Author Archives: Albert Bordons
12th August 2017
Probiotics are living microorganisms that, when ingested in adequate amounts, can have a positive effect on the health of guests (FAO / WHO 2006; World Gastroenterology Organization 2011, Fontana et al., 2013). Guests can be humans but also other animals. Lactic acid bacteria, especially the genus Lactobacillus and Bifidobacterium, both considered as GRAS (Generally recognized as safe), are the microbes most commonly used as probiotics, but other bacteria and some yeasts can also be useful. Apart from being able to be administered as medications, probiotics are commonly consumed for millennia as part of fermented foods, such as yoghurt and other dairy products (see my article “European cheese from 7400 years ago..” “December 26th, 2012). As medications, probiotics are generally sold without prescription, over-the-counter (OTC) in pharmacies.
I have already commented on the other posts of this blog the relevance of probiotics (“A new probiotic modulates microbiota against hepatocellular carcinoma” August 24th, 2016), as well as the microbiota that coexists with our body (“Bacteria in the gut controlling what we eat” October 12th, 2014; “The good bacteria of breast milk” February 3rd, 2013) and other animals (“Human skin microbiota … and our dog” December 25th, 2015; “The herbivore giant panda …. and its carnivore microbiota” September 30th, 2015).
Besides lactic acid bacteria and bifidobacteria, other microorganisms that are also used to a certain extent as probiotics are the yeast Saccharomyces cerevisiae, some strains of Escherichia coli, and some Bacillus, as we will see. Some clostridia are also used, related to what I commented in a previous post of this blog by March 21st, 2015 (“We have good clostridia in the gut ...”).
In fact, Bacillus and clostridia have in common the ability to form endospores. And both groups are gram-positive bacteria, within the taxonomic phylum Firmicutes (Figure 1), which also includes lactic acid bacteria. However, bacilli (Bacillus and similar ones, but also Staphylococcus and Listeria) are more evolutionarily closer to lactobacillalles (lactic acid bacteria) than to clostridia ones. The main physiological difference between Clostridium and Bacillus is that the first are strict anaerobes while Bacillus are aerobic or facultative anaerobic.
Figure 1. Phylogenetic tree diagram of Gram-positive bacteria (Firmicutes and Actinobacteria). Own elaboration.
Bacterial endospores (Figure 2) are the most resistant biological structures, as they survive extreme harsh environments, such as UV and gamma radiation, dryness, lysozyme, high temperatures (they are the reference for thermal sterilization calculations), lack of nutrients and chemical disinfectants. They are found in the soil and in the water, where they can survive for very long periods of time.
Figure 2. Endospores (white parts) of Bacillus subtilis in formation (Image of Simon Cutting).
Bacillus in fermented foods, especially Asian
Several Bacillus are classically involved in food fermentation processes, especially due to their protease production capacity. During fermentation, this contributes to nutritional enrichment with amino acids resulting from enzymatic proteolysis.
Some of these foods are fermented rice flour noodles, typical of Thailand and Burma (nowadays officially Myanmar). It has been seen that a variety of microorganisms (lactic acid bacteria, yeasts and other fungi) are involved in this fermentation, but also aerobic bacteria such as B. subtilis. It has been found that their proteolytic activity digests and eliminates protein rice substrates that are allergenic, such as azocasein, and therefore they have a beneficial activity for the health of consumers (Phromraksa et al. 2009).
However, the best-known fermented foods with Bacillus are the alkaline fermented soybeans. As you know, soy (Glycine max) or soya beans are one of the most historically consumed nourishing vegetables, especially in Asian countries. From they are obtained “soy milk”, soybean meal, soybean oil, soybean concentrate, soy yogurt, tofu (soaked milk), and fermented products such as soy sauce, tempeh, miso and other ones. Most of them are made with the mushroom Rhizopus, whose growth is favoured by acidification or by direct inoculation of this fungus. On the other hand, if soy beans are left to ferment only with water, the predominant natural microbes fermenting soy are Bacillus, and in this way, among other things, the Korean “chongkukjang” is obtained, “Kinema” in India, the “thua nao” in northern Taiwan, the Chinese “douchi”, the “chine pepoke” from Burma, and the best known, the Japanese “natto” (Figure 3). Spontaneous fermentation with Bacillus gives ammonium as a by-product, and therefore is alkaline, which gives a smell not very good to many of these products. Nevertheless, natto is made with a selected strain of B. subtilis that gives a smoother and more pleasant smell (Chukeatirote 2015).
These foods are good from the nutritional point of view as they contain proteins, fibre, vitamins, and they are of vegetable or microbial origin. In addition, the advertising of the commercial natto emphasizes, besides being handmade and sold fresh (not frozen), its probiotic qualities, saying that B. subtilis (Figure 4) promotes health in gastrointestinal, immunologic, cardiovascular and osseous systems (www.nyrture.com). They say the taste and texture of natto are exquisite. It is eaten with rice or other ingredients and sauces, and also in the maki sushi. We must try it !
Figure 3. “Natto”, soybeans fermented with B. subtilis, in a typical Japanese breakfast with rice (Pinterest.com).
Figure 4. Coloured electronic micrograph of Bacillus subtilis (Nyrture.com).
Bacillus as probiotics
The endospores are the main advantage of Bacillus being used as probiotics, thanks to their thermal stability and to survive in the gastric conditions (Cutting 2011). Although Clostridium has also this advantage, its strict anaerobic condition makes its manipulation more complex, and moreover, for the “bad reputation” of this genus due to some well-known toxic species.
Unlike other probiotics such as Lactobacillus or Bifidobacterium, Bacillus endospores can be stored indefinitely without water. The commercial products are administered in doses of 10^9 spores per gram or per ml.
There are more and more commercial products of probiotics containing Bacillus, both for human consumption (Table 1) and for veterinary use (Table 2). In addition, there are also five specific products for aquaculture with several Bacillus, and also shrimp farms are often using products of human consumption (Cutting 2011).
For use in aquaculture, probiotic products of mixtures of Bacillus (B. thuringiensis, B. megaterium, B. polymixa, B. licheniformis and B. subtilis) have been obtained by isolating them from the bowel of the prawn Penaeus monodon infected with vibriosis. They have been selected based on nutrient biodegradation and the inhibitory capacity against the pathogen Vibrio harveyi (Vaseeharan & Ramasamy 2003). They are prepared freeze-dried or microencapsulated in sodium alginate, and it has been shown to significantly improve the growth and survival of shrimp (Nimrat et al., 2012).
As we see for human consumption products, almost half of the brands (10 of 25) are made in Vietnam. The use of probiotic Bacillus in this country is more developed than in any other, but the reasons are not clear. Curiously, as in other countries in Southeast Asia, there is no concept of dietary supplements and probiotics such as Bacillus are only sold as medications approved by the Ministry of Health. They are prescribed for rotavirus infection (childhood diarrhoea) or immune stimulation against poisoning, or are very commonly used as a therapy against enteric infections. However, it is not clear that clinical trials have been carried out, and they are easy-to-buy products (Cutting 2011).
Table 1. Commercial products of probiotics with Bacillus, for human consumption (modified from Cutting 2011).
|Product||Country where it is made||Species of Bacillus|
|Bactisubtil ®||France||B. cereus|
|Bibactyl ®||Vietnam||B. subtilis|
|Bidisubtilis ®||Vietnam||B. cereus|
|Bio-Acimin ®||Vietnam||B. cereus and 2 other|
|Biobaby ®||Vietnam||B. subtilis and 2 other|
|Bio-Kult ®||United Kingdom||B. subtilis and 13 other|
|Biosporin ®||Ukraine||B. subtilis + B. licheniformis|
|Biosubtyl ®||Vietnam||B. cereus|
|Biosubtyl DL ®||Vietnam||B. subtilis and 1 other|
|Biosubtyl I and II ®||Vietnam||B. pumilus|
|Biovicerin ®||Brazil||B. cereus|
|Bispan ®||South Korea||B. polyfermenticus|
|Domuvar ®||Italy||B. clausii|
|Enterogermina ®||Italy||B. clausii|
|Flora-Balance ®||United States||B. laterosporus *|
|Ildong Biovita ®||Vietnam||B. subtilis and 2 other|
|Lactipan Plus ®||Italy||B. subtilis *|
|Lactospore ®||United States||B. coagulans *|
|Medilac-Vita ®||China||B. subtilis|
|Nature’s First Food ®||United States||42 strains, including 4 B.|
|Neolactoflorene ®||Italy||B. coagulans * and 2 other|
|Pastylbio ®||Vietnam||B. subtilis|
|Primal Defense ®||United States||B. subtilis|
|Subtyl ®||Vietnam||B. cereus|
|Sustenex ®||United States||B. coagulans|
* Some labelled as Lactobacillus or other bacteria are really Bacillus
Table 2. Commercial products of probiotics with Bacillus, for veterinary use (modified from Cutting 2011).
|Product||Animal||Country where it is made||Species of Bacillus|
|AlCare ®||Swine||Australia||B. licheniformis|
|BioGrow ®||Poultry, calves and swine||United Kingdom||B. licheniformis and B. subtilis|
|BioPlus 2B ®||Piglets, chickens, turkeys||Denmark||B. licheniformis and B. subtilis|
|Esporafeed Plus ®||Swine||Spain||B. cereus|
|Lactopure ®||Poultry, calves and swine||India||B. coagulans *|
|Neoferm BS 10 ®||Poultry, calves and swine||France||B. clausii|
|Toyocerin ®||Poultry, calves, rabbits and swine||Japan||B. cereus|
The Bacillus species that we see in these Tables are those that really are found, once the identification is made, since many of these products are poorly labelled as Bacillus subtilis or even as Lactobacillus (Green et al. 1999; Hoa et al. 2000). These labelling errors can be troubling for the consumer, and especially for security issues, since some of the strains found are Bacillus cereus, which has been shown to be related with gastrointestinal infections, since some of them produce enterotoxins (Granum & Lund 1997; Hong et al. 2005)
The probiotic Bacillus have been isolated from various origins. For example, some B. subtilis have been isolated from the aforementioned Korean chongkukjang, which have good characteristics of resistance to the gastrointestinal tract (GI) conditions and they have antimicrobial activity against Listeria, Staphylococcus, Escherichia and even against B. cereus (Lee et al. 2017).
