Have the special properties of bird mitochondria influenced the evolution of the avian lung?
John B. West

For the comparative physiologist, a fascinating question is why the lungs of birds and mammals have taken such different evolutionary paths. These two classes of vertebrates are the only ones capable of sustained high oxygen consumptions, and much of their physiology is similar, but they have radically different lungs. Birds have a through-put system of ventilation whereby the inspired air is taken in by air sacs and pumped directly through the gas-exchanging tissue (Duncker,1972). An analogy is the flow of air through the radiator of a car for heat exchange. By contrast, mammals have a reciprocating system of ventilation in which the inspired air is mixed with a large volume of alveolar gas, the result being a lower alveolar and arterial PO2. This means that the mammalian lung is inherently less efficient and it is not clear what lead to this evolutionary path being taken.

The two evolutionary paths probably began to diverge about 300 million years ago in the Carboniferous (Gauthier, 1986). The atmospheric oxygen concentration was much higher then than now, perhaps as great as 35 percent (Dudley, 1998). Such high oxygen levels result in increased generation of reactive oxygen species (ROS) which are potentially damaging to tissues (Turrens, 2003). The lung may be particularly vulnerable because of its extreme vascularity.

However birds and mammals differ greatly in their handling of ROS, most of which are generated by mitochondria (Turrens, 2003). Avian mitochondria produce far less ROS compared with mammalian (Barja, 2004), and furthermore, many bird tissues tolerate ROS better than their mammalian counterparts (Ogburn et al.,1998; Hickey, 2007).

In this respect, the reciprocating ventilation system of the mammalian lung confers a selective advantage because the alveolar gas-exchanging tissue is not exposed to the very high oxygen concentration of the inspired air, but to a lower value as a result of mixture with the alveolar gas.
By contrast, the gas-exchanging tissue in the avian lung with its lower ROS production and less susceptibility to damage by ROS, is able to tolerate the high concentrations of the pure inspired air. In other words the reciprocating ventilation pattern of mammals protects the gas-exchanging tissue from the potentially damaging high inspired oxygen levels, but the penalty is a lower alveolar and arterial PO2.

John West is in the School of Medicine, University of California San Diego
email: jwest@ucsd.edu

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"The mitochondrial genome is practically immune to decay over evolutionary time, despite a mutation rate that is 10 to 20 times faster than nuclear genes in vertebrates, and little, if any, recombination."

I read somewhere (trying to find the post) that the mutation rate is about 17 to 31 times faster than nuclear genes in vertebrates.

A recent article by Berlin et al. (2007) (also reviewed by Marais (Marais, 2007)) reported that the low mitochondrial variability in birds (relative to mammals) is most easily explained by Hill–Robertson effects on the W chromosome. These authors suggest selection and linkage between the W chromosome (in heterogametic females) and mtDNA act to lower avian mtDNA diversity. This may well be correct; however a well known biological phenomenon which appears unique to birds was excluded from these analyses, and may in fact provide a much simpler and more plausible explanation.
By comparison to mammals, birds are remarkable in several physiological parameters such as athletic performance, capacity to regenerate neuronal damage and their high respiratory efficiencies. However, perhaps more remarkable are several metabolic avian features. Birds have metabolic rates that are 2-2.5 fold greater, and estimated lifetime energy expenditures 15 times that of mammals of equivalent body mass. Not only do birds maintain body temperatures 3oC hotter than mammals, but many birds have blood glucose levels two to four times that of mammals, which in part defines them as diabetic (Holmes et al., 2001). This last feature occurs without the associated pathological complications seen in mammals and highlights considerable protection from oxidative damage.
With few exceptions, birds are also very long living relative to body mass, as many birds live three times longer, or more, than mammals of equivalent mass, and birds age slower at the cellular level (Holmes et al., 2001). Parrots may live over one hundred years, and even the tiny 5 gram Broad-billed hummingbird (Selasphorus platycercus) can live for 14 years (Holmes and Austad, 1995), while the maximum recorded life span of a 20 gram house mouse is only 4 years (Holmes et al., 2001). Zoo and wild tagging data also can mostly eliminate confounding influences of reduced predation through flight (Ricklefs, 1998), and the “exception species”, which are generally domesticated species, still live relatively long (e.g. chickens 20 years, Cortunix quail 6-7 years) (Holmes and Austad, 1995).
Just how birds achieve such exemplary resistance to age appears to be largely explained by different mitochondrial properties. In mammals 2-4% of all consumed oxygen is released as reactive oxygen species (ROS, e.g. superoxide O2-., hydroxyl radical OH-, hydrogen peroxide H2O2) from the electron transport system (ETS) complexes I and III. In health (and more so with numerous pathologies) mitochondria are generally the largest source of ROS (Turrens, 2003). Avian mitochondria produce considerably less ROS than mammalian mitochondria, with pigeon liver and heart mitochondria producing up to 10-fold less H2O2 than rats (this depends on respiration state, and ROS predominates from complexes I and III as O2-., which is converted to H2O2, (Barja, 2004; Herrero and Barja, 1997)). Furthermore, parrots and canaries show considerable resistance to lipid peroxidation relative to rodents, and isolated kidney epithelial cells from other long-lived bird species are much more resilient to pro-oxidant challenge by paraquat, H2O2 and 95% O2 with markedly less DNA damage than mouse cells (Ogburn et al., 1988). The differences between birds and mammals should not also be assumed to be adaptive. ROS release is not necessarily a byproduct of less efficient ETS function, as ROS provides feedback to cells and mitochondria (Barja, 2004), which explains why antioxidants can often be of detriment (Lane, 2005). These data do however illustrate increased ROS protection and lower ROS production in numerous bird species, which results in less DNA damage (Barja, 2004).
These avian physiological features were overlooked by Berlin et al. (2007). This is surprising given that the mitochondrial ETS is juxtaposed to mtDNA, and that ETS derived ROS makes the greatest contribution to mtDNA damage, and hence diversity (Barja, 2004). Admittedly ROS production would be a difficult parameter to measure for many species, although ROS production correlates more tightly with longevity than body mass (Barja, 2004). Therefore, correlation of π with mammalian and avian longevity may provide greater insight (note that this assumes the metabolic theory of ageing). Potentially the lower mitochondrial ROS output of bird mitochondria may provide another and potentially stronger physiological explanation for the low mtDNA diversity of birds.

