1. INTRODUCTION

In the county of Cornwall in the South West of England there is a visitor centre called the Eden Project. Visitors can observe biomes, representatives of most of the world’s land ecosystems, the world’s largest greenhouse, and a variety of plant life. These ecosystems are enclosed in plastic domes and are thereby sealed off from the biosphere and the sky above. The Eden Project provides a suitable analogy of how most biologists regard the Earth, i.e. as an enclosed, individual unit, separated from space, just like these domes. Such “edenists” appear to be blissfully unaware of the obvious fact that the Earth is connected to space, that it is continually exposed to material incoming from the cosmos including meteors and organic dust, and that material is also exiting the planet and is ejected into space. Given this continual exchange between Earth and space, why shouldn't life also be incoming to and exiting from Earth? This is the possibility which we discuss here.

The theory of panspermia suggests that life did not originate on Earth, but instead came from space (Arrhenius 1908/2009; Hoyle and Wickramasinghe, 2000; Wainwright 2010; Wickramasinghe et al., 2009). The possibility that life originated here on Earth, but was supplemented by space-derived microorganisms also cannot be ruled out.

Another variant of panspermia, “neopanspermia” refers to the contemporary arrival of life from space (Wainwright, 2003). The idea that life originated from space has a long history, while the theory of neopanspermia is relatively new. However, the entire concept of panspermia, in its modern guise is based on the seminal work of Sir Fred Hoyle and Chandra Wickramasinghe (Hoyle and Wickramasinghe, 2000).

Until recently most of the work on panspermia has been theoretical. However, there is now laboratory evidence to support the view that microbes can be transferred across the cosmos, and which suggests that, at this moment, life is entering the Earth’s atmosphere from space.

One might imagine that the proposition that life is incoming to Earth from space could be easily be demonstrated, simply by sampling space at a height above the Earth where there is no possibility of contamination from below. One also might have assumed that NASA or another space agency would have looked for the presence of microbes in near space and would have determined at what height above the Earth’s surface they eventually peter out. This has not happened. Surprisingly, despite all the billions spent of space research we still do not know how high the Earth’s biosphere extends into space, nor have answers been provided to the apparently simple question- are microorganisms present in near space?

During the 1970s Russian workers found bacteria of the genera, Micrococcus and Mycobacterium, and fungi Circinella muscae, Aspergillus niger and Penicillium notatum, in the stratosphere, at heights of 48-77-km (Imshenetsky, 1978). However, no attempt was made to detect or isolate organisms above this height or to determine whether these microorganisms were coming in from space, or exiting from Earth.

More recently, the presence of bacteria and fungi in the stratosphere has been confirmed by Harris et al., (2002), Wainwright et al., (2003, 2004a,b), Narlikar, et al., (2003), Griffin (2004, 2008), Yang et al. (2008), Shivaji et al. (2009). Specifically, workers in Cardiff, Sheffield and India collaborated in sending balloons to the stratosphere to determine if a high cold biosphere exists at heights up to 41km (Harris et al., 2002, Wainwright et al., 2003, 2004). Using scanning electron microscopes, clumps of bacteria-like forms have been found at this height, either alone or associated with dust particles. Vital fluorescent stains, visualized using fluorescent microscopes have confirmed that these clumps consist of living cells, i.e. bacteria. Thus we have evidence that microbial life is readily isolatable from this region, the so-called “high cold biosphere” (Wainwright, 2008).

The obvious assumption is that these bacteria are carried to the stratosphere from Earth. It is has also been suggested that the transfer of bacteria from Earth to the stratosphere could act as means of “negative or geo-panspermia”, by which space is seeded with Earth bacteria. For example, Joseph and Schild (2010), argue that microbes are also commonly lifted into the stratosphere by monsoons and tropical storms, clinging to dust and debris. As pointed out by these authors, monsoons commonly funnel dust, water, gases, and pollutants to deep within the stratosphere where they stay aloft and circulate the globe for years. According to Joseph and Schild (2010) these microbes may then be ejected into space when Earth is struck by particularly powerful solar winds.

