shown a “neuroprotective affect” using anti-mycobacterial antibiotics [3,4], and evidence suggests that patients aged 50-64 with infection with M. tuberculosis have a significantly higher risk of dementia [5].

And now still other evidence presents itself.

The discovery that the apolipoprotein (APOE) ε4 allele happens to be the strongest genetic risk for the most common form of AD led in turn to the consideration that the identification of the noncoding or microRNAs that control APOE as well as brain lipid metabolism might provide both a way to identify AD diagnostically and a way to treat it. But true understanding of genetic test results also requires attention to potential detail to avoid inaccurate results by exclusion. For example, APOE ε4 alleles themselves are known to show a distinct increase in tuberculosis (TB) [6]. And should this finding carry through to the various microRNAs established to be important to AD, still another argument can be made for the mycobacterial origins of AD.

The discovery of the first microRNA (miRNA) over 20 years ago has ushered in a new era in molecular biology. miRNAs are attractive molecules to be considered as one of the blood-based biomarkers for neurodegenerative disorders such as AD. And searching for non-invasive AD biomarkers is currently one of the most rapidly growing areas in AD research [7]. miRNA profiles have been extensively studied as potential bio-indicators for AD in blood, cerebrospinal fluid (CSF), and brain tissue. However, due to the high variability between the reported data, stemming from the lack of methodological standardization and the heterogeneity of AD, narrowing the field of the many miRNA to just a few of the most promising biomarker candidates has been slow to realize. Of special interest in this respect were two recent studies: first a large, 2019 review by Nagaraj et al. which found only 3 miRNAs: miRNA-146a, miRNA-125b, and miRNA-135a that were consistently reported in AD blood, CSF, and brain tissue [8]. This was after doing an extensive literature review that showed that out of 137 miRNAs found to be altered in AD blood, 36 have been replicated in at least one independent study, and out of 166 miRNAs reported as differential in AD CSF, 13 have been repeatedly found. But only 3 miRNAs were consistently reported as altered in three analyzed specimens: blood, CSF, and the brain (hsa-miRNA-146a, hsa-miRNA-125b, hsa-miRNA-135a).

Also, just prior to the Nagaraj paper, Yang’s study, which also found miRNA-135, in addition to miRNA-193b and miRNA-384, as potential early biomarkers for the early diagnosis of AD [9].

What will become obvious here, is that all of the proposed miRNA markers from both the Nagaraj and Yang studies find their miRNA counterparts in similar biomarker diagnostics for TB and the mycobacteria.

The increase in miRNA-384, noted by Yang to be indicative of AD, is also significantly increased in mice with experimental autoimmune encephalomyelitis (EAE) using complete Freund’s adjuvant containing heat-killed Mycobacterium tuberculosis H37Ra. The use of heat-killed TB is mandatory in the induction protocols of many experimental models of EAE [10].

Both Nagaraj’s and Yang’s studies point to the importance of miRNA-135a in AD diagnosis with Yang pointing to the fact that in his study serum levels of miRNA-135a, increased significantly like miRNA-384 in patients with mild cognitive impairment and AD. Also, it was miRNA-135 that yielded the highest diagnostic accuracy in discriminating mild cognitive impairment patients from normal controls. All of Yang’s preferred AD markers in the serum (miRNA-135, miRNA-193b, and miRNA-384) were found previously to regulate the expression of amyloid-β protein precursor (AβPP) which eventually becomes amyloid-β (Aβ), whose amyloid fibrillar form is the primary component of amyloid plaques found in the brains of AD patients.

Yang proved miRNA-135a to be abnormally expressed in AD [9], but its value toward detecting tuberculous meningitis (TBM) had to wait for Lin et al. [11] who both confirmed its miRNA-135a’s diagnostic value for TBM as well as its worth in indicating prognosis. To Lin, miRNA-135a exerts a vital effect on the development, progression, as well as prog¬nosis of TBM. In fact, miRNA-135a exerts a vital effect on the development, progression, as well as prog¬nosis of TBM [11].

TBM remains a serious, often undetected health threat. Chronic neuropsychological sequelae may occur even after appropriate treatment, often in the form of cognitive impairment as seen in AD [12], though the mechanisms that lead to and mediate cognitive and behavioral outcomes in TBM remain unclear [13]. Due to its hidden onset and long course of disease, clinical manifestations and laboratory tests lack specificity [14]. It is because the diagnosis of TBM mainly rests on pathogen detection, which has the limitations of long cycle, poor sensitivity, and low specificity that delay in diagnosis and treatment result in the high fatality and high morbidity of TBM [15]. Therefore, it is of great significance to explore methods for the early diagnosis of TBM [15]. One of these is through the diagnostic value of miRNA-135a for TBM and its association with prognosis.