One of the more known probiotics pharmaceuticals is Enterogermina ® (Figure 5), with B. subtilis spores, which is recommended for the treatment of intestinal disorders associated with microbial alterations (Mazza 1994).
Figure 5. Enterogermina ® with spores of Bacillus subtilis (Cutting 2011)
Bacillus in the gastrointestinal tract: can they survive there ?
It has been discussed whether administered spores can germinate in the GI tract. Working with mice, Casula & Cutting (2002) have used modified B. subtilis, with a chimeric gene ftsH-lacZ, which is expressed only in vegetative cells, which can be detected by RT-PCR up to only 100 bacteria. In this way they have seen that the spores germinate in significant numbers in the jejunum and in the ileum. That is, spores could colonize the small intestine, albeit temporarily.
Similarly, Duc et al. (2004) have concluded that B. subtilis spores can germinate in the gut because after the oral treatment of mice, in the faeces are excreted more spores that the swallowed ones, a sign that they have been able to proliferate. They have also detected, through RT-PCR, mRNA of vegetative bacilli after spore administration, and in addition, it has been observed that the mouse generates an IgG response against bacterial vegetative cells. That is, spores would not be only temporary stagers, but they would germinate into vegetative cells, which would have an active interaction with the host cells or the microbiota, increasing the probiotic effect.
With all this, perhaps it would be necessary to consider many Bacillus as not allochthonous of the GI tract, but as bacteria with a bimodal growth and sporulation life cycle, both in the environment and in the GI tract of many animals (Hong et al. 2005).
Regarding the normal presence of Bacillus in the intestine, when the different microorganisms inhabiting the human GI tract are studied for metagenomic DNA analysis of the microbiota, the genus Bacillus does not appear (Xiao et al., 2015). As we can see (Figure 6), the most common are Bacteroides and Clostridium, followed by various enterobacteria and others, including bifidobacteria.
Figure 6. The 20 bacterial genera more abundant in the mice (left) and human (right) GI tract (Xiao et al. 2015).
In spite of this, several species of Bacillus have been isolated from the GI tract of chickens, treating faecal samples with heat and ethanol to select only the spores, followed by aerobic incubation (Barbosa et al. 2005). More specifically, the presence of B. subtilis in the human microbiota has been confirmed by selective isolation from biopsies of ileum and also from faecal samples (Hong et al. 2009). These strains of B. subtilis exhibited great diversity and had the ability to form biofilms, to sporulate in anaerobiosis and to secrete antimicrobials, thereby confirming the adaptation of these bacteria to the intestine. In this way, these bacteria can be considered intestinal commensals, and not only soil bacteria.
Security of Bacillus as probiotics
The oral consumption of important amounts of viable microorganisms that are not very usual in the GI treatment raises additional doubts about safety. Even more in the use of species that do not have a history of safe use in foods, as is the case of sporulated bacteria. Even normal bowel residents may sometimes act as opportunistic pathogens (Sanders et al. 2003).
With the exception of B. anthracis and B. cereus, the various species of Bacillus are generally not considered pathogenic. Of course, Bacillus spores are commonly consumed inadvertently with foods and in some fermented ones. Although Bacillus are recognized as GRAS for the production of enzymes, so far the FDA has not guaranteed the status of GRAS for any sporulated bacteria with application as a probiotic, neither Bacillus nor Clostridium. While Lactobacillus and Bifidobacterium have been the subject of numerous and rigorous tests of chronic and acute non-toxicity, and a lot of experts have reviewed data and have concluded that they are safe as probiotics, there is no toxicity data published on Bacillus in relation to their use as probiotics. When reviewing articles on Medline with the term “probiotic” and limited to clinical studies, 123 references appear, but Bacillus does not appear in any of them (Sanders et al. 2003).
Instead, there are some clinical studies where Bacillus strains have been detected as toxigenic. All this explains that some probiotic Bacillus producers refer to them with the misleading name of Lactobacillus sporogenes, a non-existent species, as can be seen from NCBI (https://www.ncbi.nlm.nih.gov/taxonomy/?term = Lactobacillus + sporogenes).
Finally, we should remember the joint report on probiotics of FAO (United Nations Food and Agriculture Organization) and WHO (World Health Organization) (FAO / WHO 2006), which suggests a set of Guidelines for a product to be used as a probiotic, alone or in the form of a new food supplement. These recommendations are:
- The microorganism should be well characterized at the species level, using phenotypic and genotypic methods (e.g. 16S rRNA).
- The strain in question should be deposited in an internationally recognized culture collection.
- To evaluate the strain in vitro to determine the absence of virulence factors: it should not be cytotoxic neither invades epithelial cells, and not produce enterotoxins or haemolysins or lecithinases.
- Determination of its antimicrobial activity, and the resistance profile, including the absence of resistance genes and the inability to transfer resistance factors.
- Preclinical evaluation of its safety in animal models.
- Confirmation in animals demonstrating its effectiveness.
- Human evaluation (Phase I) of its safety.
- Human evaluation (Phase II) of its effectiveness (if it does the expected effect) and efficiency (with minimal resources and minimum time).
- Correct labelling of the product, including genus and species, precise dosage and conservation conditions.
The use of Bacillus as probiotics, especially in the form of dietary supplements, is increasing very rapidly. More and more scientific studies show their benefits, such as immune stimulation, antimicrobial activities and exclusive competition. Their main advantage is that they can be produced easily and that the final product, the spores, is very stable, which can easily be incorporated into daily food. In addition, there are studies that suggest that these bacteria may multiply in GI treatment and may be considered as temporary stagers (Cutting 2011).
On the other hand, it is necessary to ask for greater rigor in the selection and control of the Bacillus used, since some, if not well identified, could be cause of intestinal disorders. In any case, since the number of products sold as probiotics that contain the sporulated Bacillus is increasing a lot, one must not assume that all are safe and they must be evaluated on a case-by-case basis (Hong et al. 2005).
Barbosa TM, Serra CR, La Ragione RM, Woodward MJ, Henriques AO (2005) Screening for Bacillus isolates in the broiler gastrointestinal tract. Appl Environ Microbiol 71, 968-978.
Casula G, Cutting SM (2002) Bacillus probiotics: Spore germination in the gastrointestinal tract. Appl Environ Microbiol 68, 2344-2352.
Chukeatirote E (2015) Thua nao: Thai fermented soybean. J Ethnic Foods 2, 115-118.
Cutting SM (2011) Bacillus probiotics. Food Microbiol 28, 214-220.
Duc LH, Hong HA, Barbosa TM, Henriques AO, Cutting SM (2004) Characterization of Bacillus probiotics available for human use. Appl Environ Microbiol 70, 2161-2171.
FAO/WHO (2006) Probiotics in food. Health and nutritional properties and guidelines for evaluation. Fao Food and Nutrition Paper 85. Reports of Joint FAO/WHO expert consultations.
Fontana L, Bermudez-Brito M, Plaza-Diaz J, Muñoz-Quezada S, Gil A (2013) Sources, isolation, characterization and evaluation of probiotics. Brit J Nutrition 109, S35-S50.
Granum, P. E., T. Lund (1997) Bacillus cereus and its food poisoning toxins. FEMS Microbiol. Lett. 157:223–228.
Green, D. H., P. R. Wakeley, A. Page, A. Barnes, L. Baccigalupi, E. Ricca, S. M. Cutting (1999) Characterization of two Bacillus probiotics. Appl Environ Microbiol 65, 4288–4291.
Hoa, N. T., L. Baccigalupi, A. Huxham, A. Smertenko, P. H. Van, S. Ammendola, E. Ricca, A. S. Cutting (2000) Characterization of Bacillus species used for oral bacteriotherapy and bacterioprophylaxis of gastrointestinal disorders. Appl Environ Microbiol 66, 5241–5247.
Hong HA, Dic LH, Cutting SM (2005) The use of bacterial spore formers as probiotics. FEMS Microbiol Rev 29, 813-835.
Hong HA, Khaneja R, Tam NMK, Cazzato A, Tan S, Urdaci M, Brisson A, Gasbarrini A, Barnes I, Cutting SM (2009) Bacillus subtilis isolated from the human gastrointestinal tract. Res Microbiol 160, 134-143.
Lee S, Lee J, Jin YI, Jeong JC, Hyuk YH, Lee Y, Jeong Y, Kim M (2017) Probiotic characteristics of Bacillus strains isolated from Korean traditional soy sauce. LWT – Food Sci Technol 79, 518-524.
