Collaborative Research: MSB: Biological stoichiometry of microbes under severe P-limitation

Project: Research project

Description

Intellectual Merit: Understanding the mechanisms that link genomes to biomes is one of the most exciting challenges facing modern biology. In this project we test a set of ideas that connect environmental phosphorus (P) supplies to dynamics and structure of microbial communities via mechanisms associated with the structure and function of the genes that control ribosome biogenesis (the rrn genes and their regulatory domains). To test this Growth Rate Hypothesis (GRH), we will perform lab and field experiments involving microorganisms and microbial communities in springs and saline ponds in the Cuatro Cinegas basin (Mxico; CCB) where previous work has found both highly biodiverse microbial communities and severe in situ P-limitation. Lab studies using isolates of Bacillus from CCB will evaluate associations among growth rate, macromolecular composition (especially RNA allocation), and C:N:P stoichiometry and how these are generated by variations in the rrn genes (copy number, nature of rrn promoters). Carefully controlled chemostat experiments will evaluate how differences in microbial rrn genes or biomass N:P ratios translate to differences in competitive outcomes under nutrient supply conditions differing in N:P ratio or in the timing of P supply. Field studies at CCB will test the GRH in microbe-dominated saline ponds (salterns), the Pozas Rojas. A microcosm experiment will manipulate nutrient supply N:P ratio and evaluate response of the microbial community. We will also take advantage of a NASA-funded whole-ecosystem P fertilization that will take place at the Pozas Rojas in which 3 salterns will receive P fertilization while 3 others will be left unmanipulated as controls. In both the microcosms and the experimental pozas we will use a suite of molecular methods (clone libraries and ribotyping; qPCR; single-cell genotyping) to characterize the response of the microbial communities to these manipulations and relate these changes to aspects of the rrn genes of species that are most responsive to N:P ratio and P fertilization. Our project offers an opportunity for a transformative integration of genomics, evolutionary ecology, and biogeochemistry by spotlighting ribosomes as central not only to cellular growth but also to microbial evolution and to stoichiometric coupling of nutrient cycles in ecosystems. Broader Impacts: Our study will provide a comprehensive interdisciplinary and international experience for undergraduate, graduate, and postdoctoral students and thus provide training that is uniquely appropriate for students entering an increasingly integrated and globalized research environment. Furthermore, the bi-national nature of our project offers a special opportunity to attract and retain Hispanic American students to microbiological research; we will target underrepresented groups for these lab and field research opportunities. The spectacular desert springs of CCB provide a compelling platform for educational outreach. We will work with the Houston Museum of Natural Science to develop an ultra-high resolution educational video about microbial ecology and evolution at CCB for presentation at permanent and portable planetariums. Following up on successful showings during past visits to Cuatro Cinegas, we will deploy the new planetarium show both at Cuatro Cinegas and in schools in Houston and Phoenix. The video will be produced both in Spanish and English, providing a unique opportunity for enhanced science education for Hispanic American students who are an important demographic group in Arizona and Texas. Prior to distribution to other planetariums, the video will be premiered on YouTube together with a public live discussion using the Skype Public Chat feature. Finally, the CCB is a global biodiversity hot spot for both macro- and micro-biota and its conservation status has recently received high-profile attention in Mxico. Our data will contribute further to t