If correct, then the transfer of bacteria from Earth to the stratosphere may have played a major role in the evolution of life on Earth. Not all stratospheric bacteria would be ejected into space and most would likely fall back to Earth; but only after exposure to mutagenic radiation (notably UV) in the high cold biosphere. Specifically, as horizontal gene transfer is commonplace between archae, bacteria, eukaryotes, and viruses (reviewed in Joseph and Schild 2010), then mutagenic genes may also be transferred thereby triggering genetic experiments in evolutionary innovation.

If we assume that bacteria may also be ejected into space, such as following episodes of increased solar wind activity, we can also assume that bacteria on other planets may also be ejected into space. In fact, most of the bacteria that finds its ways into the stratosphere may not have come from Earth, but from space. This is because the tropopause acts a barrier to the free movement of particles above 17km thereby making such transfer from Earth to the stratosphere very difficult. For example, bacterial cell masses exceeding ten micron in diameter have been found in stratosphere samples. Since it seems unlikely that such large clumps could have been carried up from Earth even by monsoons, we assume that these are incoming from space and that they contribute to a mixed stratospheric biosphere made up of space and Earth-derived organisms (Wainwright et al., 2004, Wainwright, 2008). These incoming bacteria may continually join those bacteria which already exist on Earth.

In addition, bacteria incoming to Earth from space may also promiscuously exchange genes with indigenous bacteria, thereby enriching the Earth’s gene pool with genes from a cosmic gene pool (Wainwright et al., 2004).

2. THE HIGH COLD BIOSPHERE AND EVOLUTION

The highest point at which we know that microbial life exists is 77 km (Imshenetsky, 1978). However, we know nothing about the biology, if it exists, at heights above this. If microorganisms continue to be isolated as at even greater heights then there must come a point when it is acknowledged that they are incoming to Earth from space.

The existence of a stratospheric biosphere may have had an important effect on the evolution of life on Earth. Any bacteria transferred from Earth to the stratosphere will be exposed to high levels of mutagenic UV rays and other forms of radiation (Wainwright, 2008). Such exposure will induce mutations in bacteria passing into the stratosphere. The ability of UV to cause mutations in microbial genome has long been recognised, and is used in biotechnology to improve the production of important biochemicals like penicillin. Moreover, the ability of the effected host to survive in varied environments can also be impacted. For example, UV induced mutations in Lactobacillus enables them to survive high concentrations of sodium chloride and sodium nitrate (Arrahar and Itoh, 2002). Thus, mutations may pave the way for bacteria to colonize even toxic planets.

Therefore, naturally enhanced mutation in the stratosphere may speed up evolution rates in microbes which survive a period of UV exposure in this region and then return to Earth. This would also be true of microbes which arrive on Earth from space. Such mutagenesis will be far greater than that which occurs on Earth, where the amount of UV is reduced by the atmosphere, clouds, and the ozone layer.

The high cold biosphere may therefore act as a huge mutation- generator, a vast laboratory where new microbial genomes are created and returned to Earth where this new “information” can be promiscuously transferred to microbes which have not journeyed to the stratosphere. This process may be ongoing with microorganisms being continually returned to the stratosphere for a new dose of mutagenic radiation.

The acquisition of an atmosphere and ozone layer was absolutely essential for the development of complex multicellular life on Earth, thereby allowing life to explore and conquer diverse environments and to evolve and diversify. However, this protective layer also reduced the level of mutagenesis in prokaryotes and eukaryotes. Given that the protective ozone layer was not sufficiently established until around 540 million years ago, coupled with the explosion of complex life which followed, it could be said that UV-induced mutagenesis may have promoted microbial evolution and diversity for the first 4 billion years of Earth's history, but hindered eukaryotic evolutionary development.

