In response to growing concerns regarding the sustainability of human civilization, rising effects of climate change, and worldwide resource inequalities , researchers have developed a variety environmental impact assessment techniques to reduce and manage the environmental footprint of human-made systems. One technique in particular, Life Cycle Assessment , is used to investigate the environmental impacts of industrial products and production systems. Major issues such as water pollution, greenhouse gas emissions, and eutrophication tend to be a result of resource intensive processes such as the extraction of raw materials, consumption of materials and energy, transformation of one material into another, and the manufacture of products. LCA involves the quantification of these resource flows in a system to calculate the environmental impacts that the system incurs. Models of agricultural products and production systems are commonly created using LCA. Farmers, along with environmental analysts, can conduct LCAs to quantify environmental impacts, and subsequently make improvements in the farming processes to reduce undesired impacts. Interest in tailoring LCA for agricultural systems has led to the identification of many substantive problems at the intersection of agricultural modeling and environmental impact assessment. This has led to the development of many new modeling techniques, creation of software tools, and interdisciplinary collaborations. As a result, myriads of models are produced to address these problems. The creation of LCA models is often money and time intensive, requiring input from many stakeholders, large quantities of data to be collected, and analyses to be conducted by environmental experts trained in LCA methods and tools.

The resulting LCA models are instrumental to understanding and improving the environmental performance of the agricultural system of interest. These LCA models are therefore highly place and time specific: they often represent a single product system in a particular geographic region using data collected during a certain period of time. By the time the environmental impacts are calculated and translated into meaningful recommendations for a change in the real world system, the models may be out of sync with the real world. This is not conducive to making recommendations regarding the assessment, design, or optimization of other similar systems.LCA is a technique used to assess the environmental impacts of products and the processes by which they are constructed. The system of interest typically involves the development of a particular product: a farm where tomatoes are grown, a factory where shoes are made, or a wastewater treatment plant where clean water is produced. There exist numerous LCA variants. Each has a different purpose, system modeling technique, domain applicability, and data requirements. Standardization: The generic LCA process is governed by a suite of standards: ISO 14040 details the principles and overall framework, and ISO 14044 specifies the requirements and guidelines for conducting an LCA. These standards are part of a greater family, ISO 14000, regarding Environmental Management, that provide guidance on environmental assessment, performance, and responsibility. ISO standards are reviewed every five years: the most recent versions of ISO 14040 and ISO 14044 were published in 2006 and reviewed in 2010. These reviews aim to maintain the capacity of LCA models to represent systems resulting from advances in the real world. Not all LCA studies are ISO compliant.

However, compliance tends to occur in government funded LCAs, when analysts intend for their data to be incorporated into national LCA databases , some LCA research, and certain LCA studies conducted for companies and organizations. LCA standards are written in a domain agnostic manner, enabling for LCAs conducted in any domain to potentially be ISO compliant. For instance, an LCA of a beef producing agricultural system, plastics and metals recycling, and a printer cartridge can all be ISO compliant, as long as they adhere to the requirements and guidelines laid out in ISO 14040. For the rest of this section, I will briefly overview the generic LCA process. Figure 3.1 outlines the main phases of an LCA: Goal Definition and Scope, Inventory Analysis, Impact Assessment, and Interpretation. There are varying techniques for conducting both the overall LCA, as well as individual phases.Define the purpose of the LCA study: One is describing a problem in the real world: what environmental impacts are incurred by producing specific quantities of a product, how great are the impacts , and what are the causes? Therefore, the goal is to assess the environmental footprint of the system. Alternatively, the goal could be to compare the environmental performance of competing systems. Depending on the problem domain, reasons for the analysis being conducted, and the familiarity of the analyst with the domain, the analyst may, from experience, know which environmental impacts can and should be calculated. There exist environmental impact categories that can be selected as specific areas of interest, but they are not always selected during Phase One, as they are only used in Phase Three when impact calculation occurs. Define system boundaries and scope: The production systems and stages to be included in the system of interest, and therefore the assessment, must be selected. For example, a cradle-to-grave LCA includes everything that went into the manufacturing of the final product, starting from the extraction of raw materials, to the disposal of the product after use. It takes into account the full life cycle of a product. This also requires specifying the time scale of the analysis: for example, the life cycle of a solar panel includes 25 years for the operational/use phase, in addition to the time spent in the manufacture and disposal of the product. In contrast to the solar panel, the use phase of a cup of coffee depends on how it is made and consumed. The undercradle-to-grave LCA stages for which raw materials and energy are accounted for are: raw material acquisition,transportation, grow racks manufacturing processes, the use, reuse, and maintenance of the product, and finally, recycling and waste management. Alternatively, a cradle-to-gate LCA, accounts for everything up until the product is ready to be transported to the customer. This method of scoping an LCA study works particularly well for analyzing production systems for physical products with a well-defined life cycle, clear-cut inputs and outputs, and a limited set of uses. More recently, it has been suggested that life cycle considerations should extend from “cradle-to-cradle”, taking into account both the production, but eventual disposal, and potential reuse of physical products. A decision must also be made as to how many levels of indirection from the core system are to be included in the study. Attributional LCA studies focus on accounting only the immediate and direct impacts of the system of interest. This has been the traditional approach to LCA, where one quantifies the effects of the system as it currently is.The Life Cycle Inventory Analysis phase involves gathering a list of all the processes occurring within the system, along with the material and energy resources utilized, useful products created, and waste produced. In this phase, the system of interest is decomposed into a set of discrete processes, with inputs, outputs, and quantifiable flows of material and energy resources through each process. A comprehensive LCA relies on the ability of the analyst to find the environmental impact of each process within each subsystem during the LCI Analysis phase. This approach is known as process LCA. Alternatively, in an input-output LCA, the system is treated as a whole, with aggregate inputs and outputs into the system. One of the most popular variants is the economic inputoutput LCA: it uses aggregate economic inputs and outputs into the system of interest to estimate environmental impacts.

