SPECIAL SECTION: MICROBIAL DIVERSITY Extremophilic microbes: Diversity and perspectives T. Satyanarayana1,*, Chandralata Raghukumar2 and S. Shivaji3 1Department of Microbiology, University of Delhi South Campus, New Delhi 110 021, India 2Biological Oceanography Division, National Institute of Oceanography, Dona Paula, Goa 403 004, India 3Centre for Cellular and Molecular Biology, Uppal Road, Hyderabad 500 007, India Life in extreme environments has been studied inten- A variety of microbes inhabit extreme environments. Extreme is a relative term, which is viewed compared sively focusing attention on the diversity of organisms to what is normal for human beings. Extreme envi- and molecular and regulatory mechanisms involved. The ronments include high temperature, pH, pressure, salt products obtainable from extremophiles such as proteins, concentration, and low temperature, pH, nutrient enzymes (extremozymes) and compatible solutes are of concentration and water availability, and also condi- great interest to biotechnology. This field of research has tions having high levels of radiation, harmful heavy also attracted attention because of its impact on the pos- metals and toxic compounds (organic solvents). Cul- sible existence of life on other planets. ture-dependent and culture-independent (molecular) The progress achieved in research on extremophiles/ methods have been employed for understanding the thermophiles is discussed in international conferences diversity of microbes in these environments. Extensive held every alternate year in different countries. The last global research efforts have revealed the novel diver- conference on extremophiles was held in USA in 2004, sity of extremophilic microbes. These organisms have evolved several structural and chemical adaptations, and that on thermophiles in England in 2003, and the forth- which allow them to survive and grow in extreme en- coming one is due in Australia in 2005. In this article, an vironments. The enzymes of these microbes, which attempt has been made to review the diversity of microor- function in extreme environments (extremozymes), ganisms that occur in extreme environments, their adaptations have several biotechnological applications. Antibiotics, and potential biotechnological applications. The ‘Great compatible solutes and other compounds obtainable Plate Count Anomaly’ wherein only a miniscule fraction from these microbes are also finding a variety of uses. of microorganisms (<5%) are detected using the culture- dependent methods as compared to in situ detection MODERATE environments are important to sustain life. methods and culture independent methods, such as the Moderate means environments with pH near neutral, ‘rRNA approach’ is now well known. The latter approach temperature between 20 and 40°C, air pressure 1 atm and has indeed revealed a greater degree of biodiversity but adequate levels of available water, nutrients and salts. suffers from the fact that other characteristics of living Many extreme environments, such as acidic or hot springs, organisms cannot be ascertained. Therefore, there is a saline and/or alkaline lakes, deserts and the ocean beds need to have methods, which not only reflect the phylogeny are also found in nature, which are too harsh for normal of the bacteria, but also their biotechnological potential life to exist. Any environmental condition that can be and this indeed is possible using the ‘metagenome’ approach perceived as beyond the normal acceptable range is an discussed elsewhere in this special section. extreme condition. A variety of microbes, however, survive Metagenome represents the genomes of the total micro- and grow in such environments. These organisms, known biota found in nature, and involves cloning of total DNA as extremophiles, not only tolerate specific extreme con- isolated from an environmental sample in a bacterial arti- dition(s), but usually require these for survival and fical chromosome (BAC) vector and expressing the genes growth. Most extremophiles are found in microbial in a suitable host. Thus the metagenome approach, apart world. The range of environmental extremes tolerated by from providing phylogenetic data based on 16S rRNA gene microbes is much broader than other life forms. The lim- sequence analysis and enumerating the extent of micro- its of growth and reproduction of microbes are, –12° to bial diversity, also provides an access to genetic informa- more than +100°C, pH 0 to 13, hydrostatic pressures up tion of uncultured microorganisms. In fact, metagenome to 1400 atm and salt concentrations of saturated brines. libraries could form the basis for identification of novel Besides natural extreme environments, there are man- genes from uncultured microorganisms. Recognizing