While Yang struggled to explain why AD biomarker miRNA-193 decreased significantly in the serum of those with mild cognitive impairment and AD, Lyu posited that such a dip in miRNA-193 could be expected as a manifestation of the standard differentially expression of miRNA-193 were such mental impairments due to active infection with Mycobacterium tuberculosis (Mtb) [17], a disease which Lyu validated miRNA-193 to be specifically expressed in.

As for Nagaraj et al. [8] who found only 3 miRNAs: miRNA-146a, miRNA-125b, and miRNA-135a that were consistently reported in AD blood, CSF, and brain tissue these also find their counterparts in other studies.

microRNA-146a and microRNA-193
Besides being premier AD marker candidates, Miotto et al. found both Nagaraj’s miRNA-146 and Yang’s miRNA-193 to be among the 15 miRNAs identified as a signature for discriminating between healthy controls and patients with pulmonary TB [18].

Despite the existence of various anti-mycobacterial therapies, TB remains one of the world’s major causes of illness and death. Approximately one third of the world’s population is thought to be infected with Mtb, and more than 9 million develop “active” TB each year [19]. As the first line of host defense, macrophages are responsible for intracellular killing of Mtb. Paradoxically, they are also its principal target cells. It has been well-accepted that Mtb has evolved a serious of strategies to subtly modulate host immunity and create a microenvironment favoring its replication and growth—of which, regulation of miRNAs expression is considered as an important one. miRNAs are non-coding, single-stranded RNAs of ∼22 nt in length that regulate gene expression. In mycobacteria-infected macrophages, miRNA-146a is not only robustly increased, but leads to a higher mycobacterial burden in infected macrophages. miRNA-146 expression significantly increased in mouse macrophages post Mycobacterium bovis Bacille Calmette-Guérin infection (unpublished data), suggesting that miRNA-146a may play a role in TB-associated inflammation [20].

Besides being proposed as a test to differentiate between healthy individuals and those with pulmonary TB, miRNA-146a has been proposed as both a biomarker for human and cattle TB [21]. miRNA-146a can increase up to 20-fold in mycobacterial infection, which promotes mycobacterial survival in macrophages by suppressing nitric oxide production [22].

TB is an inducer of miRNA-125b [23]. And Fu et al. found miRNA-125b increased in the serum of TB patients [24].

Similarly, lipomannan from virulent Mtb stimulated a high expression of miRNA-125b in human macrophages [25]. Lipomannan is a glycolipid in TB’s cell wall which plays a critical role in the pathogenesis of TB [26].

The etiology of AD is still of course unknown, but it is good to see the substantial progress through different avenues such as the paper by Lowry and colleagues [1] to better define it.

