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Question continued: A variety of notions exist about MTB’s evolution and interaction with the humans. While on one hand some evolutionary biologists in TB believe it was a disease of the cattle that got transferred to humans during domestication, others believe it was the humans who gave  it to the cattle. Is it really possible that MTB colonised humans sometime early in the evolutionary stages and co-evolved within us?

Tirumalai Kamala’s answer:

It’s eminently possible that MTB co-evolved with humans. Problem is co-evolution is difficult to prove.

First, some commonly used techniques and abbreviations in TB genomics (1, 2).

Additionally, MTC: Mycobacterium tuberculosis complex; includes Mycobacterium tuberculosis (MTB), M. canettii, M. africanum, M. bovis, M. microti, M. pinnipedii, M. caprae
LSP: Large Sequence Polymorphisms; also called chromosomal deletions; RD, i.e. Regions of Difference. Number and distribution of deletions offer a TB strain-specific genomic signature used to build maps of phylogenetic relationships.
WGS: Whole Genome Sequencing

Is Human-MTB co-evolution possible to prove?
Why is co-evolution difficult to prove? According to Woolhouse (3), at least three conditions are necessary for co-evolution

  • Relevant traits need to vary in both host and pathogen. For e.g., resistance and infectivity.
  • Such traits should have reciprocal effects on the fitness of host and pathogen populations.
  • Combination of host-pathogen genotypes determines the outcome of their interactions.

Sounds logical and reasonable, no? Yet not so simple in the case of TB. Why?

  • TB lacks classic virulence factors, e.g. toxins.
  • On the one hand, molecular approaches are starting to reveal TB’s inner workings in terms of what helps establish successful infections and what doesn’t but they’re being evaluated largely in irrelevant mouse model studies.
  • OTOH, the host side of the equation remains largely a blank slate, especially the human side.
  • What’s an effective human immune response against TB? Not known so human fitness traits relevant for TB disease resistance remain unknown as well.
  • As a result, TB virulence studies are an abundant literature morass that serve to confuse and distract.

Geographic patterns could provide indirect evidence. In particular, pathogen and host variants co-existing in particular geographic regions suggest local adaptation (4). Before getting to MTB’s geographic patterns, let’s examine the players and their defining features, and unresolved issues of evolutionary TB studies.

The Mycobacterium tuberculosis complex (MTC)

  • Each species of MTC behaves as an ecotype, i.e. has a preferred mammalian host species (5).
  • Have more than 99.9% nucleotide similarity (6)
  • Identical 16S rRNA sequence (7).
  • Little or no evidence of ongoing horizontal gene transfer (HGT) between current MTC members (8).
  • MTC thus interpreted as being relatively young pathogens, clonal progeny of an ancestral strain that underwent an evolutionary bottleneck ~20000 to 35000 years ago (9).
  • MTC presumably evolved from M. prototuberculosis. Which brings us to M. canetii, assumed to be the extant (surviving) progeny of M. prototuberculosis.

Mycobacterium canetti

  • Georges Canetti first isolated it in 1969 from a 20-year old French farmer suffering from pulmonary TB (10).
  • Isolated from patients living in or presumably infected in East Africa.
  • <100 strains identified thus far.
  • All from humans with TB disease, mostly from East Africa.
  • Striking morphological difference in culture, smooth rather than rough which is the characteristic of MTB.
  • Mosaic housekeeping genes, with individual segments found in MTC members, suggesting HGT in MTC ancestral species (11, 12)
  • In a comparison study, 5 M. canetti strains grew 2 to 3 times faster both in vitro and in mouse, but persisted poorly in mouse tissues (13) compared to standard MTB strains, suggesting they’re less pathogenic.

Which came first, human or cow TB?
Until the 1990s, human TB was assumed to be a by-product of animal, specifically cattle, domestication as in a jump of cow TB (M. bovis) to human (MTB). Why? Two reasons.

  • One, archeological identification of extrapulmonary TB-like deformities in Egyptian skeletal remains. Extrapulmonary TB is common in humans infected with M. bovis.
  • Two, M. bovis is a generalist with a broader host range beyond cattle. MTB is a specialist, infecting mostly humans.

