Chapter 5 Horizontal Gene Transfer Elements: Plasmids in Antarctic Microorganisms Matías Giménez, Gastón Azziz, Paul R. Gill, and Silvia Batista Abstract Plasmids play an important role in the evolution of microbial communi- ties. These mobile genetic elements can improve host survival and may also be involved in horizontal gene transfer (HGT) events between individuals. Diverse culture-dependent and culture-independent approaches have been used to character- ize these mobile elements. Culture-dependent methods are usually associated with classical microbiological techniques. In the second approach, development of spe- cific protocols for analysis of metagenomes involves many challenges, including assembly of sequences and availability of a reliable database, which are crucial. In addition, alternative strategies have been developed for the characterization of plas- mid DNA in a sample, generically referred to as plasmidome. The Antarctic continent has environments with diverse characteristics, including some with very low temperatures, humidity levels, and nutrients. The presence of microorganisms and genetic elements capable of being transferred horizontally has been confirmed in these environments, and it is generally accepted that some of these elements, such as plasmids, actively participate in adaptation mechanisms of host microorganisms. Information related to structure and function of HGT elements in Antarctic bac- teria is very limited compared to what is known about HGT in bacteria from temper- ate/tropical environments. Some studies are done with biotechnological objectives. The search for mobile elements, such as plasmids, may be related to improve the expression of heterologous genes in host organisms growing at very low tempera- tures. More recently, however, additional studies have been done to detect plasmids in isolates, associated or not with specific phenotypes such as drug resistance. Although various Antarctic metagenomes are available in public databases, corre- M. Giménez · G. Azziz · S. Batista (*) Unidad Microbiología Molecular, Instituto de Investigaciones Biológicas Clemente Estable, Montevideo, Uruguay e-mail: [email protected] P. R. Gill Nevada City Biolabs, Nevada City, CA, USA © Springer Nature Switzerland AG 2019 85 S. Castro-Sowinski (ed.), The Ecological Role of Micro-organisms in the Antarctic Environment, Springer Polar Sciences, https://doi.org/10.1007/978-3-030-02786-5_5 86 M. Giménez et al. sponding studies of plasmidomes are needed. The difficulties usually associated with the study of metagenomes are increased in these cases by the limited number of sequences in functionally characterized databases. Keywords Horizontal gene transfer · Plastmidome · Global climate change · Animal and human influence · Antibiotic resistance genes 5.1 Definition of Biogeographic Regions Classically, the Antarctic Polar Frontal Zone is classified into three regions with distinct climates and accompanying biota: Continental Antarctica, Maritime Antarctica, and Sub-Antarctica (Convey 2010). The well-studied McMurdo Dry Valleys of Continental Antarctica that have been ice-free for many years are consid- ered to be a polar desert. Maritime Antarctica, which includes northwestern parts of the Antarctic Peninsula and surrounding Islands, has a cool moist climate and is less extreme in terms of high irradiance and low temperature compared with Continental Antarctica. The sub-Antarctic region includes various islands located between 35 and 60°S. These islands are breeding sites for seabird and mammal species and exhibit contrasting higher biodiversity compared with both the other regions. The incorporation of new data and criteria for classification has allowed the iden- tification of 16 Antarctic Conservation Biogeographic Regions (ACBRs) (Terauds and Lee 2016). These bioregions, located in ice-free areas (comprising 0.4% of the continent’s surface), are internationally recognized and studied for different pur- poses including biodiversity analysis (Fig. 5.1). 5.2 Characteristic of Antarctic Soil in ACBRs Soil properties of different ACBR sites are diverse, depending on their location, topography, local climate, and associated biota. Some soils have low carbon (C) and nitrogen (N) content, whereas those exposed to birds, like penguins, have relatively high C and N levels (Cowan 2014). Salt content is variable, depending on distance to a coastline. Some of these soils are especially dry, like those in the Dry Valleys, while others have comparatively higher humidity. Human activities, including those associated with scientific bases and tourism, are concentrated in these regions with their inevitable effects. Global climate change is also affecting these bioregions, in which retreating glaciers are exposing new surfaces with their particular soils. 