Mazza P (1994) The use of Bacillus subtilis as an antidiarrhoeal microorganism. Boll Chim. Farm. 133, 3-18.
Nimrat S, Suksawat S, Boonthai T, Vuthiphandchai V (2012) Potential Bacillus probiotics enhance bacterial numbers, water quality and growth during early development of white shrimp (Litopenaeus vannamei). Veterinary Microbiol 159, 443-450.
Phromraksa P, Nagano H, Kanamaru Y, Izumi H, Yamada C, Khamboonruang C (2009) Characterization of Bacillus subtilis isolated from Asian fermented foods. Food Sci Technol Res 15, 659-666.
Sanders ME, Morelli L, Tompkins TA (2003) Sporeformers as human probiotics: Bacillus, Sporolactobacillus, and Brevibacillus. Compr Rev Food Sci Food Safety 2, 101-110
Vaseeharan, B., P. Ramasamy (2003) Control of pathogenic Vibrio spp. by Bacillus subtilis BT23, a possible probiotic treatment for black tiger shrimp Penaeus monodon. Lett Appl Microbiol 36, 83–87
World Gastroenterology Organisation Global Guidelines (2011) Probiotics and Prebiotics.
Xiao et al. (2015) A catalogue of the mouse gut metagenome. Nature Biotechnol 33, 1103-1108.
March 20th, 2016
The Arctic Ocean
Interestingly and coincidentally, “Arctic” comes from the Greek word αρκτος -arctos-, which means “bear” and is a reference to the constellation Ursa Minor, where is the North Star, which indicates the geographic North Pole .
The Arctic constitutes a unique ecosystem of the Earth, consisting of a large ice field, or ice-covered ocean, sometimes regarded as the northern part of the Atlantic Ocean, and it is surrounded by land, which is permafrost, with complete absence of trees. Life in the Arctic consists of organisms adapted to ice, including zooplankton and phytoplankton, fish, marine mammals, birds, land animals, plants and human societies fully adapted to the extreme conditions of the environment.
Due to global warming, isotherms are moving northward at a rate exceeding 50 km per decade over the past 30 years, so if we define the Arctic from a defined temperature or the tree line, its size is diminishing, being the reduction of sea ice the most visible effect.
Anthropogenic climate change: global warming, especially in the Arctic
Yes: climate change is here and it is generated by human activities, that is, it is anthropogenic. Previously there have been on Earth fluctuations in global temperature caused by natural phenomena, usually long-term and cyclical variations. For example, glaciations since about 2 million years are repeated every 100,000 years, and last ice age ended 15,000 years ago. So we are living now in an interglacial period and the next ice age could become not before 50,000 years. The cause of this cycle of glaciations seems to be orbital variations of the Earth, resulting in a lower insolation in high latitudes of the northern hemisphere during glacial periods.
Solar activity, like other stars, has cycles and roughly every 600 years there are periods of little activity (absence of very few solar spots and auroras), with lower energy output, which corresponds to cold periods in the Earth’s climate. The last minimal was in the period 1645-1715, and therefore from the middle of the eighteenth century we enjoy a maximum solar activity, with small cycles of minimum and maximum every 11 years.
Discounting these natural variations, it is clear that throughout the 20th century and especially since the 1960s there has been a steady increase in global average temperature (Figure 1), reaching almost 1ºC more than the beginning of the 20th century. In the early years of the current century the trend is worsening. The last 10 years have been the warmest since there are records, and the forecast is to continue increasing. Most experts agree that humans exert a direct impact on the heating process known as the greenhouse effect. The causes of this effect are some of gaseous components of the atmosphere, especially CO2, which has grown in parallel with rising temperatures, from about 300 ppm at the beginning of 20th century to nearly 400 ppm today. This CO2 and other gases as water vapour, methane and other exclusively anthropogenic absorb radiation and the result is that the atmosphere warms further.
Figure 1. Increase in average global temperature compared to the beginning of 20th century (from GISTEMP).
This global warming is particularly evident in the Arctic. The temperature increases are higher in northern latitudes, especially 60-70º N, where this past December 2015 (Figure 2) have raised to 9ºC above average in large areas of North America and Eurasia. This is called Polar Warming Amplification (PWA). The cause of this overheating in the Arctic respect of the rest of Earth is partly due to the loss of snow and ice (retroactive effect) because the largest area of land and water absorbs more solar energy than white ice (albedo effect), but also the PWA is partly due to the dynamic atmospheric transport, which transports heat energy from the clouds and subtropical regions to the north (Taylor et al 2013).
Figure 2. Thermal anomaly registered in December 2015 with respect to the average 1951-1980 (from GISTEMP).
Besides the consequences of this warming on the Arctic ice that we will comment below, another serious problem is the melting of permafrost, since then methane gas trapped under the frozen ground is released. This way, vast quantities of methane are released, and this greenhouse gas is contributing further to accelerate the global warming.
Less and less ice in the Arctic
Linear trends of sea ice extent and sea ice in the Arctic from 1979 to date are negative year after year, for any month is considered, but it is more clear by comparing Septembers, at the end of the summer when the ice is melting (Figure 3). Of the approximately 7 million km2 minimum in September (the maximum in March is about 16 million), about 100,000 km2 are melt per year, almost 9% every 10 years (Serreze et al 2007), so that there is now almost half ice than in 1979 (Figure 4).
Figure 3. Comparison of the extent of sea ice (in red): September 1979 and 2012 (from The Cryosphere Today).
Figure 4. Average monthly extension of Arctic sea ice since 1979 (Reeves et al 2013).
In addition to the reduction in surface ice, keep in mind the reduction in volume, representing now a third of what it was in September 1979.
There is a big difference between the different models for predicting the disappearance of Arctic sea ice. Half of them expect the total disappearance by September 2100. Predictions move since September 2040 the less optimistic until well past 2100 for the other (Serreze et al 2007).
Other problems resulting from the disappearance of sea ice are the ship traffic, which could shorten distances trips between the ports of northern countries, and on the other hand the exploitation of oilfields and other fossil fuels and minerals, since there is a large part of global reserves in the Arctic (Figure 5).
Figure 5. Left: forecast paths for open sea ships (blue) and for icebreakers (red) for 2040-2059. Right: Distribution of the potential major reserves of oil and gas (yellow) and licenses (red) and wells in operation or to operate (black). The dashed line indicates the limit of Conservation of Arctic Flora and Fauna (CAFF) declared by the Working Group of the Arctic Council (www.arctic-council.org). Figures from Reeves et al (2013).
Ecological consequences of the disappearance of the Arctic ice pack
There are many living beings linked to the ice. The polar bears roam on the Arctic ice, so we are feared for his fate. Many fish, seals and crustaceans (krill) form a food chain that starts from the algae that grow under the ice in a very consistent environment, rich in nutrients, especially favourable for marine life (Figure 6 A). Moreover, floating sea ice in summer is a good corridor for dispersion of terrestrial vertebrates (for instance arctic foxes) and plants.
The gradual disappearance of sea ice and warming in the Arctic coast involves a series of ecological imbalances (Figure 6 B). We see for example how walruses forced to remain grouped on the ground are more predisposed to disease transmission. The loss of sea ice diminishes dispersion by ice corridors and then the land populations are most isolated, thus gene flow is restricted. Polar bears and other predators that hunt on the sea ice have it much harder and their populations are at risk. Phytoplankton productivity decreases significantly, thereby reducing zooplankton, and then the whole food chain (fish, seals, etc.) is affected (Post et al 2013).
Figure 6. Ecological interactions influenced by sea ice. A: The distribution and seasonality of sea ice affects the abundance, distribution and interactions of the entire ecosystem in balance. B: The longest period without ice and less sea ice extent have disastrous consequences on the balance of the ecosystem (Post et al 2013).
The polar bear tries to survive
The polar bear (Ursus maritimus) is considered an endangered animal. There are only about 25,000 worldwide. The impact of climate change affects the exclusive habitat of polar regions and forecasts suggest that in a few years from now the ice of the Arctic will melt permanently and polar bears may become extinct because of warming area.
The polar bear is basically carnivorous, unlike others such as brown bears, and remains above the ice hunting seals. With the gradual disappearance of the ice it has more trouble finding preys, and some have begun to learn how to catch salmon rivers, as we see in the images (Figure 7).
Figure 7. White Bear dedicated to fishing salmons in order to survive (www.youtube.com/watch?v=9m_Q9Ojbcmw).
We have also seen groups of polar bears at sea fishing (see video) and dive emerging alternately as if they were dolphins or porpoises. Despite these small adaptations, the food is very low and it is clear that their populations are declining rapidly.
Orcas thrive north
The disappearance of the northern ice is a dramatic ecological change that is causing the disappearance of some species like the polar bear, but interestingly these imbalances benefit some other emerging species. This is the case of the killer whale (Orcinus orca), which is thriving more and more to the north (Figure 8).
Figure 8. Places (marked with numbers) of the Canadian Arctic where groups of orcas were repeatedly photographed between 2004 and 2009 (Young et al 2011).
Eskimo Inuit people live around the American Arctic (from Quebec to Alaska including Hudson Bay and adjacent islands) and the west coast of Greenland, and they are the first witnesses since the mid-twentieth century observing whales in their waters, unknown before. Moreover, in recent years scientists have made numerous orca’ sightings, they have been photographed individually (Young et al 2011), and their travels have been followed through bioacoustics (Ferguson et al 2010) and other techniques.