It is an application of a popular economic modeling approach that is used to assess the economic performance of a system. This phase has two deliverables. The first is a flow diagram, a high level overview of the main processes or components in the system. The second is the Life Cycle Inventory itself. The LCI is an all-encompassing inventory of inputs, outputs, and processes within the system of interest. Process and input-output LCA represent two different approaches to abstracting the system of interest into a model with quantitative and qualitative attributes. The level of detail in an LCA study also varies based on how much time, effort, and money the analyst has to put in, how much data is actually available or collectable, and what the results will be used for. If the level of detail is low, and the system of interest has been simplified in its representation, the study is labeled as a streamline LCA. Otherwise, every process of every stage in the scope of the system of interest must be represented in the LCI. The LCA ISO 14044 standard specifies mandatory elements within this phase: the selection of impact categories, category indicators, and characterization models; classification ; and characterization . This produces a set of Category Indicator Results, which collectively are known as the LCIA Profile, and where each result represents the actual environmental impacts of the system of interest by associating each of the processes in the LCI with the impact categories affected by the process. Impact categories may include: eutrophication, climate change, global warming potential , land use, water use. There has also been growing interest in conducting more holistic “sustainability” LCAs that include assessment with respect to economic, social, and environmental sustainability, as described by Goodland. The functional unit, the processes represented in the LCI, and selected impact categories are brought together in this phase to produce system level calculations of environmental impacts. Conclusions take the form of: X system causes Y quantity of Z impact, due to [1:n] processes.The interpretation phase of the LCA involves analysis of the results after each phase, validation to ensure that the LCA is proceeding in compliance with the originally stated method, and creation of reports regarding progress and findings. According to the ISO 14040 standard, this phase involves the reflection on the results of the Life Cycle Inventor Analysisand the Life Cycle Impact Assessment, as well as appropriate communication about findings. While there are many interpretation activities within this phase, they are usually specific to the types of environmental impact that are being calculated in a particular life cycle assessment case. Interpretation involves the production of a variety of charts and reports communicating intermediate results and checking in with the stakeholders of the study to ensure correctness and completeness. This is not actually the last phase of LCA, but instead, it occurs after each of the other phases. It allows for constant reflection on the relationships between results in different phases as they are produced.The studies analyzed in this section were constrained to those whose primary focus was on the United States, European or Australian agricultural systems, due to two reasons. First, a vast majority of published LCA literature is in English, and comes from these three regions. Second, while LCA is popular in other parts of the world, such publications are not always easily accessible, or data sources traceable, as they may be using local databases or native language resources. Figure 3.2 describes the top ten most produced agricultural commodities globally, as reported by the Food and Agriculture Organization of the United Nations Statistics Division.

These anecdotes highlight the growing social issue of homelessness in Long Beach, and bring into question whether physical barriers and policies effectively protect UA sites, or simply exclude community members. UA leaders also struggled with language barriers, which unintentionally excluded or created misunderstandings between community members who were not fluent in English. Some UA leaders had to rely on gardeners’ English-speaking children to act as interpreters. While volunteering at a community garden, I witnessed how language differences made it difficult to communicate important policies. While working with gardeners to weed a pathway, we discovered that the “weeds” were vegetables intentionally planted by a Korean gardener, who appeared very upset. Due to the language barrier, we could not identify the seedlings, nor was the gardener aware of the policy that common areas should not be used for planting. In the end, we replanted the dug-up vegetables, and the gardener shared a plate of fried perilla leaves with us .This section expands on Chapter 3, which partially addressed my research questions on how communities create, maintain, and engage in UA. To further answer questions regarding how UA fosters skills and knowledge and addresses health inequities, I specifically analyzed my findings in the context of the CCW model and SDOH domains.The six forms of capital from Yosso’s CCW model were represented in several aspects of UA. Aspirational capital allowed community members to envision the possibility of converting vacant lots into gardens and farms.

Resistant, navigational, and social capital was integral for actualizing these possibilities into reality. Through resistant capital, UA leaders and community members worked to actively change their neighborhoods despite challenges. LBO’s Zaferia Junction Community Garden is an example of this, as it was constructed on land previously used by the Pacific Electric Railway, demolished during the Termino Avenue Drain Project, then rebuilt again . Through navigational capital, UA leaders acquired permits from the city and obtained permission from private landowners to create UA sites. Community members leveraged social capital to build and maintain these sites collectively. In turn, UA sites boosted social capital by creating communal spaces for people to gather and host events. Familial capital was also important for preserving agricultural skills and knowledge through generations of family. UA leaders recognized the linguistic capital of others involved in UA , but it was the least represented form of capital in this study.UA sites increased social and community support by creating a space for community members to connect, through the experience of growing food, exchanging food at crop swaps and potlucks, and engaging in educational opportunities. In addition to the educational lessons provided by Ground Education’s school gardens, community gardens and urban farms offered educational workshops for children, adolescents, and adults. UA also provided informal opportunities for families and community members to share knowledge and skills.