Microbes have contributed crucial genes to the eukaryotic genome (Joseph 2009). As oxygen was pumped into the atmosphere by photosynthesizing microbes, a protective ozone layer was generated and greatly reduced microbial genetic mutagenesis. However, the creation of the ozone layer and increased oxygen is associated with evolutionary events leading up to the Cambrian Explosion, 540 million years ago. Therefore, it could be argued that microbes induced biospheric conditions which shifted the trajectory of evolution from microbes to eukaryotes. As most mutations kill or sicken the host, widespread UV induced mutations would not confer evolutionary advantages to eukaryotes. By contrast, as microbes make up the major biomass of this planet, the transfer of bacteria from Earth to the stratosphere would ensure that only a small sample of bacteria suffer massive UV exposure. Those who die have no effect on life on Earth, whereas those who survive with beneficial mutations can return to Earth and horizontally transfer these genes not just to other bacteria, but to pockets of eukaryotes. If those eukaryotes sicken and die, then bad mutations are eliminated. However, if the mutations are beneficial, then they would be naturally selected thereby triggering the next stage of evolution.

In fact, eukaryotes, including humans, may be the direct beneficiaries of UV induced mutation in bacteria.

It is interesting to note that Imshenetsky et al. (1978) isolated Penicillium notatum from the stratosphere, a fungus which Alexander Fleming was the first to show produces penicillin. We can speculate that perhaps this fungus was not originally a penicillin producer, or produced the antibiotic in trace amount. However, after a period of residence and mutation in the stratosphere it arrived on Fleming’s famous petri dish as a more active penicillin producer than before. Of course, we have no means of knowing if this happened, but it is by no means as outlandish an idea as it might first appear.

4. PANSPERMIA AND BACTERIAL RESISTENCE TO UV

Critics of panspermia often erroneous claim that it is impossible for naked bacteria to survive the transfer from space to Earth, because of problems related to ionising, and, particularly, UV radiation. These and other potential limitations have been discussed and accounted for in the numerous publications of Hoyle and Wickramasinghe (1982, 2000). These authors suggest that bacteria would be protected from the damaging effects of UV by layers of cosmic dust and carbonised outer cells of bacterial clumps.

The views of Hoyle and Wickramasinghe are now backed up by hard evidence. There is now considerable evidence demonstrating that bacteria can survive UV radiation, and a journey from Earth to space and back again (Burchell et al. 2004; Burchella et al. 2001; Horneck et al. 1994, 2002; Mastrapaa et al. 2001; Nicholson et al. 2000).

However, we suggest that it is not necessary for bacteria, travelling through space or present in the stratosphere, to be completely resistant to UV. What is important is that bacteria should not die and this we find can is the case, particularly for spore forming bacteria (i.e. notably species of Bacillus). Resistance to UV for even a short period of time would allow a bacterium to survive when the protective cosmic dust covering is partially exposed, until a new UV-protective dust cover is formed. In this way, a bacterium which can survive direct exposure to UV would be at a competitive advantage over one that was not; of course, if a bacterium remained permanently covered by an impenetrable UV-protective layer of cosmic dust or carbonised cells then it could remain viable in the absence of any native UV resistance.

By being permanently protected against UV exposure, or by being able to resist short periods of UV until any protective cover is reformed, bacteria should be able to survive even the extremely long time periods needed to cross space. Of course such protection would be even more readily available should the bacterium be present inside a meteorite.

Amber clay and various coals contain bacteria (Wainwright et al., 2009) thereby presenting the possibility that, as a result of past or future impact events, microbes from Earth could be ejected into space in a kind of reverse, or negative panspermia. It is generally assumed that microorganism incoming to Earth would necessarily possess unusual morphologies or physiological properties which adapt them for the period of transfer to Earth from space.

It is interesting to note that species of Deinococcus, which have the ability to survive exposure to ionising radiation, have recently been discovered in the low stratosphere (Yang et al., 2008); the temptation, in this case, is to assume that these bacteria must have come from space.

Bacteria are so small they can easily be protected by dust particles. A single bacterium, say a coccus or spore of 1-2 microns in size could be covered by a layer of protective UV dust. Even a small clump of bacteria could be under 5 micron in size, and thus completely protected by a dust particle. Therefore, bacteria clinging to dust, and especially those buried within meteors or in the heart of comets, could easily survive a journey through space, be they ejected from Earth, or incoming from other planets.

In the case of the panspermic establishment of the first life on this planet, only a single, viable bacterium need to survive and once on Earth it could multiply rapidly. In the absence of competition from other life forms could soon cover the planet. It is a remarkable thought then that all current life on this planet could have arisen from a single bacterium present in a small clump of dust which landed on a previously untenanted Earth.