Figure 9. (Top): Narwhals with the characteristic great tusk, which gave rise to the myth of the unicorn. (Low): Group of orcas attacking narwhals cornered on the beach. Watch the video of PBS Nature.
For some years attacks by orcas on narwhals (as in Figure 9) have been observed repeatedly by Inuit Eskimos and studied in detail by several scientists. Laidre et al (2006) observed that before approaching whales, the narwhals tend to group, are more quiet and swim closer to the beach in shallow waters. During the attack, the narwhals disperse significantly but nevertheless mortality is very high. After predation, which can last several hours, oily stains are observed in sea surface, which come from fat of depredated narwhals (Figure 10).
Figure 10. Group of orcas surrounded by patches of oil on the sea surface from the fat of attacked narwhals (Laidre et al 2006).
Orcas’ attacks on narwhals are so common and effective that are beginning to affect the population. The effects are even worse in other cetaceans with smaller population such as whales of Greenland or bowhead (Balena mysticetus), which are now virtually extinct (Figure 11).
Figure 11. Scheme of preys’ proportions by a group of orcas from Hudson Bay (Ferguson et al 2010).
In conclusion, anthropogenic climate change is affecting the Arctic ecosystem severely (and all the other ecosystems), and although this problem is becoming known, effective policy measures to reduce emissions of CO2 and other greenhouse gases are so scarce that hardly will arrive in time. We are leading the planet Earth to a massive extinction of species and ecological changes ever seen in the history of humans.
The picture says it all: polar bear habitat is running out.
Arctic Council: http://www.arctic-council.org
Ferguson S.H., Higdon J.W. & Chmelnitsky E.G. (2010) The rise of killer whales as a major Arctic predator. In S.H. Ferguson, et al. (eds.): A little less Arctic: top predators in the world’s largest northern inland sea, Hudson Bay. Pp. 117–136. New York: Springer
GISTEMP, Goddard Institute for Space Studies Surface Temperature Analysis (NASA-GISS): http://data.giss.nasa.gov/gistemp/
Hawkings E (2014) nov 28: http://www.climate-lab-book.ac.uk/2014/hiatuses-in-the-rise-of-temperature/
Laidre KL, Heide-Jørgensen MP, Orr J (2006) Reactions of narwhals, Monodon monoceros, to killer whale, Orcinus orca, attacks in the Eastern Canadian Arctic. Can. Field Nat., 120, 457–465
Morell V (2012) Killer whale menu finally revealed. http://www.sciencemag.org/news/2012/01/killer-whale-menu-finally-revealed
Post et al. (2013) Ecological Consequences of Sea-Ice Decline. Science. DOI: 10.1126/science.1235225: http://www.carbonbrief.org/knock-on-effects-for-wildlife-as-the-arctic-loses-ice
Reeves RR et al (2014) Distribution of endemic cetaceans in relation to hydrocarbon development and commercial shipping in a warming arctic. Marine Policy 44, 375-389
Serreze MC, Holland MM, Stroeve J (2007) Perspectives on the Arctic’s shrinking sea-ice cover. Science 315, 5818, 1533–6.
Taylor PC, Cai M, Hu A, Meehl J, Washington W, Zhang GJ (2013) Decomposition of feedback contributions to Polar Warming Amplification. J Climate 26, 7023-43
The Cryosphere Today: http://arctic.atmos.uiuc.edu/cryosphere/
Young BG, Jeff W. Higdon JW, Steven H. Ferguson SH (2011) Killer whale (Orcinus orca) photo-identification in the eastern Canadian Arctic. Polar Research Vol 30
December 25th, 2015
Diversity of the human microbiota in different parts of the body and between individuals
As I have commented in previous posts of this blog (Good Clostridia in our gut March 21st, 2015; Bacteria controlling what we eat October 12th, 2014; Bacteria of breast milk February 3rd, 2013), it becomes increasingly clear the importance of our microbiota, id est, all the micro-organisms, especially bacteria, with which we live.
The human microbiota varies from one individual to another, in relation to diet, age and the own genetic and phenotypic characteristics. Moreover, since we do not live isolated, there is also the influence of the environment, and of other people with we live, including our pets, dogs and others. They all have also their own microbiota.
The human body is home to many different microorganisms: bacteria (and archaea), fungi and viruses, that live on the skin, in the gut and in several other places in the body (Figure 1). While many of these microbes are beneficial to their human host, we know little about most of them. Early research focused on the comparison of the microorganisms found in healthy individuals with those found in people suffering from a particular disease. More recently, researchers have been interested in the more general issues, such as understanding how the microbiota is established and knowing the causes of the similarities and differences between the microbiota of different individuals.
Figure 1. Types of microorganisms that live in different parts of the human body: bacteria (large circles), fungi (small circles right) and viruses (small circles left) (Marsland & Gollwitzer 2014)
Now we know that communities of microorganisms that are found in the gut of genetically related people tend to be more similar than those of people who are not related. Moreover, microbial communities found in the gut of unrelated adults living in the same household are more similar than those of unrelated adults living in different households (Yatsunenko et al 2012). However, these studies have focused on the intestine, and little is known about the effect of the relationship, cohabitation and age in microbiota of other parts of the body, such as skin.
Human skin microbiota
The skin is an ecosystem of about 1.8 m2 of various habitats, with folds, invaginations and specialized niches that hold many types of microorganisms. The main function of the skin is to act as a physical barrier, protecting the body from potential attacks by foreign organisms or toxic substances. Being also the interface with the external environment, skin is colonized by microorganisms, including bacteria, fungi, viruses and mites (Figure 2). On its surface there are proteobacteria, propionibacteria, staphylococci and some fungi such as Malassezia (an unicellular basidiomycetous). Mites such as Demodex folliculorum live around the hair follicles. Many of these microorganisms are harmless and often they provide vital functions that the human genome has not acquired by evolution. The symbiotic microorganisms protect human from other pathogenic or harmful microbes. (Grice & Segre 2011).
Figure 2. Schematic cross section of human skin with the different microorganisms (Grice & Segre 2011).
According to the commented diversity of microbiota, this is also very different depending on the region of skin (Figure 3), and therefore depending on the different microenvironments, that can be of three different characteristics: sebaceous or oily, wet and dry.
Figure 3. Topographic distribution of bacterial types in different parts of the skin (Grice & Segre 2011)
The skin is a complex network (structural, hormonal, nervous, immune and microbial) and in this sense it has been proven that the resident microbiota collaborates with the immune system, especially in the repair of wounds (Figure 4). As we see, particularly the lipopotheicoic acid (LTA), compound of the bacterial cell wall, can be released by Staphylococcus epidermidis and stimulates Toll-like receptors TLR2, which induce the production of antimicrobial peptides, and also stimulate epidermal keratinocytes via TLR3, which trigger the inflammation with production of interleukin and attracting leukocytes (Heath & Carbone 2013). All this to ensure the homeostatic protection and the defence against the potential pathogens. More information in the review of Belkaid & Segre (2014).
Figure 4. Contribution of the resident microbiota to the immunity and wound repair (Heath & Carbone 2013)
At home we share microbiota, and with the dog
As mentioned earlier, environment influences the microbiota of an individual, and therefore, individuals who live together tend to share some of the microbiota. Indeed, it was recently studied by Song et al (2013), with 159 people and 36 dogs from 60 families (couples with children and / or dogs). They study the microbiota of gut, tongue and skin. DNA was extracted from a total of 1076 samples, amplifying the V2 region of the 16S rRNA gene with specific primers, and then it was proceeded to multiplex sequencing of high performance (High-Throughput Sequencing) with an Illumina GA IIx equipment. Some 58 million sequences were obtained, with an average of 54,000 per sample, and they were analysed comparing with databases to find out what kind of bacteria and in what proportions.
The results were that the microbial communities were more similar to each other in individuals who live together, especially for the skin, rather than the bowel or the tongue. This was true for all comparisons, including pairs of human and dog-human pairs. As shown in Figure 5, the number of bacterial types shared between different parts was greater (front, palms and finger pulps dog) of the skin of humans and their own dog (blue bars) than the human with dogs of other families (red bars), or dogs with people without dogs (green bars). We also see that the number of shared bacterial types is much lower when compared faecal samples or the tongue (Song et al 2013).
Figure 5. Numbers of bacterial phylotypes (phylogenetic types) shared between adults and their dogs (blue), adults with other dogs (red) and adults who do not have dogs with dogs. There are compared (dog-human) fronts, hands, legs pulps, and also faecal samples (stool) and tongues. Significance of being different: *p<0.05, **p<0.001 (Song et al 2013)
This suggests that humans probably take a lot of microorganisms on the skin by direct contact with the environment and that humans tend to share more microbes with individuals who are in frequent contact, including their pets. Song et al. (2013) also found that, unlike what happens in the gut, microbial communities in the skin and tongue of infants and children were relatively similar to those of adults. Overall, these findings suggest that microbial communities found in the intestine change with age in a way that differs significantly from those found in the skin and tongue.
Although is not the main reason for this post, briefly I can say that the overall intestinal microbiota of dogs is not very different from humans in numbers (1011 per gram) and diversity, although with a higher proportion of Gram-positive (approx. 60% clostridial, 12% lactobacilli, 3% bifidobacteria and 3% corynebacteria) in dogs, and less Gram-negative (2% Bacteroides, 2% proteobacteria) (García-Mazcorro Minamoto & 2013).