Supporting economic stability, UA sites created paid positions for community members. Though most sites could only afford to hire one person or a small team of staff, UA yielded career development opportunities for volunteers and interns to gain experience. According to interviewees, UA did not directly increase access to health care services or health insurance. However, UA sites were healthy environments for community members to grow fruits and vegetables, increase their physical activity, and cope with stress. These findings support the use of UA for addressing SDOH, to promote health and potentially prevent disease. The following chapter will summarize the main takeaways of this research and provide recommendations for organizations, researchers, and policymakers.In summary, this dissertation provided an overview of urban agriculture sites in Long Beach and described how communities create, maintain, and engage in UA. Furthermore, this research examined how UA relates to community cultural wealth and the social determinants of health . These findings supported previous research on the social and environmental benefits of gardening , and uniquely focused on multiple types of UA sites, including community gardens, school gardens, and urban farms. According to interviews and field observations, UA increased access to healthy food and green spaces, particularly for Long Beach’s communities of color, which are disproportionately impacted by health inequities . This research supports emerging evidence that UA contributes significant value to public health and social systems . UA provided communities with fresh produce, improved mental health, promoted outdoor activity, enhanced neighborhoods, and created educational opportunities. Interestingly, Long Beach UA sites often began as the vision of one person or a small group of community members. Individuals combined their skills and knowledge by forming volunteer groups, organizations, and businesses. However, as I discovered while researching nonoperational gardens and farms, landowners held the most control over the longevity of UA sites. Public policies that encourage agricultural land use are critical for the long-term success of UA, particularly to protect UA land from being sold. To summarize study findings, I designed Figure 32 based on Lee et al.’s socioecological model of agricultural safety and health interventions.The UA sites in this study relied heavily on partnerships between community organizations, volunteers, the City of Long Beach, and private landowners. Most community gardens and school gardens were managed by Long Beach Organic and Ground Education respectively, which are both nonprofit organizations. Long Beach Community Garden , the largest community garden, cannabis commercial was managed by the Long Beach Community Garden Association, a not-for-profit organization. Nonprofits qualify as tax exempt in exchange for providing a public benefit . In contrast, not-for-profits can solely operate for the goals of their members, not the public . Both nonprofit and not-for-profit organizations must use their funding for their mission and operations, rather than the profit of an owner. Regardless of the type of organization, this research found that it was important for community members to organize formally. Organized groups could mobilize their skills and social capital to advocate for the creation of UA sites, secure funding, hire workers, and coordinate volunteers. Interviewees emphasized that volunteers from the community or organizations were essential for constructing and maintaining UA sites. In particular, UA sites repeatedly described volunteer efforts from the Conservation Corps of Long Beach, Boy Scouts of America, and UC Master Gardener Program. UA sites can also collaborate with partners to develop programming. Long Beach gardeners and farmers created social events, developed educational workshops, and shared their harvest with family, neighbors, and the community.

UA leaders advanced food justice, accomplishing Heynen et al.’s recommendation of using UA to connect food sovereignty and food security. Long Beach UA sites empowered communities to grow food, exchange knowledge, and gain agricultural skills. UA sites increased access to fruits and vegetables for those interested in growing and purchasing food. Additionally, UA sites served local community members and under served populations through food donation programs. For example, the Peace Garden distributed produce at monthly events as part of their outreach program. Peace Garden staff members grew food around the perimeter of the garden, which is also open for community members to harvest on weekdays. Similarly, LBCG and LBO gardens designated areas to grow food for local pantries and organizations. Since 2020, LBO has donated 160,000 pounds of organic produce to California State Long Beach University’s student pantry UA leaders should consider designing sites to be accessible and inclusive. Many of Long Beach’s UA sites accommodated different populations, such as those with disabilities, seniors, and veterans. LBCG was built with paved roads, which allows gardeners with mobility aids to access their plot more easily. LBO designed their gardens with pathways large enough to accommodate wheelchairs, based on standards issued by the Americans with Disabilities Act . The Veterans Affairs Patient Garden, which serves veterans staying long-term at the VA Hospital, was accessible to those with wheelchairs and walkers. The Magic Garden was originally founded by veterans. This garden has raised beds, enabling elderly gardeners and gardeners with disabilities to tend their crops without crouching or bending over. In addition to considering the physical accessibility of gardens, UA leaders were interested in increasing language accessibility. For example, the Peace Garden translated informational materials on how to register for garden plots into English, Spanish and Khmer. The Peace Garden was managed by Long Beach’s health department. Their staff worked with a public affairs team to distribute outreach materials at community resource fairs and online via social media. By offering outreach materials in multiple languages, UA sites can attract more diverse community members.While recruiting research participants, I found that many UA leaders did not immediately respond to emails and phone calls to schedule an interview. In some cases, even UA leaders who I had met in person and verbally expressed interest in participating, did not respond. This may be explained by a lack of capacity, as UA leaders were typically responsible for multiple duties at their site. Most UA sites were managed by a single person, with occasional assistance from board members, volunteers, and/or interns. Therefore, it was important for me to visit sites, participate in events, and volunteer. Volunteering as part of fieldwork is a participatory research practice that can allow researchers to contribute to a community, instead of passively observing . Through volunteer work, I earned the trust of UA leaders and was able to work alongside community members. For future research on UA sites, there is a need for interdisciplinary collaboration between social and environmental scientists, as well as medical professionals who can analyze biometric data. Longitudinal studies could investigate the quantitative impacts of UA on food security and disease risk. Further exploration is needed to understand the relationships between UA participation and obesity, diabetes, asthma, and mental health disorders. Additionally, researchers should study UA’s potential to improve air quality by reducing greenhouse gases and producing oxygen . UA may also reduce flooding in urban areas, as permeable soil can retain and absorb rainwater .Industrial agriculture has accelerated the loss of soil organic carbon, an indicator of soil health and a key part of climate regulation . Scholars should consider implementing the traditional ecological knowledge of indigenous people, such as the Tongva. Practices linked to TEK, such as the preservation of oak groves, can be applied today to promote biodiversity, prevent droughts, decrease carbon emissions, and ensure biodiversity . When designing future studies, researchers should include more diverse perspectives of UA participants and beneficiaries, such as family members and neighbors. A limitation of this research is that study participants did not reflect the demographics of the community at large. Out of 19 interviewees, 68% identified as White, yet over two-thirds of Long Beach’s population are people of color . This may be an indication that more Whites held UA leadership roles, compared to the diversity of gardeners, volunteers, and other participants. The discrepancy in participant demographics could also be explained by distrust, resulting from researchers’ historical exploitation of people of color. Examples of this include the Tuskegee study, which led to the preventable deaths of African American men, and a genetic research study of Havasupai tribe members, who donated blood samples for a diabetes study. The tribe members never received results of the study, yet researchers continued to use the blood samples for research .