5. MICROORGANISMS IN THE STATOSPHERE INCOMING, OUTGOING-OR BOTH?

Now that the existence of a stratospheric bacterial has been established the next obvious question is- from where do these organisms originate; from Earth or from space? The application of Occam’s razor suggests that since these are microbes are commonly found on Earth they must have an Earth origin. There exists however, the possibility that some, at least, originate from space and that a mixed population of bacteria exists in the stratosphere, some outgoing from Earth and some incoming from space (Wainwright, 2003, Wainwright et al., 2006). How then might the microorganisms, which originate on Earth, reach heights of 60 km above the Earth’s surface? One possibility is that they are ejected into the stratosphere by volcanoes. However, at least two of the above cited studies (Wainwright et al., 2003, Shivaji et al., 2006) were conducted some two years after the last major volcanic eruption on Earth; since bacteria and fungi deposit under gravity, any stratosphere isolations of organisms, derived from this study are unlikely to have originated from volcanoes. A number of other mechanisms have been suggested by which bacteria might be carried into the stratosphere, including blue lightening, gravitophotophoresis and electrostatic action (Wainwright et al., 2006). However, it appears unlikely that any of these mechanisms would be capable of carrying a particle of a diameter exceeding 1micron, i.e. the usual size of bacteria when grown on nutrient–rich laboratory media.

The only other mechanisms would be powerful monsoons and tropical storms. Dust, pollutants, water, gases, and other material are lofted into the stratosphere, presumably by monsoons, and therefore microbes would also likely be lifted to these heights (Joseph and Schild 2010). However, this has not been experimentally demonstrated.

Nevertheless, the likelihood of particles like bacteria being elevated from Earth to the stratosphere is likely to increase with decreasing cell size. Very small bacteria do occur in nature (Hahn, 2004) and it is likely that these so-called “ultrasmall” or “ultrabacteria” (i.e. filterable bacteria) would be more readily easily carried to the stratosphere than would bacteria of a larger size.

It is important to recognize the difference between ultrasmall bacteria and so-called nanobacteria (or nanobes). There is now considerable debate about the nature of nanobacteria, if they are alive, or are non-living calcified entities which attract proteins, nucleic acids and other biological materials to their surfaces. Nanobe–like particles have been found in the stratosphere (Wickramasinghe and Wickramasinghe, 2006), so it is possible that such calcified particles (of if they are alive) living entities may have brought, from space, to Earth. And if nanobes are arriving from space, and given their association with nuclei acids and proteins, then these biological materials may have also arrived from space and may have played a role in the origin or evolution of life on Earth.

6. THE PARADOX OF WHY THERE ARE NO ULTRASMALL BACTERIA IN THE STRATOSPHERE

Ultrasmall bacteria, which can pass through 0.1 and 0.2 micron filters, have been found in most oceans of the world and in a variety of soils. Most are relatively “exotic species” (Bacterides, Alphaproteobacteria, Betaproteobacteria, Actinobacteria and Spirochaetes Spirillum, Leucothrix, Flavobacterium, Cytophaga,Vibrio (Hahn, 2004). Since it assumed that the smaller a bacterium is, the more likely it is to be carried into the stratosphere, by known and suggested mechanisms, one would expect these bacteria to make up a predominant portion of the stratosphere microflora. Yet no reports of the isolation of such bacteria have yet been made in the numerous reports on microbial sampling of the stratosphere (Table 1).

It could be argued that these ultrasmall bacteria are not resistant to stratospheric UV and although they might reach this region from Earth they do not survive there. However, a number of non-UV resistant bacteria, such as Staphylococcus and Microococcus have been reported to be isolated from the stratosphere, so the lack of marked UV resistance should not necessarily prevent ultrasmall bacteria from surviving in the stratosphere.