Less asthma in children living with dogs
Although the relationship with the microbiota has not fully been demonstrated, some evidence of the benefits of having a dog has been shown recently, and for the physical aspects, not just for the psychological ones. Swedish researchers (Fall et al 2015) have carried out a study of all new-borns (1 million) in Sweden since 2001 until 2010, counting those suffering asthma at age 6. As the Swedes also have registered all dogs since 2001, these researchers were able to link the presence of dogs at home during the first year of the baby with the onset of asthma or no in children, and have come to the conclusion that children have a lower risk of asthma (50% less) if they have grown in the presence of a dog.
Similar results were obtained for children raised on farms or in rural environments, and thus having contact with other animals. All this would agree with the “hygiene hypothesis”, according to which the lower incidence of infections in Western countries, especially in urban people, would be the cause for increased allergic and autoimmune diseases (Okada et al 2010). In line with the hypothesis, it is believed that the human immune system benefits from living with dogs or other animals due to the sharing of the microbiota. However, in these Swede children living with dogs and having less risk of asthma there was detected a slight risk of pneumococcal disease. This links to the aforementioned hypothesis: more infections and fewer allergies (Steward 2015), but with the advantage that infections are easily treated or prevented with vaccines.
Belkaid Y, Segre JA (2014) Dialogue between skin microbiota and immunity. Science 346, 954-959
Fall T, Lundholm C, Örtqvist AK, Fall K, Fang F, Hedhammar Å, et al (2015) Early Exposure to Dogs and Farm Animals and the Risk of Childhood Asthma. JAMA Pediatrics 69(11), e153219
García-Mazcorro JF, Minamoto Y (2013) Gastrointestinal microorganisms in cats and dogs: a brief review. Arch Med Vet 45, 111-124
Heath WR, Carbone FR (2013) The skin-resident and migratory immune system in steady state and memory: innate lymphocytes, dendritic cells and T cells. Nature Immunology 14, 978-985
Marsland BJ, Gollwitzer ES (2014) Host–microorganism interactions in lung diseases. Nature Reviews Immunology 14, 827-835
Okada H, Kuhn C, Feillet H, Bach JF (2010) The “hygiene hypothesis” for autoimmune and allergic diseases: an update. Clin Exp Immunol 160, 1-9
Song SJ, Lauber C, Costello EK, Lozupone, Humphrey G, Berg-Lyons D, et al (2013) Cohabiting family members share microbiota with one another and with their dogs. eLife 2, e00458, 1-22
Steward D (2015) Dogs, microbiomes, and asthma risk: do babies need a pet ? MD Magazine, Nov 03
Yatsunenko T, Rey FE, Manary MJ, Trehan I, Dominguez-Bello MG, Contreras M, et al. 2012. Human gut microbiome viewed across age and geography. Nature 486, 222–7
21st March 2015
Clostridia: who are they ?
The clostridia or Clostridiales, with Clostridium and other related genera, are Gram-positive sporulating bacteria. They are obligate anaerobes, and belong to the taxonomic phylum Firmicutes. This phylum includes clostridia, the aerobic sporulating Bacillales (Bacillus, Listeria, Staphylococcus and others) and also the anaerobic aero-tolerant Lactobacillales (id est, lactic acid bacteria: Lactobacillus, Leuconostoc, Oenococcus, Pediococcus, Lactococcus, Streptococcus, etc.). All Firmicutes have regular shapes of rod or coccus, and they are the evolutionary branch of gram-positive bacteria with low G + C content in their DNA. The other branch of evolutionary bacteria are gram-positive Actinobacteria, of high G + C and irregular shapes, which include Streptomyces, Corynebacterium, Propionibacterium, and Bifidobacterium, among others.
Being anaerobes, the clostridia have a fermentative metabolism of both carbohydrates and amino acids, being primarily responsible for the anaerobic decomposition of proteins, known as putrefaction. They can live in many different habitats, but especially in soil and on decaying plant and animal material. As we will see below, they are also part of the human intestinal microbiota and of other vertebrates.
The best known clostridia are the bad ones (Figure 1): a) C. botulinum, which produces botulin, the botulism toxin, although nowadays has medical and cosmetic applications (Botox); b) C. perfringens, the agent of gangrene; c) C. tetani, which causes tetanus; and d) C. difficile, which is the cause of hospital diarrhea and some postantibiotics colitis.
Figure 1. The four more pathogen species of Clostridium. Image from http://www.tabletsmanual.com/wiki/read/botulism
Clostridia in gut microbiota
As I mentioned in a previous post (Bacteria in the gut …..) of this blog, we have a complex ecosystem in our gastrointestinal tract, and diverse depending on each person and age, with a total of 1014 microorganisms. Most of these are bacteria, besides some archaea methanogens (0.1%) and some eukaryotic (yeasts and filamentous fungi). When classical microbiological methods were carried out from samples of colon, isolates from some 400 microbial species were obtained, belonging especially to proteobacteria (including Enterobacteriaceae, such as E. coli), Firmicutes as Lactobacillus and some Clostridium, some Actinobacteria as Bifidobacterium, and also some Bacteroides. Among all these isolates, some have been recognized with positive effect on health and are used as probiotics, such as Lactobacillus and Bifidobacterium, which are considered GRAS (Generally Recognized As Safe).
But 10 years ago culture-independent molecular tools began to be used, by sequencing of ribosomal RNA genes, and they have revealed many more gut microorganisms, around 1000 species. As shown in Figure 2, taken from the good review of Rajilic-Stojanovic et al (2007), there are clearly two groups that have many more representatives than thought before: Bacteroides and Clostridiales.
Figure 2. Phylogenetic tree based on 16S rRNA gene sequences of various phylotypes found in the human gastrointestinal tract. The proportion of cultured or uncultured phylotypes for each group is represented by the colour from white (cultured) passing through grey to black (uncultured). For each phylogenetic group the number of different phylotypes is indicated (Rajilic-Stojanovic et al 2007)
In more recent studies related to diet such as Walker et al (2011) — a work done with faecal samples from volunteers –, population numbers of the various groups were estimated by quantitative PCR of 16S rRNA gene. The largest groups, with 30% each, were Bacteroides and clostridia. Among Clostridiales were included: Faecalibacterium prausnitzii (11%), Eubacterium rectale (7%) and Ruminococcus (6%). As we see the clostridial group includes many different genera besides the known Clostridium.
In fact, if we consider the population of each species present in the human gastrointestinal tract, the most abundant seems to be a clostridial: F. prausnitzii (Duncan et al 2013).
Benefits of some clostridia
These last years it has been discovered that clostridial genera of Faecalibacterium, Eubacterium, Roseburia and Anaerostipes (Duncan et al 2013) are those which contribute most to the production of short chain fatty acids (SCFA) in the colon. Clostridia ferment dietary carbohydrate that escape digestion producing SCFA, mainly acetate, propionate and butyrate, which are found in the stool (50-100 mM) and are absorbed in the intestine. Acetate is metabolized primarily by the peripheral tissues, propionate is gluconeogenic, and butyrate is the main energy source for the colonic epithelium. The SCFA become in total 10% of the energy obtained by the human host. Some of these clostridia as Eubacterium and Anaerostipes also use as a substrate the lactate produced by other bacteria such as Bifidobacterium and lactic acid bacteria, producing finally also the SCFA (Tiihonen et al 2010).
Clostridia of microbiota protect us against food allergen sensitization
This is the last found positive aspect of clostridia microbiota, that Stefka et al (2014) have shown in a recent excellent work. In administering allergens (“Ara h”) of peanut (Arachis hypogaea) to mice that had been treated with antibiotics or to mice without microbiota (Germ-free, sterile environment bred), these authors observed that there was a systemic allergic hyper reactivity with induction of specific immunoglobulins, id est., a sensitization.
In mice treated with antibiotics they observed a significant reduction in the number of bacterial microbiota (analysing the 16S rRNA gene) in the ileum and faeces, and also biodiversity was altered, so that the predominant Bacteroides and clostridia in normal conditions almost disappeared and instead lactobacilli were increased.
To view the role of these predominant groups in the microbiota, Stefka et al. colonized with Bacteroides and clostridia the gut of mice previously absent of microbiota. These animals are known as gnotobiotic, meaning animals where it is known exactly which types of microorganisms contain.
In this way, Stefka et al. have shown that selective colonization of gnotobiotic mice with clostridia confers protection against peanut allergens, which does not happen with Bacteroides. For colonization with clostridia, the authors used a spore suspension extracted from faecal samples of healthy mice and confirmed that the gene sequences of the extract corresponded to clostridial species.
So in effect, the mice colonized with clostridia had lower levels of allergen in the blood serum (Figure 3), had a lower content of immunoglobulins, there was no caecum inflammation, and body temperature was maintained. The mice treated with antibiotics which had presented the hyper allergic reaction when administered with antigens, also had a lower reaction when they were colonized with clostridia.
Figure 3. Levels of “Ara h” peanut allergen in serum after ingestion of peanuts in mice without microbiota (Germ-free), colonized with Bacteroides (B. uniformis) and colonized with clostridia. From Stefka et al (2014).
In addition, in this work, Stefka et al. have conducted a transcriptomic analysis with microarrays of the intestinal epithelium cells of mice and they have found that the genes producing the cytokine IL-22 are induced in animals colonized with clostridia, and that this cytokine reduces the allergen uptake by the epithelium and thus prevents its entry into the systemic circulation, contributing to the protection against hypersensitivity. All these mechanisms, reviewed by Cao et al (2014), can be seen in the diagram of Figure 4.