Environmental justice is a movement that seeks to address health inequities, specifically by targeting the inequitable distribution of environmental hazards and benefits . This movement was a direct response to the injustices faced by communities disproportionately exposed to environmental risks, due to their race, color, national origin, or income . Though low-income communities of color typically lack the resources to politically and legally oppose or hold accountable polluting companies in their neighborhoods, they have historically made several efforts to resist environmental injustice through protest and social change .In relation to CCW, social, resistant, and navigational capital are important for communities to organize against injustice. As described in Chapter 1, Native Americans have experienced the loss of land, water, minerals, and other raw materials since the appearance of non-Native explorers and settlers in the late 15th century . The intellectual and legal traditions of Native Americans and other Indigenous peoples, such as collective systems that encourage sharing resources, share much in common with modern principles of environmental justice . The First National People of Color Environmental Leadership Summit, a multinational movement “of all peoples of color to fight the destruction and taking of [their] lands and communities,” drafted and created 17 principles of Environmental Justice . These principles affirmed the right of all species to be free from ecological destruction, and called for public policies free of discrimination, ethical uses of land, education of present and future generations on social and environmental issues, and the opposition of exploitation from multinational corporations and militaries.

Several communities across the United States reported benefits from UA, which can transform the built environment while creating positive social change . In New York, community activists took the initiative to build gardens on lots left empty by the city council. They were “squatted by local community activists” to grow flowers and vegetables in an otherwise neglected site, once occupied by gangs and drug dealers . In a study of 17 community gardens in Illinois, Lanier et al. found that the most frequently reported benefits were learning how to build and tend a garden, giving back to the community, promoting healthy living, increasing physical activity through gardening, and fostering new friendships between volunteers and the population served . In Bakersfield, California, graduate students, community members, and local agencies collaborated to transform a low socioeconomic neighborhood’s vacant lot into a garden. The community garden improved residents’ eating habits, increased property value, and “fostered a tighter-knit, stronger community” . Ron Finley of South-Central Los Angeles, a food desert where healthy food options were limited, began planting vegetables in neighborhood parkways. After being cited by the City of Los Angeles for gardening without a permit, he petitioned with fellow activists to gain the right to grow food in their community . In San Diego, the Ocean View Growing Grounds community garden was developed in 2012 from a privately owned vacant lot, which was used as parking and storage for an automotive repair facility. OVGG, now equipped with a kitchen and amphitheater, grows a variety of crops including figs, limes, and strawberries.