Lawrence Broxmeyer, MD

[1] Loupy KM, Lee T, Zambrano CA, Elsayed AI, D'Angelo HM, Fonken LK, Frank MG, Maier SF, Lowry CA (2020) Alzheimer’s disease: protective effects of Mycobacterium vaccae, a soil-derived mycobacterium with anti-inflammatory and anti-tubercular properties, on the proteomic profiles of plasma and cerebrospinal fluid in rats. J Alzheimers Dis, doi: 10.3233/JAD-200568.
[2] Mawanda F, Wallace R (2013) Can infections cause Alzheimer's disease? Epidemiol Rev 35, 161-180.
[3] Umeda T, Ono K, Sakai A, Yamashita M, Mizuguchi M, Klein WL, Yamada M, Mori H, Tomiyama T (2016) Rifampicin is a candidate preventive medicine against amyloid-β and tau oligomers. Brain 139, 1568–1586.
[4] Iizuka T, Morimoto K, Sasaki Y, Kameyama M, Kurashima A, Hayasaka K, Ogata H, Goto H (2017) Preventive effect of rifampicin on Alzheimer disease needs at least 450 mg daily for 1 year: an FDG-PET follow-up study. Dement Geriatr Cogn Disord Extra 7, 204-214.
[5] Peng YH, Chen CY, Su CH, Muo CH, Chen KF, Liao WC, Kao CH (2015) Increased risk of dementia among patients with pulmonary tuberculosis: a retrospective population-based cohort study. Am J Alzheimers Dis Other Demen 30, 629-634.
[6] Farivar TN, Moud BS, Sargazi M, Moeenrezakhanlou A (2008) Apolipoprotein E polymorphism in tuberculosis patients. J Appl Sci 4, 719-722.
[7] Lista S, Faltraco F, Prvulovic D, Hampel H (2013) Blood and plasma-based proteomic biomarker research in Alzheimer's disease. Prog Neurobiol 101-102, 1-17.
[8] Nagaraj S, Zoltowska KM, Laskowska-Kaszub K, Wojda U (2019) microRNA diagnostic panel for Alzheimer's disease and epigenetic trade-off between neurodegeneration and cancer. Ageing Res Rev 49, 125-143.
[9] Yang TT, Liu CG, Gao SC, Zhang Y, Wang PC (2018) The serum exosome derived microRNA-135a, -193b, and -384 were potential Alzheimer’s disease biomarkers. Biomed Environ Sci 31, 87-96.
[10] Billiau A, Matthys P (2001) Modes of action of Freund's adjuvants in experimental models of autoimmune diseases. J Leukoc Biol 70, 849-860.
[11] Lin J, Cao C, Jin M (2020) Expression of miR-135a in cerebrospinal fluid of patients with tuberculous meningitis and its association with clinicopathological features. Int J Clin Exp Med 13, 2863-2870.
[12] Kalita J, Misra UK, Ranjan P (2007) Predictors of long-term neurological sequelae of tuberculous meningitis: a multivariate analysis. Eur J Neurol 14, 33–37.
[13] Chen HL, Lu CH, Chang CD, Chen PC, Chen MH, Hsu NW, Chou KH, Lin WM, Lin CP, Lin WC (2015) Structural deficits and cognitive impairment in tuberculous meningitis. BMC Infect Dis 15, 279.
[14] Heemskerk AD, Bang ND, Mai NT, Chau TT, Phu NH, Loc PP, Chau NV, Hien TT, Dung NH, Lan NT, Lan NH, Lan NN, Phong le T, Vien NN, Hien NQ, Yen NT, Ha DT, Day JN, Caws M, Merson L, Thinh TT, Wolbers M, Thwaites GE, Farrar JJ (2016) Intensified antituberculosis therapy in adults with tuberculous meningitis. N Engl J Med 374, 124-134.
[15] Erdem H, Ozturk-Engin D, Tireli H, Kilicoglu G, Defres S, Gulsun S, Sengoz G, Crisan A, Johansen IS, Inan A, et al. (2015) Hamsi scoring in the prediction of unfavorable outcomes from tuberculous meningitis: results of Haydarpasa-II study. J Neurol 262, 890-898.
[16] Thao LTP, Heemskerk AD, Geskus RB, Mai NTH, Ha DTM, Chau TTH, Phu NH, Chau NVV, Caws M, Lan NH, Thu DDA, Thuong NTT, Day J, Farrar JJ, Torok ME, Bang ND, Thwaites GE, Wolbers M (2018) Prognostic models for 9-month mortality in tuberculous meningitis. Clin Infect Dis 66, 523-532.
[17] Lyu L, Zhang X, Li C, Yang T, Wang J, Pan L, Jia H, Li Z, Sun Q, Yue L, Chen F, Zhang Z (2019) Small RNA profiles of serum exosomes derived from individuals with latent and active tuberculosis. Front Microbiol 10, 1174.
[18] Miotto P, Mwangoka G, Valente IC, Norbis L, Sotgiu G, Bosu R, Ambrosi A, Codecasa LR, Goletti D, Matteelli A, Ntinginya EN, Aloi F, Heinrich N, Reither K, Cirillo DM (2013) miRNA signatures in sera of patients with active pulmonary tuberculosis. PLoS One 8, e80149.
[19] Lawn SD, Zumla AI (2011) Tuberculosis. Lancet 378, 57-72.
[20] Li S, Yue Y, Xu W, Xiong S (2013) MicroRNA-146a represses mycobacteria-induced inflammatory response and facilitates bacterial replication via targeting IRAK-1 and TRAF- 6. PLoS One 8, e81438.
[21] Iannaccone M, Cosenza G, Pauciullo A, Garofalo F, Proroga YT, Capuano F, Capparelli R (2018) Milk microRNA-146a as a potential biomarker in bovine tuberculosis. J Dairy Res 85, 178-180.
[22] Li M, Wang J, Fang Y, Gong S, Li M, Wu M, Lai X, Zeng G, Wang Y, Yang K, Huang X (2016) microRNA-146a promotes mycobacterial survival in macrophages through suppressing nitric oxide production. Sci Rep 6, 23351. Erratum in Sci Rep 6, 24555 (2016).
[23] Behrouzi A, Alimohammadi M, Nafari AH, Yousefi MH, Rad FR, Vaziri F, Siadat SD (2019) The role of host miRNAs on Mycobacterium tuberculosis. ExRNA 1, 40.
[24] Fu Y, Yi Z, Wu X, Li J, Xu F (2011) Circulating microRNAs in patients with active pulmonary tuberculosis. J Clin Microbiol 49, 4246e51.
[25] Rajaram MV, Ni B, Morris JD, Brooks MN, Carlson TK, Bakthavachalu B, Schoenberg DR, Torrelles JB, Schlesinger LS (2011) Mycobacterium tuberculosis lipomannan blocks TNF biosynthesis by regulating macrophage MAPK-activated protein kinase 2 (MK2) and microRNA miR-125b. Proc Natl Acad Sci U S A 108, 17408-17413.
[26] Fukuda T, Matsumura T, Ato M, Hamasaki M, Nishiuchi Y, Murakami Y, Maeda Y, Yoshimori T, Matsumoto S, Kobayashi K, Kinoshita T, Morita YS (2013) Critical roles for lipomannan and lipoarabinomannan in cell wall integrity of mycobacteria and pathogenesis of tuberculosis. mBio 4, e00472-12.