In conclusion, this study opens new perspectives to prevent food allergies by modulating the composition of the intestinal microbiota. So, adding these anti-inflammatory qualities to the production of butyrate and other SCFA, and the lactate consumption, we must start thinking about the use of clostridia for candidates as probiotics, in addition to the known Lactobacillus and Bifidobacterium.
Figure 4. Induction of clostridia on cytokine production by epithelial cells of the intestine, as well as the production of short chain fatty acids (SCFA) by clostridia (Cao et al 2014).
Cao S, Feehley TJ, Nagler CR (2014) The role of commensal bacteria in the regulation of sensitization to food allergens. FEBS Lett 588, 4258-4266
Duncan SH, Flint HJ (2013) Probiotics and prebiotics and health in ageing populations. Maturitas 75, 44-50
Rajilic-Stojanovic M, Smidt H, de Vos WM (2007) Diversity of the human gastrointestinal tract microbiota revisited. Environ Microbiol 9, 2125-2136
Rosen M (2014) Gut bacteria may prevent food allergies. Science News 186, 7, 4 oct 2014
Russell SL, et al. (2012) Early life antibiotic-driven changes in microbiota enhance susceptibility to allergic asthma. EMBO Rep 13(5):440–447
Stefka AT et al (2014) Commensal bacteria protect against food allergen sensitization. Proc Nat Acad Sci 111, 13145-13150
Tiihonen K, Ouwehand AC, Rautonen N (2010) Human intestinal microbiota and healthy aging. Ageing Research Reviews 9:107–16
Walker AW et al (2011) Dominant and diet-responsive groups of bacteria within the human colonic microbiota. The ISME J 5, 220-230
It seems to be so: the microbes in our gastrointestinal tract (GIT) influence our choice of food. No wonder: microbes, primarily bacteria, are present in significant amounts in GIT, more than 10 bacterial cells for each of our cells, a total of 1014 (The human body has about 1013 cells). This amounts to about 1-1.5 kg. And these bacteria have lived with us always, since all mammals have them. So, they have evolved with our ancestors and therefore they are well suited to our internal environment. Being our bodies their habitat, much the better if they can control what reaches the intestine. And how can they do? Then giving orders to the brain to eat such a thing or that other, appropriate for them, the microbes.
Well, gone seriously, there is some previous work in this direction. It seems there is a relationship between preferences for a particular diet and microbial composition of GIT (Norris et al 2013). In fact, it is a two-way interaction, one of the many aspects of symbiotic mutualism between us and our microbiota (Dethlefsen et al 2007).
There is much evidence that diet influences the microbiota. One of the most striking examples is that African children fed almost exclusively in sorghum have more cellulolytic microbes than other children (De Filippo et al 2010).
The brain can also indirectly influence the gut microbiota by changes in intestinal motility, secretion and permeability, or directly releasing specific molecules to the gut digestive lumen from the sub epithelial cells (neurons or from the immune system) (Rhee et al 2009).
The GIT is a complex ecosystem where different species of bacteria and other microorganisms must compete and cooperate among themselves and with the host cells. The food ingested by the host (human or other mammal) is an important factor in the continuous selection of these microbes and the nature of food is often determined by the preferences of the host. Those bacteria that are able to manipulate these preferences will have advantages over those that are not (Norris et al 2013).
Recently Alcock et al (2014) have reviewed the evidences of all this. Microbes can manipulate the feeding behaviour of the host in their own benefit through various possible strategies. We’ll see some examples in relation to the scheme of Figure 2.
Figure 2. As if microbes were puppeteers and we humans were the puppets, microbes can control what we eat by a number of marked mechanisms. Adapted from Alcock et al 2014.
People who have “desires” of chocolate have different microbial metabolites in urine from people indifferent to chocolate, despite having the same diet.
Dysphoria, id est, human discomfort until we eat food which improve microbial “welfare”, may be due to the expression of bacterial virulence genes and perception of pain by the host. This is because the production of toxins is often triggered by a low concentration of nutrients limiting growth. The detection of sugars and other nutrients regulates virulence and growth of various microbes. These directly injure the intestinal epithelium when nutrients are absent. According to this hypothesis, it has been shown that bacterial virulence proteins activate pain receptors. It has been shown that fasting in mice increases the perception of pain by a mechanism of vagal nerve.
Microbes can also alter food preferences of guests changing the expression of taste receptors on the host. In this sense, for instance germ-free mice prefer more sweet food and have a greater number of sweet receptors on the tongue and intestine that mice with a normal microbiota.
The feeding behaviour of the host can also be manipulated by microbes through the nervous system, through the vagus nerve, which connects the 100 million neurons of the enteric nervous system from the gut to the brain via the medulla. Enteric nerves have receptors that react to the presence of certain bacteria and bacterial metabolites such as short chain fatty acids. The vagus nerve regulates eating behaviour and body weight. It has been seen that the activity of the vagus nerve of rats stimulated with norepinephrine causes that they keep eating despite being satiated. This suggests that GIT microbes produce neurotransmitters that can contribute to overeating.
Neurotransmitters produced by microbes are analogue compounds to mammalian hormones related to mood and behaviour. More than 50% of dopamine and most of serotonin in the body have an intestinal origin. Many persistent and transient inhabitants of the gut, including E. coli, several Bacillus, Staphylococcus and Proteus secrete dopamine. In Table 1 we can see the various neurotransmitters produced by GIT microbes. At the same time, it is known that host enzymes such as amine oxidase can degrade neurotransmitters produced by microorganisms, which demonstrates the evolutionary interactions between microbes and hosts.
Table 1. Diversity of neurotransmitters isolated from several microbial species (Roschchina 2010)
|GABA (gamma-amino-butyric acid)||Lactobacillus, Bifidobacterium|
|Norepinephrine||Escherichia, Bacillus, Saccharomyces|
|Serotonin||Candida, Streptococcus, Escherichia, Enterococcus|
Some bacteria induce hosts to provide their favourite nutrients. For example, Bacteroides thetaiotaomicron inhabits the intestinal mucus, where it feeds on oligosaccharides secreted by goblet cells of the intestine, and this bacterium induces its host mammal to increase the secretion of these oligosaccharides. Instead, Faecalibacterium prausnitzii, a not degrading mucus, which is associated with B. thetaiotaomicron, inhibits the mucus production. Therefore, this is an ecosystem with multiple agents that interact with each other and with the host.
As microbiota is easily manipulated by prebiotics, probiotics, antibiotics, faecal transplants, and changes in diet, controlling and altering our microbiota provides a viable method to the otherwise insoluble problems of obesity and poor diet.
Alcock J, Maley CC, Aktipis CA (2014) Is eating behavior manipulated by the gastrointestinal microbiota? Evolutionary pressures and potential mechanisms. BioEssays 36, DOI: 10.1002/bies.201400071
De Filippo C, Cavalieri D, Di Paola M, Ramazzotti M, et al (2010) Impact of diet in shaping gut microbiota revealed by a comparative study in children from Europe and rural Africa. Proc Natl Acad Sci USA 107:14691–6
Dethlefsen L, McFall-Ngai M, Relman DA (2007) An ecological and evolutionary perspective on human-microbe mutualism and disease. Nature 449:811-818
Lyte M (2011) Probiotics function mechanistically as delivery for neuroactive compounds: Microbial endocrinology in teh design and use of probiotics. BioEssays 33:574-581
Norris V, Molina F, Gewirtz AT (2013) Hypothesis: bacteria control host appetites. J Bacteriol 195:411–416
Rhee SH, Pothoulakis C, Mayer EA (2009) Principles and clinical implications of the brain–gut–enteric microbiota axis. Nature Reviews Gastroenterology and Hepatology 6:306-314
Roschchina VV (2010) Evolutionary considerations of neurotransmitters in microbial, plant, and animal cells. In Lyte M, Freestone PPE, eds; Microbial Endocrinology: Interkingdom Signaling in Infectious Disease and Health. New York: Springer. pp. 17–52
It is really surprising, but it seems so: Italian and Austrian researchers have published a paper (Campisano et al. 2014) which shows that the bacterial species Propionibacterium acnes, related to human acne, can be found as obligate endophytes in bark tissues of Vitis vinifera, the grapevine.
Some bacterial pathogens of humans, such as Salmonella, are able to colonize plant tissues but temporarily and opportunistically (Tyler & Triplett 2008). In fact, there is a temporary mutual benefit between plants and bacteria, so some of these enterobacteria pathogenic to plants do not live endophytically and can be beneficial for them. These pathogens to humans, in its life cycle, use plants as alternative hosts to survive the environment, passing to the plants through contaminated irrigation water. Therefore, some bacteria are often temporary endophyte guests of plants.
But on the other hand, there are relatively rare cases of bacteria changing the host and adapting to the new host, finally being endophytes. This horizontal transfer happens mostly between evolutionarily close hosts, such as symbiotic bacteria of aphids (insects), which has proven to transfer to other species of aphids (Russell & Moran 2005). It has also been suggested the horizontal transfer of beneficial lactic acid bacteria (Lactobacillus reuteri) in the intestinal tract of vertebrates, since strains of this L. reuteri are similar in several species of mammals and birds.
Well, going beyond, the work of Campisano et al. subject of this review, concludes that bacteria associated with human acne should have passed on the vine, that is, the bacteria would have made a horizontal transfer interregnum, from plants to mammals.