Cooper et al. note the risk of converting spaces for urban gardening, as they found detectable levels of lead in the leaves of fruit trees and arsenic in leafy green crops grown directly in the ground. However, edible fruit tissues did not accumulate heavy metals and arsenic, and crops grown in raised soil beds had no detectable contamination. The study recommended soil and food sample testing but concluded that UA can minimize health risks by providing a source of fruits and vegetables, as well as a space for cultural exchange and social connections in the community .Research suggests that UA can influence health by improving the environment, increasing access to healthy food, and providing a space for educational workshops and classes in nutrition education and physical activity . In general, UA increases access to green space. Residents with access to green space have lower asthma prevalence, reduced mortality from circulatory disease, and greater longevity . Studies suggest that access to green space helps urban dwellers increase their access to physical activity, cope with stress, ease symptoms of attention-deficit/hyperactivity disorder and depression, and reduce aggression and crime . Spaces with trees remove air pollution, while also providing shade and preventing heat-related illnesses . Additionally, green space in the form of gardens and urban farms can increase access to affordable food . UA can generate economic benefits by providing an affordable source of fruits and vegetables, promoting entrepreneurship, and alleviating poverty and social exclusion . Ferris et al. documented the efforts of St. Mary’s Youth Farm, which offered affordable organic produce, employment , and training opportunities to residents of a San Francisco housing project. A 2004 study of Latino community gardens in New York City found that an investment of $5 to $10 in plants for a garden plot can yield $500 to $700 worth of fruits and vegetables, which some participants sold to markets or restaurants . In 2017, Saha and Eckelman conducted a geospatial analysis of potential UA spaces at ground level and rooftops in Boston and found that UA could yield enough fruits and vegetables for the city’s population. UA sites can also provide employment opportunities, which increase job and interpersonal skills such as respectfulness and teamwork . There is evidence that UA provides educational opportunities, which can also influence health habits . A study of 845 residents from Flint, Michigan found that respondents with a household member who participated in a community garden consumed fruits and vegetables 1.4 more times per day, compared to those without household participation in a community garden. In addition, households with a community gardener were significantly more likely to have participated in a neighborhood cleanup or beautification project . According to a California study of six community garden programs in the cities of Berkeley, Escondido, Loma Linda, Oceanside, San Bernardino, and West Hollywood, community gardens increased participants’ average fruit and vegetable consumption and provided space for nutrition education and physical activity workshops . Through funding from California Healthy Cities and Communities, these cities established garden programs to serve residents, children at schools and day care centers, or seniors. Programs offered classes in gardening, exercise, vertical racks and cooking. Twiss et al. found that staff and gardeners also had opportunities to build their skills in leadership, community organizing, program planning, and cultural competency .

Multiple studies documented UA’s influence on children’s knowledge, attitudes, and behaviors related to nutrition . In 2008 and 2009, Growing Healthy Kids established three community gardens in Carrboro, North Carolina . This program hosted weekly gardening sessions, cooking and nutrition workshops, and family events. According to the evaluation report, participating families reported a 146% increase in fruit and vegetable availability and increases in fruit and vegetable consumption. At the end of GHK, 17% of obese or overweight children had improved their BMI classification. According to parental reports, there was an increase of 146% in the availability of fruits and vegetables and a 28% increase children’s in fruit and vegetable intake. Following GHK’s implementation, Carrboro changed its planning policies to include community gardens at designated parks, require recreation spaces in private developments, and periodically review plans with the GHK program team to sustain current garden sites . Youth gardening programs were found to improve access and consumption of healthy foods, promote development through social relationships and respect for others, and increase science achievement and environmental attitudes . One example of such a program is AmeriCorps Farm to School , which was established in Wisconsin in 2008 . The F2S program evaluation included nine school sites, with 1,191 students. F2S incorporated locally grown ingredients into school meals, and consisted of activities such as gardening, field trips, and nutrition education. F2S schools served more fruits and vegetables to students, and students from schools with one year or more of F2S demonstrated increased knowledge and improved attitudes on food, nutrition and agriculture. . Another program, Sustainability via Active Garden Education , was developed as a community-based participatory research intervention, to increase physical activity and produce consumption among children aged 3 to 5 years old at early childhood education centers in Phoenix, Arizona and Houston, Texas. In addition to garden maintenance and taste tests, children participated in interactive learning activities, such as songs and games. Though participants did not report significant changes in fruitand vegetable consumption, participants had a higher amount of physical activity post intervention . Table 2 provides an overview of the conceptual framework for this dissertation, which aims to further explore this concept, through a case study of community-led UA in Long Beach, California.Chapter 3 will partially answer my first two research questions regarding how communities create, maintain, and engage in UA, and what forms of skills and knowledge do they draw on to create UA sites. This chapter provides context for what UA sites in Long Beach look like, where they are, when they were formed, what they grow, and how they are managed. From May to January 2024, I visited 26 active UA sites in Long Beach, including 17 community gardens, five school gardens, and four urban farms. During these field visits, I also identified 14 gardens and farms that are either no longer operational or under new ownership. In this chapter, I share photographs from my field visits and discuss descriptive findings from UA sites: location, year when the site was built, who owns the property, who manages the property, amenities, and food grown. Long Beach is in USDA Plant Hardiness Zone 10b, which means that extreme winter temperatures range from 35 to 40 degrees Fahrenheit . From my observations, all UA sites planted seasonal crops that thrive in the warm Southern California climate, such as the following fruits and vegetables: arugula, beans, beets, bok choy, broccoli, carrots, chard, collard greens, cucumber, eggplant, kale, lettuce, mustard greens, peas, peppers, radishes, strawberries, tomatoes, zucchini and other types of squash, and wheat. Due to the similarity of crops, this chapter will only discuss plants that were unique to a site or specifically mentioned by gardeners. It is important to note that I intentionally wrote about UA sites in the present tense, based on the concept of the ethnographic present . Though UA sites will inevitably change over time, I chose to distinguish operational sites from nonoperational sites .The Veterans Affairs Hospital Patient Garden is a two-acre site that has served long-term patients since the 1960s. The Patient Garden is behind the main hospital, the Tibor Rubin VA Medical Center , and next to the Fisher House, which provides temporary accommodations at no cost for families and caregivers of Veterans and Service members receiving treatment through a VA Medical Facility. Long Beach’s VA Hospital was established in 1941 as a Naval hospital and became part of the Veterans Administration in 1950 . The hospital fulfills the health care needs of over 50,000 veterans as part of the VA Long Beach Healthcare System .