We are grateful to Dr. Broxmeyer’s Letter to the Editor regarding the article by Loupy et al. [1]. The idea that Alzheimer’s disease might have, in some cases, a mycobacterial etiology merits further study. Tuberculosis infection remains one of the most common infections worldwide, with an estimated 2 billion people infected with M. tuberculosis [2]. In the United States, a total of 9,287 new cases of tuberculosis were reported in 2016 [3]. However, based on tuberculin skin tests (TST) in 2011-2012, it was estimated that 4.7% of the US population was TST positive (compared to the point estimate in foreign-born persons of 20.5%) [4]. Also of concern are nontuberculous mycobacterial (NTM) infections, chronic infections that appear to be increasing in the United States, particularly among older age groups [5]. This is of potential interest given the similar patterns of geographic variation of NTM infection [5] and Alzheimer’s disease [6], which could reflect shared environmental risk factors, such as the abundance of NTM, including potential pathogenic M. mucogenicum/phocaicum, M. avium complex, M. fortuitum complex, and M. abscessus complex, in municipal water sources [7]. Of these, M. avium was recently classified as belonging to an emended genus, Mycobacterium (“Tuberculosis-Simiae” clade), whereas M. abscessus was classified as Mycobacteroides gen. nov. (“Abscessus-Chelonae” clade), and M. vaccae, M. mucogenicum, and M. fortuitum were classified as Mycolicibacterium gen. nov. (“Fortuitum-Vaccae” clade), highlighting the phylogenetic diversity of mycobacteria, and foreshadowing potential differences in their impacts on the human host.

Mycobacteria are intracellular parasites, and, thus, may utilize a well-documented “Trojan horse” mechanism to enter the brain, wherein mycobacteria enter the central nervous system following infection of host immune cells [8-12]. The extent to which NTM can enter the central nervous system in this manner and the effects on host neurophysiology remain to be determined.

It is also possible that chronic mycobacterial infection increases risk of Alzheimer’s disease through signaling of peripheral inflammation from the periphery to the central nervous system though afferent signaling pathways. We have shown that intratracheal administration of a heat-killed preparation of M. vaccae NCTC 11659 (coupled to nitrocellulose beads in order to localize the immune activation to the airways) in mice acutely activates a subset of serotonergic neurons located within the interfascicular part of the dorsal raphe nucleus (DRI) [13]. This subset of serotonergic neurons projects to the hippocampus and prefrontal cortex and we have suggested previously that it modulates affective, cognitive, and stress resilience functions [14]. Importantly, we have also shown that infection of mice with live, virulent M. tuberculosis (H37Rv) activates DRI serotonergic neurons 3 days and 7 days following infection, but this activation is absent 14, 21, and 28 days after infection, suggesting adaptation of the DRI serotonergic system following chronic infection [15]. In convergence with Dr. Broxmeyer’s Letter to the Editor, we have shown an essential role for microRNA 135 (miR135a) for chronic stress resiliency, antidepressant efficacy, and serotonergic activity [16]. Dysregulation of an miR135a-serotonin signaling pathway following chronic mycobacterial infection could conceivably impact the etiology and pathophysiology of Alzheimer’s disease.