Propionibacterium acnes type Zappae
Acne, as you know, is a common human skin disease, consisting of an excess secretion of the pilosebaceous glands caused by hormonal changes, especially teenagers. The glands become inflamed, the pores obstructed and scarring appears. The microorganism associated with these infections is the opportunistic commensal bacterium P. acnes, a gram-positive anaerobic aero tolerant rod, which fed fatty acids produced by the glands.
Young with acne (Wikimedia, public)
Propionibacterium acnes at the scanning electron microscope (left) and dyed with violet crystal (right). From Abate ME (2013) Student Pulse 5, 9, 1-4.
Interestingly, other species of the same genus Propionibacterium well known in microbial biotechnology industry are used for the production of propionic acid, vitamin B12, and the Swiss cheeses Gruyere or Emmental.
Campisano et al. have made a study of the vineyard endomicrobioma by the sequencing technique (Roche 454) amplifying the V5-V9 hyper variable region of the bacterial 16S rDNA present in the tissues of vine. In 54 of the 60 plants analyzed, between 0.5% and 5% of the found sequences correspond to the species Propionibacterium acnes. This observation has been confirmed by fluorescent in situ hybridization (FISH) with fluorochromes and specific probes of P. acnes.
Location of P. acnes (fluorescent blue spots) in the bark of a vine stem, seen with FISH microscopy with specific probes for this bacterium (Campisano et al 2004).
The authors of this work proposed for this bacterium the name of P. acnes Zappae, in memory of the eccentric musician and composer Frank Zappa, to emphasize the unexpected and unconventional habitat of this type of P. acnes.
Frank Zappa (1940-1993), the eccentric and satiric singer, musician and composer. Photo: Frank Zappa reviews.
And how did this human bacteria arrive into the vineyard?
To solve this riddle, Campisano et al. have taken the 16S rDNA sequences and from other genes (recA and tly) from these strains of P. acnes Zappae found in vine and have compared with those P. acnes of human origin in databases. Comparing phylogenies and clusters deducted from them, these researchers have concluded that P. a. Zappae has diversified evolutionarily recently. Studying in detail the recA gene sequences of P. a. Zappae, and taking into account the likely mutation rate and generation time (about 5 hours), they deduce that the diversification from other P. acnes occurred 6000-7000 years ago.
This date coincides with the known domestication of the vine by humans, which is believed to have occurred about 7000 years ago in the southern Caucasus, between the Black Sea and the Caspian Sea, the area of modern Turkey, Georgia, Armenia and Iran (Berkowitz 1996). The vineyard has its origins in a wild subspecies of Vitis that survived the Ice Age and was domesticated. This plant came out to three subspecies, and one of them, Vitis vinifera pontica, spread in the mentioned area and further south in Mesopotamia and then to all south Europe thanks to the Phoenicians.
Therefore, the conclusion is that P. acnes Zappae originated from human P. acnes 7000 years ago, by contact of human hands with grapes and other parts of the vineyard during the harvest and carrying them. As the authors say, this case would be the first evidence of horizontal transfer interregnum, from humans to plants, of a obligate symbiotic bacterium. This also makes more remarkable the adaptability of bacteria. Their ability to exploit new habitats can have unforeseen impacts on the evolution of host-symbiont relationship or even host-pathogen.
Harvesting by hand in Chile (Fine Wine and Good Spirits)
Berkowitz M (1996) World’s earliest wine. Archaeology 49, 5, Sept./Oct.
Campisano Aet al. (2014) Interkingdom transfer of the acne-causing agent, Propionibacterium acnes, from human to grapevine. Mol Biol Evol 31, 1059-1065.
Gruber K (4 march 2014) How grapevines got acne bacteria. Nature News 4 march 2014.
Russell JA, NA Moran (2005) Horizontal transfer of bacterial symbionts: heritability and fitness effects in a novel aphid host. Appl Environ Microbiol 71, 7987-7994.
Tyler HL, EW Triplett (2008) Plants as a habitat for beneficial and/or human pathogenic bacteria. Ann Rev Phytopathol 46, 53-73.
Walter J, RA Britton, S Roos (2011) PNAS 108, 4645-4652.
A few weeks ago I saw the following headline in “Recercat” (the electronic newsletter of research in Catalonia): “Científics descobreixen com es formen les molècules bàsiques de la vida“, that is, “Scientists discover how the basic molecules of life are made”. Oops what a heading ! I was very surprised, of course, because I have always been passionate for this subject of life’s origin. So, I quickly read it, and looked for the original article .
Well, once seen in detail, it is clear that the heading is very, very exaggerated, as supposed at first glance. Before discussing why I think it is exaggerated, I want to mention another detail of this headline that I think is superflous: Why the word “scientists” ? Whoelse might be working on the study of the formation of the basic molecules of life besides scientists ? Politicians perhaps? or economists or maybe the bishops ? It is clear that must be scientists and therefore it is no need to say it. The heading would be enough like this: “It has been discovered how…. ” . Or else, it could tell where scientists are from: “English scientists, or American, or Japanese, have discovered … “.
Well, going to the specific discovery, and as Recercat itself summarizes, some Americans and British researchers (Martins et al. 2013) have published in Nature Geoscience their laboratory work, where they simulated the impact of comets on the surface of a planet, shooting a projectile with a compressed air pistol at speeds of 7 km/s (25,000 km/h). They have seen that by the impact and the heat generated, amino acids are synthesized from water, CO2 and ammonia. This is what they call “shock synthesis”. Among others, the amino acids detected were glycine, D-alanine, L-alanine, amino isobutyric acid, isovaline, norvaline and other precursors of amino acids, both isomers D and L. The amounts detected were between nanograms and one microgram.
This synthesis process shows that a simple mechanism, such as the impact of comets on the surface of a rocky planet, can transform basic inorganic molecules to more complex organic molecules such as amino acids, which are the monomers of proteins, basic constituents of all living beings. And therefore, this shock synthesis could have been a step in the appearance of terrestrial life.
Now, is this the first work to demonstrate the possible formation of basic molecules of life ? Well, absolutely not ! This work has its value but does not deserve this headline so exaggerated.
So, let’s briefly review what is already known, since more than 50 years, many different researchers have worked on this topic and have been discovering aspects that reinforce the scientific hypotheses of abiogenesis. This, also known as biopoesis, is the natural process by which living organisms originated on earth from simple molecules about 3,700 million years ago.
Biopoesis, the process of emergence of living beings on Earth
This process of biopoesis necessarily involved several steps:
1) The formation and appearance of basic organic molecules of living beings, it is, the monomers such as amino acids, monosaccharides, fatty acids and nitrogenous bases.
2) From the above monomers, the formation of biogenic macromolecules, it is, polysaccharides, polypeptides, lipoids and others, by polymerization, probably on the surface of inorganic substrates such as clay or iron minerals.
3) And the formation of the protobionts, the precursors of first cells, from macromolecules. This key step, the most difficult to prove, was probably linked to the parallel acquisition of the three basic properties of living beings: i) a structure envelope (the membrane) of lipidic consistency; ii) some reactions transforming nutrients and energy (rudimentary metabolism); and iii) the ability of transfering characteristics to the offspring (hereditary mechanism) with an information-carrying molecule, probably RNA.
By the moment I will leave the steps 2 and 3, maybe for discussing in a future post, and I will focus on the first one, related to the mentioned article of Martins et al. (2013). This formation and appearance of compounds in the early Earth may have been by three mechanisms: a) in situ production, b) contributions from abroad: c) synthesis due to impacts.
These three categories of mechanisms were already raised as an inventory of the possible origins of life in 1992 in an article of Nature by Christopher Chyba and Carl Sagan, the famous pioneer of exobiology and science writer, very known by the extraordinary TV series Cosmos, and author of the phrase “We are stardust“. Coincidentally, he was the first husband of Lynn Margulis, who spread the endosymbiotic theory of the bacterial origin of mitochondria and chloroplasts.
Endogenous synthesis of organic compounds on the primitive Earth
Well, as I said, it has been demonstrating since many years the possibility of the formation of organic molecules in situ, id est, endogenous synthesis in the primitive Earth, without external inputs. The Oparin’s hypothesis that the reducing anaerobic conditions of the early atmosphere, along with solar energy, should favoured the synthesis of organic molecules forming the “prebiotic soup”, was demonstrated as possible by the known experiments of Miller and Urey:
In 1952, the graduate student Stanley Miller with his professor Harold Urey introduced a mixture of water, hydrogen, methane and ammonia in a cyclical container, where electrical sparks were applied. A week later, when analyzing the components, they found that 15% of the carbon coming from methane had been transformed in various organic compounds, including 5 amino acids, both D-and L-.
Schema of the experiments of Miller and Urey. From GCSE-Bitesize (BBC).
Recently (Parker et al 2011) the tubes with the extracts of original Miller- Urey experiments have been analyzed again with current sophisticated analytical techniques and equipment, and many more compounds that the found originally in the 1950s have been discovered, in particular 23 other amino acids.
The synthesis of these organic molecules in the early Earth was probably facilitated by energy sources of the atmospheric activity such as electrical discharges that were used by Miller and Urey experiments, but there were other possibilities, such as the sunlight, with more UV radiation than at present (there was no ozone layer since it was formed later, from oxygen), and also more volcanic activity and radioactivity in a younger Earth, and impacts of meteorites and comets, which is related to the mentioned work of Martins et al. (2013).