We specifically avoid these issues by considering multiple, independent, neutral regions from the autosomes. In sum, the ,1000-fold increase in human population size that has taken place over the last 10 kya is unlikely to be detectable with current resequencing data. The finding that autosomal resequencing data from all sub-Saharan African populations so far tested contain a signal of exponential size increase beginning in the Late Stone Age is concordant with archaeological data showing intensification in the number of LSA sites on the African landscape, an increased abundance of blade-based lithic technologies, and enhanced long-distance exchange after 50 Kya. Interestingly, there is mounting evidence that many of the individual elements of complex behavior first appear earlier in the Middle Stone age, 70–100 Kya. This suggests that the demographic effects manifest in these indicators of modern culture were felt only sporadically in the MSA, and that they did not become the general condition until the LSA, coincident with the significant population growth that is detectable in the autosomes of contemporary sub-Saharan Africans.Antimicrobial therapy is critical for the treatment of bacterial infections in veterinary medicine; however, resistance to these treatments has been increasing for decades and, as a result, there are concerns about the efficacy of antimicrobials in food-producing animals. It has been well documented that use of antimicrobial drugs is associated with increases in antimicrobial resistant bacteria , but increases in antimicrobial resistance genes can also develop in the absence of antimicrobial drug use .

There is consensus that applying pressure on a population of bacteria through antimicrobials will enrich ARGs within that population ; however, AMR cannot be attributed to exposure to antimicrobials alone . Although Enterococcus spp. are part of the normal flora in the bovine gastrointestinal tract, there are reports of Enterococcus spp. as a causative agent of mastitis in cattle and diarrhea in neonatal calves . Enterococcus has, however, been primarily used as a sentinel and potential source of AMR genes for other Gram-positive pathogens. Enterococcus spp. are known to be intrinsically resistant to many antibiotics and can develop and confer AMR status to other pathogens . Thus, Enterococcus spp. have the potential to cause disease and serve as sentinels for the status of AMR pathogens in an environment. E. coli is another component of the normal flora of the bovine enteric system with many strains that have varying pathogenicity. Various E. coli strains can cause mastitis or metritis in cows as well as neonatal diarrhea or septicemia in calves . E. coli has also been used as an indicator of AMR prevalence in fecal bacteria and a potential source for transmission of ARGs to other Gram-negative organisms , as it acquires resistance easily and can inhabit many types of animals . California is an important contributor to the U.S. beef industry, with approximately 590,000 beef cows and contributing $3.4 billion in cash receipts from total cattle and calf sales in 2015 . In beef cow-calf operations in the U.S., calves are born and stay at the same location with their dam usually until 6–8months of age, at which point they are weaned, removed from the dam, and are often placed in a group with animals of approximately the same age and/or size.

At this stage of the beef production cycle, there are many sectors that can involve the movement and mixing of animals. Some calves may be moved directly from cow-calf operations to feedlots after weaning until they reach slaughter weight. Alternatively, if there is high forage availability, they may be moved temporarily to a stocker facility before ultimately finishing at a feedlot facility. The time spent in each sector, size of group, and management of the animals are highly variable and depend on many factors including geographic location, producer goals, and access to pasture and/or facilities. Rates for multi-drug resistance have previously been shown to be higher in California cattle from various types of beef production systems when compared to nearby states, Washington and Oregon . Additionally, many AMR studies thus far have focused on feedlot, stocker, and calf ranch operations, where there may be increased selection pressure on bacterial populations through antimicrobial therapy from treatment and/or prevention of disease that develops likely as a result of mixing of animals, transport, and stress . A 2010 study found that the highest proportion of MDR E. coli isolates originated from calf ranches, followed by feedlots, while the least MDR was found in isolates from adult beef cows . Prophylactic or metaphylactic use of antibiotics and occurrence of disease requiring antibiotic treatment is less common in cow-calf operations than other operation types in the beef production chain . There are far fewer studies investigating the levels of AMR that exist in cow-calf operations and not yet one that exclusively investigated the levels of AMR that exist on cow-calf operations in California. Nevertheless, characterization of AMR in cow-calf operations is essential for evaluating and understanding the contribution of the cow-calf sector to AMR in the beef industry, as well as whether specific management and antimicrobial use patterns may be associated with AMR during this production stage before the calves are moved to feedlots where higher selection pressures exist. Previous studies in cow-calf herds have indicated that management and operation dependent factors can influence the presence of AMR in a group. Specifically, season of collection of samples for testing , age of animal , and intensity of operation  have all been shown to be associated with increased AMR detection in beef operations. As part of the ongoing effort on surveillance for AMR, the objective of this cross-sectional study conducted on cow-calf operations in northern California was to assess prevalence of AMR in E. coli and Enterococcus spp. in beef cattle of different life stages, pasture types, and antimicrobial drug exposure on a herd and individual level. The hypothesis was that the prevalence of AMR in fecal samples varies according to the age of the animal as well as the drugs commonly used in the treatment of sick animals on those operations. The results of this study may serve as reference for future studies on the prevalence of AMR genes in the population of cow-calf operations in California and lead to a better understanding of risk factors for shedding of fecal pathogens carrying ARGs. Further surveillance, risk assessment, and interpretation of these results will help to derive more informed decisions and directions for combatting AMR in the future.A convenience sample of beef cow-calf operations in northern California was enrolled in this study either through the network of University of California Cooperative Extension livestock advisors or as clients of the Veterinary Medical Teaching Hospital at the University of California, Davis.