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Scott A. Hodges1 and Justen B. Whittall2

1Department of Ecology, Evolution and Marine Biology, University of California, Santa Barbara, CA 93106, email: hodges@lifesci.ucsb.edu

2Department of Biology, 500 El Camino Real, Santa Clara University, Santa Clara, CA 95053, email: jwhittall@scu.edu

Though the term ‘coevolution’ was coined less than 50 years ago (Ehrlich and Raven 1964), over 100 years prior to that Darwin first described the process to account for the matching of tongue and tube lengths of bee species and the clover species they visited (Darwin 1859). He expounded on this process when describing the exceptionally long spur of Angraecum sesquipedale and predicted a moth pollinator with an equally long tongue (Darwin 1862). Darwin envisioned a ‘race’ in which long-tongued individuals had a fitness advantage in gaining food from exceptionally long-spurred flowers and long-spurred flowers had a fitness advantage in reproduction due to increased pollination. Without a countervailing force, the tongue length of the pollinator and the spur length of the plant would increase due to the reciprocating fitness advantages. When traits of two interacting species such as tongue and floral tube length match tightly, it is easy to assume that Darwin’s race, i.e., coevolution, is responsible.

We recently tested predictions of Darwin’s coevolutionary race model and an alternative, the ‘pollinator shift’ model, as applied to the evolutionary history of spurs among species in the North American Aquilegia clade (Whittall and Hodges 2007). We concluded that the pollinator shift model is a better fit for the majority of spur-length evolution, i.e., that species of Aquilegia have primarily evolved to fit the already established tongue-lengths of their pollinators. Ennos (2008), provides an excellent summary of our results, and then offers an alternative hypothesis to account for them that combines aspects of both Darwin’s coevolutionary race and the pollinator shift hypotheses. He envisions that pollinator shifts do occur but that the process begins when a new pollinator has a similar tongue length as the original pollinator, and thus both are effective pollinators. Then, Darwin’s race causes the evolution of longer tongues in the new pollinator class but the original pollinator does not evolve because of some countervailing evolutionary force (e.g., body size constraints). Darwin’s race then ensues between the plant and the new pollinator until another counter-balancing force prevents the lengthening of the new pollinator’s tongue. This counter-balancing force then causes stasis of tongue and spur length evolution until yet another pollinator class appears with a similar tongue length to carry on Darwin’s coevolutionary race.

Ennos’ model predicts pollinator shifts followed by stasis in spur length as does the pollinator shift model, which envisions that plants adapt to fit the pre-established tongue length of a pollinator with no reciprocal evolution by the pollinator. Ennos (2008) goes on to claim that the pollinator shift hypothesis “requires a rather implausible ecological scenario” because it requires that the original pollinator must be absent from the population for many generations. We disagree. Instead, the pollinator shift model only requires that in some part of the plant species’ range a longer-tongued pollinator becomes the predominant visitor, and the cause of most plant reproduction (Whittall and Hodges 2007). First, bees and hummingbirds do not have to have similar tongue lengths to be similarly effective at pollen transfer of ‘bee-adapted’ flowers (Castellanos et al 2003) as Ennos claims. Thus changes in the abundance of these pollinator classes could substantially change the selection regime for floral morphology (Castellanos et al 2003). Furthermore, temporal and spatial variation in pollinator abundances has been found to be substantial and is thus certainly not implausible (e.g., Herrera 1996; Herlihy and Eckert 2005; Price et al 2005; Brunet and Sweet 2006). Changes in pollinator abundance may occur over long time periods owing to a number of factors including changes in pollinator migration patterns, extinction of pollinators, changes in community composition, or adaptation of plants to a new habitat (see Thomson & Wilson 2008). Though such situations may be rare, this does not make them unimportant to plant evolution.