Another important contribution in the search for prebiotic organic synthesis was the demonstration made by Joan Oró (born in Lleida, Catalonia) working at NASA (Oró 1961) that adenine can be synthesized heating ammonium cyanide solutions. Similarly, recently it has been demonstrated the synthesis of pyrimidines (cytosine and uracil), adenine and triazines (other nitrogenous bases) from urea by freeze-thaw cycles and electric shocks (Menor-Salván et al. 2007).
As demonstrated by Joan Oró, adenine can be synthesized from 5 molecules of hydrogen cyanide. Adenine is a key molecule for life, since it is part of nucleic acids and ATP.
Contribution of organic molecules by extraterrestrial objects
In addition to the in situ synthesis of organic compounds in the primitive Earth, they could also have come from outside. The contribution of organic molecules by extraterrestrial objects, comets or meteorites or other, it is increasingly more evident scientifically. Recent studies suggest that the so-called massive bombing that took place 3.5 billion years ago provided an amount of organic compounds comparable to those produced in situ.
Simulation of meteor shower similar to this of the early Earth. From AZ-Revista de Educación y Cultura
It has been shown that organic compounds are relatively common in extraterrestrial space, especially in the outer solar system where volatile compounds are not evaporated by solar heat. Many comets have an outer layer of a material with appearance of tar, which contains organic compounds formed by reactions caused by radiation, especially UV. Apart from that long ago they had been detected by telescope spectrography, a few years ago it has been identified in situ the amino acid glycine in comet Wild -2 in samples taken by NASA’s Stardust spacecraft (Dolmetsch 2006) .
The Murchison meteorite, about 100 kg, fell in Australia in 1969 and broke in several fragments that have been well studied. This meteorite is of type carbonaceous chondrites, which are rich in carbon, and indeed contains amino acids, both common (glycine, alanine and glutamic acid) as the most unusual (isovaline, pseudoleucine), with concentrations up to 60 ppm (Kvenvolden et al. 1970). It also contains aromatic and aliphatic hydrocarbons, alcohols and other organic compounds such as carboxylic acids and fullerenes.
A fragment of the Murchison meteorite, fell in Australia in 1969, of the type carbonaceous chondrites, which contains amino acids and other organic compounds. Image from Wikipedia
The isotope ratio 12C/13C of uracil and other organic compounds in the Murchison indicates an origin no terrestrial (Martins et al. 2008). In fact, besides the Murchison, the analyses made with many other meteorites show that organic compounds can be formed in outer space.
Model studies done with computer suggest that prebiogenic organic compounds might be formed in the protoplanetary disk of dust that surrounded the sun before the formation of the Earth, and that the same process may happen around other stars (Moskowitz 2012).
Moreover, studies of infrared emission spectra (Kwok & Zhang 2011) of cosmic dust have concluded that complex organic molecules are produced in the supernova stars, and that these molecules are expelled to the interstar space by effect of the explosion of the star. Surprisingly, this organic dust is similar to compounds found in meteorites. Since meteorites are remnants of the early solar system, we can now suggest that organic compounds found in meteorites had formed in distant stars .
Infrared spectrum of organic compounds, superimposed on an image of the Orion Nebula where these complex organic compounds (with the formulas) have been found. Image taken from NASA (C.R. O’Dell and S.K. Wong, Rice University.
In recent years there has been a breakthrough in the detection of organic molecules in galactic space thanks to the radio telescopes such as the Green Bank (100 m diameter) in West Virginia (USA), and the ALMA (Atacama Large Millimeter Array, at 5000 m in the Atacama Desert, northern Chile) which is composed of 66 radio antennas of 12 m diameter connected to each other by optical fiber. These astronomical radio telescopes or interferometers capture wavelengths around mm.
With these telescopes, as commented by Pere Brunet last month in his post post “Som pols d’estrelles ?” (Are we stardust ?, of the blog Fractal Ara-Ciència), different compounds have been detected around other stars: propenal, cyclo-propenone, acetamide, and glycolaldehyde (CHO-CH2OH). The latter is quite significant, since it is the smallest sugar and it is necessary for the formation of RNA. It has been detected with ALMA, around a young binary sun-type star (IRAS 16293-2422 ), 400 light years from Earth, relatively close, within the Milky Way (Jørgensen et al. 2012) .
Synthesis of organic compounds due to impacts of extraterrestrial objects
Finally, we have this third mechanism related with the article object of this post (Martins et al. 2013). Well, as we said before, this possibility was already reviewed by Chyba & Sagan (1992), because experiments in this direction had been made years ago. Carl Sagan itself, with other authors (Bar-Nun et al. 1970) had shown that applying a thermal shock, simulating impacts of comets and micrometeorites, to a gas mixture similar to the primitive atmosphere, there appeared amino acids.
However, thinking about the impacts of extraterrestrial bodies the first idea is the opposite, that they are antagonists of life on Earth because we remember the impact which caused extinctions and catastrophes such as the asteroid 65 million years falling in the Chicxulub crater in Yucatan, or a much smaller scale, the meteorite that fell last year in Chelyabinsk, Russia, with the appearance of fireballs. However, although these collisions can cause adverse effects on living beings where they fall, at the same time the energy released by the shock can be a source of reactions that generate prebiogenic organic compounds, as evidenced by the work of Martins et al. (2013).
At the same time, it has been shown that biological compounds that were present in the meteorite can “survive” impacts. Indeed, it has been shown that these compounds may be captured in the carbonaceous pores, inside the molten material due to the temperature and pressure of impact, particularly in analyses made with material of Darwin Crater in Tasmania, coming from a meteorite that impacted 800,000 years ago (Howard et al. 2013).
Besides from Martins et al., other studies have also simulated the effects of impacts. Furukawa et al. (2009) simulated the impact of a meteorite chondrite-type in a primitive ocean. They used a propulsion gun to create a high-speed impact in a mixture of carbon, iron, nickel, nitrogen and water, and then they recovered several organic molecules, including fatty acids, amines and one amino acid.
Thus, these experiments suggest that the impact of frequent extraterrestrial bodies in the primitive Earth had resulted in a great contribution to the formation of many different organic compounds, and as I said, adding that to the contribution of the already previously synthesized in outer space and to in situ synthesis in the same Earth.
Bar-Nun A, Bar-Nun N, Bauer SH, Sagan C. 1970. Shock synthesis of amino acids in simulated primitive environments. Science 168, 470-472.
Chyba C, Sagan C. 1992. Endogenous production, exogenous delivery and impact-shock synthesis of organic molecules: an inventory for the origins of life. Nature 355, 125–32
Dolmetsch C. 2006. NASA Spacecraft Returns With Comet Samples After 2.9 Bln Miles. Bloomberg.com. 2006-01-15
Editorial. 2013. The upside of impacts. Nature Geoscience 6, 987.
Furukawa Y, Sekine T, Oba M, Kakegawa T, Nakazawa H. 2009. Biomolecule formation by oceanic impacts on early Earth. Nature Geoscience 2, 62–66
Generalitat de Catalunya. 2014. “Científics descobreixen com es formen les molècules bàsiques de la vida”. Recercat 94, gener 2014.
Howard KT et 12 al. 2013. Biomass preservation in impact melt ejecta. Nature Geoscience 6, 1018-1023.
Jørgensen JK, Favre C, Bisschop SE, Bourke TL, van Dishoeck EF, Schmalzl M. 2012. Detection of the simplest sugar, glycolaldehyde, in a solar-type protostar with ALMA. Astrophysical Journal Letters 757, L4, 1-13.
Kvenvolden KA, Lawless J, Pering K, Peterson E, Flores J, Ponnamperuma C, Kaplan IR, Moore C. 1970. Evidence for extraterrestrial amino-acids and hydrocarbons in the Murchison meteorite. Nature 228, 923–926
Kwok S, Zhang Y. 2011. Mixed aromatic–aliphatic organic nanoparticles as carriers of unidentified infrared emission features. Nature, DOI:10.1038/nature10542
Martins Z, Botta O, Fogel ML, Sephton MA, Glavin DP, Watson JS, Dworkin JP, Schwartz AW, Ehrenfreund P. 2008. Extraterrestrial nucleobases in the Murchison meteorite. Earth and Planetary Science Letters 270, 130–136
Martins Z, MC Price, N Goldman, MA Sephton, MJ Burchell. 2013. Shock synthesis of amino acids from impacting cometary and icy planet surface analogues. Nature Geoscience 6, 1045-1049.
Moskowitz C. 2012. Life’s Building Blocks May Have Formed in Dust Around Young Sun. Space.com
Menor-Salván C, Ruiz-Bermejo DM, Guzmán MI, Osuna-Esteban S, Veintemillas-Verdaguer S. 2007. Synthesis of pyrimidines and triazines in ice: implications for the prebiotic chemistry of nucleobases. Chemistry 15, 4411–8.
Oparin A. 1952. The origin of life. New York: Dover.
Oró J. 1961. Mechanism of synthesis of adenine from hydrogen cyanide under possible primitive Earth conditions. Nature 191, 1193–4.
Parker ET, Cleaves HJ, Dworkin JP et al. 2011. Primordial synthesis of amines and amino acids in a 1958 Miller H2S-rich spark discharge experiment. PNAS 108, 5526–31.
Wikipedia: http://en.wikipedia.org/wiki/Abiogenesis (very good review of life’s origin andthe different hypothesis)