Description

Like most semesters, I currently have three undergraduate students in my lab. Ms. Nicole Nevarez and Ms. Sarah Schimpp are conducting independent research for their honors thesis. Mr. Alek Van Houghton was last years REU student who continues to work in my lab. I continuously encourage undergraduate participation in research through Arizona State University internal programs such as the School of Life Sciences Undergraduate Research (SOLUR) program, Barrett, The Honors College at ASU, NSF-funded Interdisciplinary Training for Undergraduates in Biological and Mathematical Sciences (UBM) and the ASU/NASA Space Grant. The undergraduate student for this REU project will be recruited primarily through these programs. Our effort within this NSF-funded project focuses on linking genome characteristics with life strategies of microorganisms. We are currently designing chemostat system to compete Bacillus species with different rrn operon copy number during growth in medium with different nitrogen:phosphorus (N:P) ratios. This competition experiment aims to determine how rrn operon copy number influences the biological stoichiometry of bacteria and their response to nutrient availability. This chemostat setup also provides a great platform for studying expression of rRNA genes in a plethora of Bacillus species growing under different nutrient regime. Differential contribution of each rRNA gene to the total cellular rRNA pool has been documented in model laboratory organisms such as Bacillus subtilis and Streptomyces coelicolor (Kim et al. 2008, Rosenberg et al. 2012). The variability in the expression of the different rRNA gene is suggested to be more apparent under low-phosphorus condition (Samarrai et al. 2011). Can bacteria with high rrn operon copy number down-regulate expression of some rrn operons when P becomes limiting? An undergraduate student can take on the task to further explore this phenomenon, building upon our goal to link genome to microbial life strategies. By using promoter specific primers, quantitative RT-PCR for the different rRNA operons can be performed on the continuous cultures exposed to different nutrient regime. The outcome of this project will help determine if the differential expression pattern of rRNA operons previously characterized in model laboratory organisms are universal or specific to exponential growth. Measuring rRNA operon expression of cultures growing under extremely P-limiting conditions will help determine the contribution of rRNA synthesis on the minimal P requirement for growth. Furthermore, the undergraduate student will have the opportunity to work with unique organisms because the collection of Bacillus strains was isolated from the biodiversity oasis, Cuatro Cinegas basin in Coahuila, Mexico (Perez-Gutierrez et al. 2012). These strains have rrn operon copy number that range from 4 to 9 and yet are highly resistant to long term P-limitation. In addition to the suggested project, the student will be encouraged to discuss any ideas he/she may have. I will also take into consideration any technique or experience the undergraduate student seeks to gain. I will assist in experimental design to ensure that any positive outcome from the project can contribute to a manuscript. The undergraduate student will gain laboratory experience in microbiology, molecular biology, biochemistry and potentially bioengineering. The recruitment and project timeline is summarized in Table 1. Recruitment will take place in January through March. The student will have a clear idea of the project by the middle of May and will start working in the lab in June. The student will meet with me and all lab members involved in the project weekly to discuss the project in detail. The undergraduate 4 of 8 student will also be expected to participate in our biweekly lab meeting and other lab activities. The student will be required to present his/her result at the end of the 12-week duration.

Description

The current project (DEB-0950179, Biological stoichiometry of microbes under severe P-limitation) aims to establish a link between the genome and life strategies of microorganisms, with a focus on life under unbalanced nutrient availability. Bacteria are known to have between 1 to 15 copies of ribosomal DNA genes (rrn) and preliminary data have found that this copy number to correlates with growth capacity and the capability to respond to changes in growth conditions (Condon et al. 1995, Klappenbach et al. 2000, Weider et al. 2005). Based on the Growth Rate Hypothesis (GRH) established by my research team (Elser et al. 2003), it is predicted that organisms with higher rRNA genes copy number will require more phosphorus (P) for growth and hence, will have lower biomass C:P and N:P ratios. On the other hand, organisms with fewer rRNA genes will have high C:P and N:P rations and will be more competitive under low P environment. The main site for this project is the Cuatro Cienegas basin in Coahuila, Mexico, where many of the ponds and streams have very low phosphorus content and high N:P ratios (Elser et al. 2005, Escalante et al. 2008). We are currently investigating the influence of changes in nutrient stoichiometry on the dynamics of the microbial communities in a P-limited pond, particularly populations with different life strategies as indicated by their genomic rrn copy number. Along with characterizing the microbial communities, we characterize the water and sediment chemistry of the pond throughout the field season to elucidate the nutrient dynamics in these P-limited pond system. This project will be a great opportunity for an undergraduate student to participate in an interdisciplinary, international collaborative experiment that involves both field and laboratory components and that engages microbiology, ecology and biogeochemistry perspectives. We request funding to support a summer REU student to participate in the project. There will be many significant questions that can be asked because important aspects of nutrient cycling and microbial ecology at CCB remain largely unknown. We have an in situ nutrient amendment experiment setup in addition to sampling sites with different nutrient characteristics the student can take access. The student will be working with a team of scientists with diverse backgrounds, such as limnology, hydrology, microbiology, and genomics, and will have the opportunity to interact with our collaborators from the Insituto de Ecologia in the Universidad Nacional Autonoma de Mexico and CINESTAV.
StatusFinished
Effective start/end date9/1/108/31/16

Funding

  • NSF: Directorate for Biological Sciences (BIO): $975,749.00

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cooperative research
stoichiometry
college students
students
microorganisms
operon
ribosomal RNA
microbial communities
nutrients
genes
organisms
Bacillus (bacteria)
phosphorus
Hispanic Americans
microbiology
gene dosage
genome
biogeochemistry
microbial ecology
Mexico