Christopher A. Lowry, PhD, and
Kelsey M. Loupy, BA, MS

[1]     Loupy KM, Lee T, Zambrano CA, Elsayed AI, D'Angelo HM, Fonken LK, Frank MG, Maier SF, Lowry CA (2020) Alzheimer’s disease: protective effects of Mycobacterium vaccae, a soil-derived mycobacterium with anti-inflammatory and anti-tubercular properties, on the proteomic profiles of plasma and cerebrospinal fluid in rats. J Alzheimers Dis, doi: 10.3233/JAD-200568.
[2]    U.S.Department of Health and Human Services Centers for Disease Control and Prevention (2019) Epidemiology of Tuberculosis. Atlanta, Georgia.
[3]    Schmit KM, Wansaula Z, Pratt R, Price SF, Langer AJ (2017) Tuberculosis - United States, 2016. MMWR Morb Mortal Wkly Rep 66, 289-294.
[4]    Miramontes R, Hill AN, Yelk Woodruff RS, Lambert LA, Navin TR, Castro KG, LoBue PA (2015) Tuberculosis infection in the United States: Prevalence estimates from the National Health and Nutrition Examination Survey, 2011-2012. PLoS One 10, e0140881.
[5]     Winthrop KL, Marras TK, Adjemian J, Zhang H, Wang P, Zhang Q (2020) Incidence and prevalence of nontuberculous mycobacterial lung disease in a large U.S. managed care health plan, 2008-2015. Ann Am Thorac Soc 17, 178-185.
[6]    Kirson NY, Meadows ES, Desai U, Smith BP, Cheung HC, Zuckerman P, Matthews BR (2020) Temporal and geographic variation in the incidence of Alzheimer's disease diagnosis in the US between 2007 and 2014. J Am Geriatr Soc 68, 346-353.
[7]    Gebert MJ, Delgado-Baquerizo M, Oliverio AM, Webster TM, Nichols LM, Honda JR, Chan ED, Adjemian J, Dunn RR, Fierer N (2018) Ecological analyses of mycobacteria in showerhead biofilms and their relevance to human health. MBio 9, e01614-18.
[8]    van Leeuwen LM, Boot M, Kuijl C, Picavet DI, van Stempvoort G, van der Pol SMA, de Vries HE, van der Wel NN, van der Kuip M, van Furth AM, van der Sar AM, Bitter W (2018) Mycobacteria employ two different mechanisms to cross the blood-brain barrier. Cell Microbiol 20, e12858.
[9]    Schwerk C, Tenenbaum T, Kim KS, Schroten H (2015) The choroid plexus-a multi-role player during infectious diseases of the CNS. Front Cell Neurosci 9, 80.
[10]    Jain SK, Tobin DM, Tucker EW, Venketaraman V, Ordonez AA, Jayashankar L, Siddiqi OK, Hammoud DA, Prasadarao NV, Sandor M, Hafner R, Fabry Z; NIH Tuberculous Meningitis Writing Group (2018) Tuberculous meningitis: a roadmap for advancing basic and translational research. Nat Immunol 19, 521-525.
[11]    Cain MD, Salimi H, Diamond MS, Klein RS (2019) Mechanisms of pathogen invasion into the central nervous system. Neuron 103, 771-783.
[12]    Dando SJ, Mackay-Sim A, Norton R, Currie BJ, St John JA, Ekberg JA, Batzloff M, Ulett GC, Beacham IR (2014) Pathogens penetrating the central nervous system: infection pathways and the cellular and molecular mechanisms of invasion. Clin Microbiol Rev 27, 691-726.
[13]    Lowry CA, Hollis JH, de Vries A, Pan B, Brunet LR, Hunt JR, Paton JF, van Kampen E, Knight DM, Evans AK, Rook GA, Lightman SL (2007) Identification of an immune-responsive mesolimbocortical serotonergic system: Potential role in regulation of emotional behavior. Neuroscience 146, 756-772.
[14]    Hale MW, Lowry CA (2011) Functional topography of midbrain and pontine serotonergic systems: implications for synaptic regulation of serotonergic circuits. Psychopharmacology (Berl) 213, 243-264.
[15]    Hollis JH, Goosen K, Orozco H, Wilkinson A, Lightman SL, Hernández-Pando R, Rook GA, Lowry CA (2009) A murine model of acute and chronic bronchopulmonary infection with Mycobacterium tuberculosis: Delayed and persistent activation of serotonergic neurons within the brainstem raphe complex. Brain Behav Immun 23 (Suppl 2), S40-S41.
[16]    Issler O, Haramati S, Paul ED, Maeno H, Navon I, Zwang R, Gil S, Mayberg HS, Dunlop BW, Menke A, Awatramani R, Binder EB, Deneris ES, Lowry CA, Chen A (2014) MicroRNA 135 is essential for chronic stress resiliency, antidepressant efficacy, and intact serotonergic activity. Neuron 83, 344-360.