5.3 Effect of Global Climate Change Effects of global climate change have been analyzed on the continent. Recently, measurements of changes in Antarctic ice sheets under the surface of the Southern ocean was determined using novel strategies, including satellite altimeter 5 Horizontal Gene Transfer Elements: Plasmids in Antarctic Microorganisms 87 Fig. 5.1 Location of biogeographic regions in Antarctica, numbered from 1 to 16. The arrows show approximately the sampling sites mentioned in the studies included in the chapter determinations of ice-elevation changes and ice geometry measurements (Konrad et al. 2018). They showed that 1463 km2 of ice melted between 2010 and 2016. In about 10.7% of oceanfront glaciers, the “grounding line,” where ocean, ice, and bedrock meet, glaciers are melting at a significant rate, probably because of the influence of warmer ocean water. In contrast, only 1.9% of these glaciers are actu- ally still expanding. Some Antarctic regions are especially vulnerable to global warming. Between 1979 and 1997, mean annual temperature in the Antarctic Peninsula increased about 0.32 °C per decade but dropped −0.47 °C per decade, from 1998 to 2014 (Sancho et al. 2017). These changes have further influenced public opinion especially with the occurrence of dramatic events, like the collapse of relatively large ice shelves (Shepherd et al. 2012). 88 M. Giménez et al. The effects of climate change on biota have been analyzed at different sites. One such study focused on six lichen species growing on surfaces of the Antarctic Peninsula that became deglaciated more than 20 years ago (Sancho et al. 2017). Some microbiota found to be in association with these organisms has also been evaluated. Antarctic hair grass (Deschampsia antarctica) has been spreading in some Maritime Antarctic sites (Smith 1994), affecting other organisms including mosses and soil microbiota. Microbes associated with these vascular plants incorporate soil N at higher rates compared with microbes associated with nonvascular plants. The increase in temperature in these regions promotes processes such as soil organic matter decomposition, thereby allowing for the further spread of hair grass. During winter, winds disperse remaining surface organic matter, affecting more areas. 5.4 Animal and Human Influence on Maritime Antarctica Some Maritime Antarctic locations are particularly affected by significant biotic and abiotic influence. King George Island, the largest island in Maritime Antarctica, e.g., is home to numerous marine mammals, including elephant, Weddell and leop- ard seals, as well as chinstrap and gentoo penguins. Whales sometimes frequent the nutrient-rich shoreline, but animal life on the island is restricted to seabirds and very small invertebrates. This island is one of the most inhabited or visited sites on the continent. Nine permanent research stations are in operation, and several research refuges have been constructed. More than 500 persons may visit the island during summer, and numerous research projects in many fields and by many countries are carried out in the region. 5.5 Strategies for the Study of Microbial Communities Microorganisms are essential for maintenance of diverse processes including soil structure formation, decomposition of xenobiotics, metabolism of organic matter, and support of biogeochemical cycles of specific elements such as C, N, and phos- phorous (P). Also, the study of microorganisms and their interactions with biotic and abiotic factors could be used for the development of bioremediation strategies, biotechnological processes, evaluation of soil management, etc. The first microbiological studies in Antarctica were initiated during the twentieth century (Darling and Siple 1941; Straka and Stokes 1960). These studies involved culture and isolation of microorganisms, and results obtained supported the idea of there being relatively low abundance and diversity in microbial assemblages. Microscopy has also been used to evaluate abundance, succession, and distribu- tion of microorganisms in Antarctic soil, microbial mats, etc. Ramsay (1983) stud- ied bacterial presence in ornithogenic soils by fluorescence microscopy using acridine orange, a dye that emits green fluorescence when bound to dsDNA and red 5 Horizontal Gene Transfer Elements: Plasmids in Antarctic Microorganisms 89 fluorescence when bound to ssDNA or RNA. Samples were collected at Cape Bird (Ross Island), and differences in numbers and forms of bacteria were found, depend- ing on sampling location and particularly near rookeries. In that study, the author mentions problems that some investigators had when attempting to recover cultured bacteria from these types of sampling locations.