the made extreme conditions such as coolhouses, steam- potential of this approach, molecular biologists and chem- heated buildings and acid mine waters. ists have undertaken a project to establish the metagenome of ‘Whole earth’ (soil) to catalogue genes for useful natu- *For correspondence. (e-mail: [email protected]) ral products such as enzymes, antibiotics, immunosup- 78 CURRENT SCIENCE, VOL. 89, NO. 1, 10 JULY 2005 SPECIAL SECTION: MICROBIAL DIVERSITY pressants and anticancer agents. Thus it may not be too growing above boiling point of water have been isolated long before this approach is also customized to hunt for from hydrothermal vents, where hydrostatic pressure genes of interest in metagenomes from various extreme keeps the water in liquid state; however, majority of them environments. are anaerobes. Thermal vent sites have recently been found in the Indian Ocean. The primary areas with pH lower than 3 are those where Extreme environments relatively large amounts of sulphur or pyrite are exposed to oxygen. Both sulphur and pyrite are oxidized abioti- Low temperature (cold) environments prevail in fresh and cally through an exothermic reaction where the former is marine waters, polar and high alpine soils and waters, oxidized to sulphuric acid, and the ferrous iron in the lat- glaciers, plants and animals. The major man-made cold ter to ferric form. Both these processes occur abiotically, environments are refrigerators and freezers of industrial but are increased 106 times through the activity of acido- and domestic food storage. Oceans represent 71% of earth’s philes. Most of the acidic pyrite areas have been created surface and 90% by volume, which are at 5°C or colder. by mining and are commonly formed around coal, lignite At altitudes >3000 m, the temperature of the atmosphere or sulphur mines. All such areas have very high sulphide is consistently <5°C. As the altitude increases, the tem- concentrations, and pH values as low as 1. These are very perature decreases progressively and temperatures below low in organic matter, and are quite toxic due to high –40°C have been recorded. Low temperatures are charac- concentrations of heavy metals. In all acidic niches, the teristic of mountains where snow or ice remains year-round. acidity is mostly due to sulphuric acid. Due to spontane- The temperature is cold, part of the year, on mountains ous combustion, the refuse piles are self-heating and pro- where snow or ice melts. Thus, cold environment dominates vide the high-temperature environment required to sustain the biosphere. According to Morita1, cold environments thermophiles. The illuminated regions, such as mining can be divided into two categories: psychrophilic (perma- outflows and tailings dams, support phototrophic algae. nently cold) and psychrotrophic (seasonally cold or where In alkaline environments such as soils, increase in pH temperature fluxes into mesophilic range) environments. is due to microbial ammonification and sulphate reduction, Although habitats with elevated temperatures are not as and by water derived from leached silicate minerals. The widespread as temperate or cold habitats, a variety of high pH of these environments fluctuates due to their limited temperature, natural and man-made habitats exist. These buffering capacity and therefore, alkalitolerant microbes include volcanic and geothermal areas with temperatures are more abundant in these habitats than alkaliphiles. The often greater than boiling, sun-heated litter and soil or best studied and most stable alkaline environments are sediments reaching 70°C, and biological self-heated envi- soda lakes and soda deserts (e.g. East African Rift valley, ronments such as compost, hay, saw dust and coal refuse Indian Sambhar Lake). These are characterized by the piles. In thermal springs, the temperature is above 60°C, presence of large amounts of Na2CO3 but are significantly and it is kept constant by continual volcanic activity. Be- depleted in Mg2+ and Ca2+ due to their precipitation as sides temperature, other environmental parameters such carbonates. The salinity ranges from 5% (w/v) to saturation as pH, available energy sources, ionic strength and nutri- (33%). Industrial processes including cement manufac- ents influence the diversity of thermophilic microbial ture, mining, disposal of blast furnace slag, electroplating, populations. The best known and well-studied geothermal food processing and paper and pulp manufacture produce areas are in North America (Yellowstone National Park), man-made unstable alkaline environments. Iceland, New Zealand, Japan, Italy and the Soviet Union. Soils and oceans represent typical low nutrient envi- Hot water springs are situated throughout the length and ronments. Open oceans and