Similarly, it might be argued that the isolation media employed in stratosphere-bacteria isolation studies select against ultrasmall bacteria. However, since these bacteria can be readily isolated from Earth environments employing the same, or similar, media used in stratosphere-bacteria isolation studies, this is not the case. To date, the majority of bacteria which have been isolated from the stratosphere are spore forming members of the genus Bacillus. How do these relatively large bacteria and their spores reach the stratosphere from Earth when ultrasmall bacteria cannot? Ultrasmall forms of Bacillus may, perhaps, be formed on Earth under starvation conditions and these might be small enough to be elevated to the stratosphere. Miteva and Brenchley (2004) have also reported the isolation, from 120,000 year old Greenland Glacier ice core, of several species of Bacillus isolates (described as being distinctly related to Bacillus mucilaginosus) possessing small filterable cells and spores).

However, if ultrasmall starvation forms of Bacillus can reach the stratosphere from Earth, then the more widely abundant ultrasmall bacteria should also do so, and would be represented in the reported stratosphere-bacteria isolation studies. The fact that ultrasmall bacteria have not been isolated at high frequency from the stratosphere remains an enigma. If bacteria, no matter how small they are, are not carried to the stratosphere from Earth, this would imply that the bacteria and fungi which have been isolated from these heights originate from space.

7. HOW DO FUNGI GET TO THE STRATOSPHERE?

The presence of fungi in the stratosphere presents an even greater enigma than does the presence of bacteria. This is because fungal hyphae and spores are generally much larger than bacteria, Fungal spores range from around 5 microns, for species of Penicillium, to 100 microns for species of Alternaria, both of which have been isolated from the stratosphere (Table 2).

Fungal mycelia commonly exceed 100 microns in length and are often above 10 micron in width. These dimensions relate to the size of fungi when grown on nutrient-rich media. Naturally growing fungi can be much smaller, and in nutrient-poor conditions, so-called microcycle conidiation produce small spores and fungal hyphae can be extremely fine (1-2 micron). As yet however, no ultrasmall fungi have been reported in the literature.

Clearly, under known mechanisms, it is extremely difficult to explain how these fungi reach the stratosphere. In the case of samples obtained at 41km, the sampling protocols used excluded the possibility of elevation by volcanic action. Therefore we are left with the reality that sections of fungal hyphae and, or spores can reach the stratosphere via unknown mechanisms. If these unknown mechanism include monsoons, then it is difficult to understand they selectively target large particles which are lifted to the stratosphere. This suggests that fungi may be incoming from the stratosphere. The idea that eukaryotes are incoming to Earth from space is probably even less acceptable to most microbiologists than is the idea that bacteria can make the same journey. The presence of fungi in the stratosphere therefore presents us with an even greater enigma than does the presence of bacteria.

8. CONCLUSIONS

The evidence presented above, clearly shows that a wide range of bacteria and fungi can be isolated from the stratosphere. This microflora is not dominated by one or two species, and both spore and non-spore forming bacteria have been isolated from these heights.

Are these organisms being transferred from Earth to heights above 41km, or are they incoming to Earth from space? If we accept that the tropopause effectively acts as a barrier to the upward movement of particles of the size of bacteria and fungi we need to explain how these organisms can reach the stratosphere from Earth, especially in the cases where volcanic transfer has been excluded. One way of avoiding this problem is to assume that ultrasmall forms of bacteria and fungi are carried up into the stratosphere by some mechanism, such as monsoons. If this is the case, then we would expect to find ultrasmall bacteria, known to exist in the Earth’s oceans to be the dominant bacteria isolated from the stratosphere, and this is not the case. The alternative possibility is that the bacteria and fungi present in the stratosphere are incoming from space to Earth; a hypothesis which would probably be dismissed by most microbiologists who instead might argue that there exists an unknown mechanism for transporting particles of the size of bacterial and fungal components up to the stratosphere.

We believe the best explanation for the mixed population of microorganisms which exists in the stratosphere, is that some are incoming to Earth from space (as represented by the observed particle masses in excess of 10 microns) and some are moving in the opposite direction. It is possible that the incoming bacteria may make up a considerable portion of the viable, but non-culturable, bacteria found on Earth. Although the findings presented above do not prove that bacteria and fungi are incoming to Earth from space, the evidence seems to favor this proposition and the reality of neopansermia.