Enrollment criteria included farms with a geographic location in northern California and primary production sector as beef cow-calf. No restrictions were placed on the type of operation , herd size, grazing practices, breed of beef cattle, planting racks or previous antimicrobial use. All experimental protocols regarding animal use were approved by the Institutional Animal Care and Use Committee at the University of California, Davis.Fecal samples were collected between June 2019 and August 2020 from cows and/or calves on each farm as a convenience sample, by either the herd veterinarian, extension veterinarian, or cooperative extension livestock advisor. Fecal samples were collected from a combination of calves aged between 1week to 1year and adult cows aged between 2 and 10 years either from the rectum or from freshly voided manure after the animal was observed to defecate. Number and life stage of animal samples was based on number of animals available for sampling and that could be observed defecating within an hour of observation at the time of farm visit with the goal of sampling 5 cows and 5 calves per farm. Individual animal identifier, age, life stage , and breed were recorded when available.Fecal samples were collected during a single visit to each farm. Individual disposable gloves were used for collection of each sample and samples were stored in individual 15 mL polypropylene sample tubes. Rectal samples were collected on 8 farms from the recto-anal junction with individual rectal palpation sleeves. Pasture samples were collected as fresh feces via gloved hand from the top and center of a freshly voided fecal pat, where the individual animal was observed to defecate. Samples were transported on ice to the laboratory at the University of California, Davis where they were kept refrigerated at 4°C if culture could be performed within 48 h or stored at −80°C in tryptic soy broth with 25% glycerol.At time of fecal sample collection, an in-person survey regarding management and production practices was conducted. Information was collected on herd size, breed, certification status , whether any farm personnel were Beef Quality Assurance certified , type of pasture , whether the farm incorporated feeding of byproducts, water trough cleaning practices, existence of a current veterinarian-client-patient relationship, and whether the farm had submitted samples in the previous 12months to a diagnostic laboratory. The survey also included detailed questions regarding antibiotic practices on the farm, specifically use of antibiotics in feed, use of intramammary antibiotics, use of injectable antibiotics, practices related to injectable antibiotics including indication for treatment, method for determining treatment duration and dosage, information recorded regarding treatment, and specific antibiotic used in each method listed above. Treatment with antimicrobials in the past 6months before sample collection were recorded for all sampled animals based on ranch records, markings on treated animals, or rancher’s recollection of treatments.Selective growth media, E. coli Chromoselect Agar B and Rapid Enterococci Chromoselect Agar, following manufacturer guidelines , were used for culture and isolation of the respective bacterial types as previously described . Briefly, each fecal sample was streaked on the respective selective media using sterile cotton tipped applicators and incubated at 44°C or 35°C for 24h. Both E. coli and Enterococcus colonies were identified by characteristic blue green colony types on the Chromoselect plates. Two discrete colonies of each bacterial type were selected and purified on 5% sheep blood agar plates . The pure colonies were stored in tryptic soy broth with 25% glycerol at −80°C until all farm sampling was complete.After initial culture, a total of 698 bacterial isolates were stored. From these, 528 bacterial isolates were selected for antimicrobial susceptibility testing . Exclusion criteria included isolates from a farm where the rectal sleeve was accidentally not changed between samplings, all fecal samples which did not yield at least 2 identifiable isolates for each bacterial type after two culture attempts, and any samples with missing or unknown treatment information. Of the 528 selected, 482 were selected for antimicrobial susceptibility testing. See Figures 1A,B for flow charts of the isolate selection process for E. coli and Enterococcus isolates, respectively.Antimicrobial susceptibility testing was conducted by the broth microdilution method using Sensititre system against a panel of 19 antibiotics on a commercially available BOPO7F Vet Antimicrobial Susceptibility Testing Plate .Quality control steps included checking for bacterial growth and colony purity by plating 1μL of the inoculated MuellerHinton broth on TSA with 5% sheep blood.

The bacteria were collected from the first filter unit and divided evenly into three aliquots and recombined with either: sterile 10mM MgCl2 buffer ; 10mM MgCl2 containing the phage fraction from the 100K MWCO unit ; or an equal volume of the leaf wash filtrate from the 100K MWCO unit , which should represent another phage-depleted inoculum, but includes any molecules or particles smaller than 100 kDa in the sample.For field experiment 1, we prepared the bacteria only and bacteria plus 0.22µm flow through fractions, but we did not perform the third separation of the phage fraction from the rest of the leaf wash using the 100K MWCO centrifugation unit . After observing the significant effect that the 0.22µm fraction had on bacterial abundance, we then sought to dissect the effects of phage particles away from that of other small molecules and chemicals that were also removed from the leaved by sonication. In field experiment 2 and the constructed community experiment, therefore, we tested the effect of the leaf wash 100K MWCO filtrate treatment separately from that of the phage fraction. For field experiment 1 we verified that starting inoculum across the six different sources was within the same order of magnitude using OD600 measurements, and for the second experiment, we quantified starting bacterial density using CFU plating on Kings Medium, which was determined to be ≈5*105 CFU/mL. Size fractions and bacteria were combined to create each inoculum immediately before application onto tomato plants to avoid any impact of phages as a result of interactions outside of the host.