Thus, for a while the prevailing narrative was that M. bovis spread to humans from animals with man’s domestication of animals in the Neolithic era (14), i.e. the generalist M. bovis evolved into the specialized MTB.
However, whole genome analysis showed instead that MTB is more ancient than M. bovis (15).

  • M. bovis has lost several genes still present in all other MTC lineages.
  • M. bovis genome is ~60000bp smaller than MTB genome (16).
  • M. bovis does not contain any unique gene.
  • Comparative genomics show animal MTB strains are related to human disease-causing MTB (9, 17).
  • MTB strains associated with major epidemics such as the ‘Beijing’ and ‘Haarlem’ strains share the TbD1 deletion, absent in M. africanum, MTB’s closest phylogenetic neighbor, as well as in M. bovis.
  • Instead MTC animal strains such as M. bovis and M. africanum share the RD9 delection, absent in MTB strains.
  • Animal MTB strains cluster together while human MTB strains represent different human populations (18,19, 20).
  • MTB is clonal, unable to repair genomic deletions by recombination.
  • M. bovis is rarely found in the archeological record (1), found only in one group of Iron Age (4th century BC to 4th century AD) remains consisting of both humans and their animals (21).
  • Thus M. bovis now assumed to originate from a familial species with a larger genome (22, 23, 24).

For these reasons, M. bovis and MTB are now assumed to have evolved independently (see below).

Limitations of paleo-epidemiology, -pathology, -microbiology in reconstructing human-TB history

  • TB does not produce toxins, classic virulence factors.
  • Virulence factors typically help a pathogen transmit from human to human.
  • Pulmonary TB and ensuing lung cavities give MTB its best chance for human-to-human transmission.
  • Connection to paleo studies? Soft tissues degrade quickly and are generally unavailable for paleological studies, except in the case of some mummies. With soft tissues gone, what remains for analysis? Bone.
  • Yet bone TB, like other extrapulmonary TB, has two major limitations w.r.t. reconstructing human-TB history
    • One, bone TB constitutes less than 5% of total TB, i.e. provides a window into a tiny, perhaps irrelevant, sliver of historical human-TB interactions.
    • Two, bone TB reduces rather than maximizes human-to-human transmission, an evolutionary dead end for TB.

Biological relevance of ancient TB strains identified in ancient human bones is thus an open question, a point often overlooked/minimized/ignored.

Ancient TB strains identified in ancient human remains are instead relevant as an archeological record of MTC strains.

  • For example, TB strains in ancient Egyptian remains include those without the TbD1 deletion (25). Today such strains, presumed ancient relics of the MTB lineage, are restricted to the Far East around the Pacific Rim.
  • Today the most widespread TB strain is TbD1-deleted.
  • TbD1-deleted TB have been found in 9000-year old remains in the pre-pottery Neolithic site of Atlit-Yam in the Eastern Mediterranean (26).

The evolutionary bottleneck hypothesis for TB

  • Ancestral TB strains had extensive horizontal gene transfers (HGTs).
  • Then came a proposed bottleneck, ~35000 years ago (see below).
  • Post-bottleneck, MTC strains appear to follow a linear evolutionary pattern with key clonal deletions (1, 14).
  • Yet, the oldest MTC was found in cranial frontal bone of a 500,000 Homo erectus fossil excavated in Turkey (27).
  • Thus we come to the current controversial notion that TB’s progenitor predated modern humans.
  • Ancestral TB, M.prototuberculosis, is thus speculated to have emerged 3 million years ago.

Current TB lineages and how they evolved. Africa is the likeliest origin for MTB for following reasons

  • All major MTC lineages have been found in Africa (18, 23).
  • Unique presence of M. canetti and other smooth TB bacilli.

What M. africanum could teach about human-MTB co-evolution

  • M. africanum causes ~50% human TB in West Africa.
  • No animal reservoir identified thus far.
  • M. africanum contacts much less likely to get active disease compared to MTB contacts (28), even though both transmit equally well to contacts.
  • Millions of West Africans were forcibly taken to the Americas during slavery and yet, M. africanum never established itself in the Americas. Why? Outcompeted by MTB (29)?
  • The IRGM-261TT polymorphism in Ghana protects against Euro-American MTB but not against M.africanum (30).

