Interactions between Bacillus anthracis and Plants May Promote Anthrax Transmission Holly H. Ganz 1¤a *, Wendy C. Turner 1¤b , Eoin L. Brodie 1,2 , Martina Kusters 3 , Ying Shi 4 , Heniritha Sibanda 5 , Tamas Torok 2 , Wayne M. Getz 1,6 1 Department of Environmental Science, Policy and Management, University of California, Berkeley, Berkeley, California, United States of America, 2 Ecology Department, Earth Science Division, Lawrence Berkeley National Laboratory, Berkeley, California, United States of America, 3 Independent researcher, Windhoek, Namibia, 4 Department of Statistics, University of California, Berkeley, California, United States of America, 5 Ministry of Fisheries and Marine Resources, Inland Aquaculture, Katima Mulilo Regional Office, Katima Mulilo, Namibia, 6 School of Mathematical Sciences, University of KwaZulu-Natal, Durban, South Africa Abstract Environmental reservoirs are essential in the maintenance and transmission of anthrax but are poorly characterized. The anthrax agent, Bacillus anthracis was long considered an obligate pathogen that is dormant and passively transmitted in the environment. However, a growing number of laboratory studies indicate that, like some of its close relatives, B. anthracis has some activity outside of its vertebrate hosts. Here we show in the field that B. anthracis has significant interactions with a grass that could promote anthrax spore transmission to grazing hosts. Using a local, virulent strain of B. anthracis, we performed a field experiment in an enclosure within a grassland savanna. We found that B. anthracis increased the rate of establishment of a native grass (Enneapogon desvauxii) by 50% and that grass seeds exposed to blood reached heights that were 45% taller than controls. Further we detected significant effects of E. desvauxii, B. anthracis, and their interaction on soil bacterial taxa richness and community composition. We did not find any evidence for multiplication or increased longevity of B. anthracis in bulk soil associated with grass compared to controls. Instead interactions between B. anthracis and plants may result in increased host grazing and subsequently increased transmission to hosts. Citation: Ganz HH, Turner WC, Brodie EL, Kusters M, Shi Y, et al. (2014) Interactions between Bacillus anthracis and Plants May Promote Anthrax Transmission. PLoS Negl Trop Dis 8(6): e2903. doi:10.1371/journal.pntd.0002903 Editor: Joseph M. Vinetz, University of California San Diego School of Medicine, United States of America Received December 6, 2013; Accepted April 14, 2014; Published June 5, 2014 This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication. Funding: This project was funded by NIH Grant GM083863 to WMG. The PhyloChip laboratory work was performed at Lawrence Berkeley National Laboratory under contract number DE-AC02-05CH11231 with the Department of Energy, Office of Science. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: holly.h.ganz@gmail.com ¤a Current address: Genome Center and Department of Evolution and Ecology, University of California, Davis, Davis, California, United States of America ¤b Current address: Centre for Ecological and Evolutionary Synthesis, Department of Biology, University of Oslo, Oslo, Norway Introduction The causative agent of anthrax in humans, livestock and wildlife, Bacillus anthracis was long considered an obligate pathogen that is dormant in the environment [1,2]. This view was supported by early laboratory studies conducted by Minett and Dhanda [3] and Minnet [4] that suggest that B. anthracis is unable to compete with other soil bacteria. However, a growing number of laboratory studies indicate that, like some of its close relatives, B. anthracis may interact with other members of grassland communities, including plants [5], earthworms [6], flies [7], and soil amoeba [8]. Bacillus anthracis belongs to the closely related Bacillus cereus sensu lato, which contains both obligate and opportunistic animal pathogens, including B. cereus sensu stricto and Bacillus thuringiensis [9]. A common soil saprobiont [10], B. cereus causes opportunistic infections in humans [11] and lives symbiotically in invertebrate guts [12,13]. An insect pathogen commonly used as a pesticide in agriculture [14], B. thuringiensis also occurs in soil and can be found living symbiotically in caterpillar guts [9]. In his pioneering research, Pasteur proposed that earthworms vector B. anthracis from buried livestock carcasses [15,16]. Following up on these observations, Schuch and Fischetti [6] found that bacteriophages generate phenotypic changes in B. anthracis that enable it to persist as an endosymbiont in earthworms and to act as a saprobiont in soil and water. In a laboratory study, Saile and Koehler [5] demonstrated that B. anthracis germinates, survives as a saprobiont and undergoes horizontal gene transfer in the plant rhizosphere. Significantly, Dey et al. [8] showed that the virulent Ames strain of B. anthracis germinates and multiplies intracellularly within a free-living soil amoeba living in moist soils and that the pXO1 plasmid was essential for growth. Transmission of B. anthracis is environmentally mediated and such ecological interactions in the environment likely affect its transmission to subsequent hosts that contact and ingest spores while grazing [17]. In the late 19 th century, Louis Pasteur identified carcass sites as a key area for anthrax transmission [15]. At these locations, after a vertebrate host has succumbed to an anthrax infection, vegetative cells of B. anthracis are released (along with blood and other body fluids) into the environment and produce infectious spores capable of long-term survival [3,18–20]. The carcass also provides a pulse of nutrients into the soil, promoting plant growth [21–23] and supporting B. anthracis survival and replication [3,4]. In addition to nutrients, the inoculation of B. anthracis into carcass site soils may also promote PLOS Neglected Tropical Diseases | www.plosntds.org 1 June 2014 | Volume 8 | Issue 6 | e2903