In addition to ecological changes, floral characters can affect the frequency of pollinator visitation. In Aquilegia, hawkmoths do not discriminate between control flowers and those with artificially shortened spurs, but variation in flower orientation and color have very large effects on visitation (Fulton and Hodges 1999; Hodges et al 2002). Similarly in two species of Mimulus, introgression of alternate QTL for flower color caused dramatic changes in the frequency of visitation by bees and hummingbirds (Bradshaw and Schemske 2003). Thus, changes in traits other than nectar spurs may precipitate shifts in pollinator visitation rates. Such changes would be especially likely to cause shifts when the new pollinator was already particularly abundant (Bradshaw and Schemske 2003). Once the setting for more frequent visitation is created then there would be very strong selection for other floral traits, such as nectar spur length, to become optimized for plant reproduction via the new pollinator (Thompson and Wilson 2008).

There are further aspects of Darwin’s coevolutionary race model, alone or as part of Ennos’s hybrid model, which seem to us to make it an unlikely explanation for the majority of spur-length evolution in the North American Aquilegia clade (or more generally, floral tube length evolution). For example, pollinators such as hummingbirds likely already had tongue lengths similar to those found today prior to their first association with bee-adapted Aquilegia. One clade of hummingbirds, called bee-hummingbirds, is comprised of two sister clades, one including all North American species and the other including primarily South American species (McGuire et al 2007). Phylogenetic analysis indicates that the ancestor of bee-hummingbirds occurred in South or Central America (McGuire et al 2007). This ancestor also likely had a bill morphology similar to extant bee-hummingbird species because species in both sister clades have similar bill morphology (around 1.5 to less than 2.0 cm (Colwell 2000)). Thus, bill size of hummingbirds was likely established prior to their invasion of North America and an interaction with Aquilegia (or other North American species that became adapted to hummingbirds). Furthermore, molecular dating indicates that bee-hummingbirds originated about 6 mya (Bleiweiss 1998) while the origin of hummingbird pollination in Aquilegia was likely to have been far more recent at around 1.7 mya or less (Hodges et al 2004; Kay et al 2006). Taken together these results strongly suggests that bill morphology was established prior to an association with Aquilegia and that it did not substantially change thereafter. We have already made a similar argument concerning hawkmoth tongue lengths (Whittall and Hodges 2007).

Another reason we doubt coevolution hypotheses as a major explanation for most spur length evolution is because the interaction between plant and pollinator is likely to be very asymmetrical. Hummingbirds and hawkmoths visit a large number of species (e.g., Grant 1983; Grant 1994) and thus they do not have a tight dependency on any one plant-species. For example, the longest spurred Aquilegia species, A. longissima, belongs to a guild of unrelated species from six distinct genera in the desert Southwestern US, all with floral tubes greater than 9 cm, which by and large share the same hawkmoth pollinators (Grant 1983; Grant and Grant 1983). Thus, the ability of the longest-tongued individuals to reach a bit more nectar from the longest-spurred individuals of a single species would provide a rather small fitness advantage for the pollinator, at best. In contrast, reproduction in plants is dependent on the visitation of pollinators and thus selection will be quite strong to optimize floral morphology to maximize pollen transfer. Thus, unrelated plant species visited by the same pollinator will likely converge on a similar floral morphology, such as tube or spur length. Similar arguments have been made during a previous discussion of Darwin’s coevolutionary race and the pollinator shift hypothesis (Jermy 1999). Finally, Ennos’ model requires a new species of hummingbird or hawkmoth for each transition to a new pollinator so that Darwin’s race can be run each time anew. Given the large number of shifts from bee to hummingbird and from hummingbird to hawkmoth pollinated plant species in North America (e.g., Grant 1994; Whittall and Hodges 2007; Thomson & Wilson 2008) and the much smaller number of hummingbird and hawkmoth species, this seems unlikely as a general explanation for the evolution of tube and tongue lengths.

While we believe that the pollinator shift hypothesis accounts for the majority of spur length evolution in Aquilegia, and likely many other species as well, some spur and tongue lengths may well be due to Darwin’s coevolutionary race. How would one identify such instances? We agree with Ennos that complementary, and more specifically, phylogenetic studies of plants and pollinators will be needed. If evolution among species or populations of both plants and pollinators indicate increasing lengths during the same time period and in the same geographical area, then this would provide strong evidence for Darwin’s race. Even still, it will be necessary to establish that the respective fitness advantages are due to the plant-pollinator interaction rather than some other factor (e.g., Wasserthal 1997; Borrell 2005).

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