Is M. africanum a red herring or is the observed human-M. Africanum dynamic an indicator of human-MTB co-evolution? An open question at present but one where compelling data (see above) suggests the latter, and not the former.

How TB Virulence relates to Human-TB co-adaptation

  • What is the purpose of virulence? To maximize pathogen transmission.
  • How is virulence defined? Pathogen infection reduces host fitness.
  • TB presents two major problems w.r.t virulence definition.
    • One, TB disease ranges from pulmonary TB, the most common form to TB meningitis and extrapulmonary TB. Which disease type maximizes human-to-human transmission? Data since the classic studies of Koch and Canetti (31) suggest that transmission is related to lung cavities (32), for the simple reason that the caseating necrosis at the center of lung tuberculous granulomas is the optimal breeding ground for TB bacilli, which can reach up to a billion in a single cavity. Thus, fast rates of disease progression may not necessarily be to the benefit of TB. For example, TB meningitis kills more quickly than pulmonary TB so a TB strain associated with the former would be considered more virulent but such strains may not transmit more efficiently.
    • Two, TB lacks classical virulence factors (toxins) so TB strain virulence is defined using a variety of animal models, ranging from passably suitable (rabbit) to entirely unsuitable (mouse).
  • TB virulence studies primarily focus on bacterial growth rates and quantification, and host cell cytokine release.
  • Animal model studies also use a variety of routes of infection, doses and TB strains.
  • In vitro human studies add further variables. Some use whole blood, others PBMC (peripheral blood mononuclear cells), yet others, blood-derived monocytes (selected by targeting the CD14 surface molecule; such selection itself is capable of modulating the cells’ responses).
  • Comparisons of TB strain virulence differences across such studies? Apples and oranges.
  • Extrapolation from such studies to propensity for human-to-human transmission? Also apples and oranges.
  • Back on planet earth, how is TB virulence measured in humans (or should be)?
    • Likelihood and rate of disease progression.
    • Type of TB disease (pulmonary >TB meningitis = extrapulmonary TB for transmissibility).
    • # of TB bacilli in the sputum, presence and # of lung cavities.
    • During active TB disease, patients are mobile and do not need respiratory support because disease is typically confined to one lobe of one lung (33). Chronic low-grade disease and long-term cough without life-threatening pathology is thus the most optimal adaptive outcome for MTB, i.e. capacity for human-to-human spread.

Thus, most modern TB studies are hobbled because their virulence measurements do not assess transmission potential.

  • TB and human anti-TB immunity offer compelling clues for co-evolution (33).
  • Most pathogens specialize in immune escape through some form of antigenic variation, i.e. changing the sequence of (protein) antigens that are targets of immune responses. MTB does the opposite.
  • MTB antigens that are targets of human T cells are the products of the most conserved and invariant MTB genes (34). Rather than evade immune responses, MTB seems to goad them, akin to waving a red flag at a bull. Or does it?
  • HIV/AIDS co-infection is a natural experiment that also reveals human-TB co-adaptation (35). How? HIV/AIDS culminates in impaired immune responses. Logic dictates that HIV-infected TB patients should be more contagious, yet they are less (36).
  • Less than 1% of TB-infected persons proceed to active disease and death yet TB-infected humans number in the billions, the vast majority of whom never develop active TB disease.
  • Ancient co-travelers, could the optimal human-TB state instead be an entente, TB the goad, the human immune response the muscle? Appropriate response then means neither tissue damage nor TB elimination, with TB using the human’s appropriate immune response to learn to stay within bounds, bounds meaning a dormant state within the lung, except for a little perturbation every now and then so TB could spread through coughs.
  • Could TB carriage offer selective advantages to the human carriers? (37) Maybe protect against malaria/CMV, for example?
  • What about TB carriers who proceed to active disease? Collateral damage of the evolutionary human-TB bargain? After all evolutionary agreements necessitate trade-offs for the involved parties.
  • Final word goes to humans. Non-compliance to multi-drug anti-TB antibiotic therapy is today a wild card driving, nay disrupting, the human-TB consensus. Nothing new there. After all, human agency associated with the Industrial Revolution and its attendant appallingly unhygienic and teeming urban human conglomerations disrupted ancient human-TB equilibrium, driving the TB epidemics of the 19th and 20th centuries.


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