For culture-independent community analyses, sequence files were demultiplexed by QB3 sequencing facility. Reads were combined into contigs using VSearch and the remainder of the analysis was carried out in Mothur following their MiSeq SOP. Data were quality filtered and chimeras were removed using UChime. The Mothur pipeline uses de novo cluster-based OTU picking that does not rely on taxonomic assignment, and we used a 97% similarity cutoff for defining OTUs. Archaeal, chloroplast, mitochondrial, and unknown domain DNA sequences were removed. Once an OTU table was generated in Mothur via Galaxy web application. The same filtered data were used to calculate Bray-Curtis distances, which were used for beta diversity and Principle Coordinates analysis. Graphs were plotted using the GG2 plot package in R . All other statistical tests were performed in SPSS version 24. For field experiment 2, statistical analyses were first run with all treatments, including the 100K MWCO filtrate treatment. However, the a priori predictions we sought to test with this experiment were comparing phage-depleted and phage coinoculated treatments. We had no expectations about what would happen in the 100K MWCO filtrate treatment. Therefore, in some cases , the bacteria plus filtrate samples were removed from statistical analysis in order to allow direct testing of our predictions.

This was especially important given the surprisingly large amount of among-replicate variation in both abundance and composition observed in the bacteria plus filtrate treatment.A final experiment was conducted using a constructed bacterial community of tomato phyllosphere isolates. We sought to include representative isolates of the most abundant families based on bacterial community sequencing data, and sixteen isolates were chosen . Isolates were grown separately overnight in Kings Broth, and the cultures were normalized to an OD600 of 0.0013, approximately 1×106 CFU/mL. To generate the non-bacterial fractions of inoculum, tomato leaves were collected from the UC Berkeley Student Organic Garden, and leaf wash and fractionation was performed as previously described. The bacterial portion of the leaf wash was discarded, and the phage fraction and leaf wash fraction were combined with the constructed community of phyllosphere isolates to generate bacteria plus phage and bacteria plus filtrate treatments. Tomato plants were inoculated and sampled as described above with the addition of four plants inoculated with sterile buffer only. Additionally, given that in this case we had a panel of culturable hosts, we sought to isolate phage from the phagefraction used in this experiment with the following method. Ten µl of phage fraction was combined with 80 µl of bacterial culture from each of the 16 isolates and 7mL of Kings Broth and grown overnight at 28°C to enrich for phage. The following day, cultures were filtered using a 0.22µm filter unit. Filtrate was spotted on soft agar overlays of each of the 16 bacterial isolates from which it was generated. Additionally, filtrate from each of the 16 enrichments was pooled and spotted onto all 16 soft agar overlays. Plates were incubated at 28°C overnight and presence of lytic phage was determined by presence of zones of clearing, i.e. plaques, on the soft-agar overlay plates.Until recently, very little was known about the role of bacteriophages in the phyllosphere community. Work from our lab on tomato leaves shows that the phage fraction of the phyllosphere microbiome is capable of decreasing bacterial abundance within the first day of colonization of a new host, and this phage fraction also impacts bacterial composition and diversity. Other work conducted in the horse chestnut tree phyllosphere demonstrated that bacteria evolve resistance to phages relatively quickly over time in a natural system. The bacterial evolution of resistance to phage predation is thought to be one mechanism by which phages can contribute to, or maintain their diversity. Phage can also increase bacterial density and diversity by releasing nutrients into the environment via cell lysis, thus increasing its availability to other members of the community. One of the more frequently discussed mechanisms by which they maintain diversity in the community is through “kill-the- winner” dynamics. This theory suggests that the most abundant bacteria are also the most susceptible to phage predation. In such a system, an increase in abundance of a particular bacterial taxon is followed by an increase in abundance of the associated phage population and therefore a subsequent decrease in abundance of the susceptible bacterial strain, effectively preventing one type of bacterium from ever dominating the community In the phyllosphere studies described above, pipp mobile storage systems only the effects of lytic phage on the bacterial communities were studied. Lytic phages are those that infect a bacterial host, and upon replication, destroy the host cell. There is also increasing evidence for the prevalence and importance of temperate phages that are integrate into the hosts’ genome also commonly referred to as lysogenic phages.

In humans, these temperate phages are thought to dominate the phageome and outnumber lytic phages. The relative abundance of lytic to lysogenic phages may depend on factors such as host density and nutrient characteristics of the environment. Like lytic phages, temperate phages are also thought to increase bacterial diversity in communities through mechanisms such as facilitating horizontal gene transfer and conferring novel traits to bacteria upon lysogeny. For example, phage transduction mediates the acquisition of virulence factors in Staphylococcus aureus MRSA. They can also lysogenize and revert to a lytic lifecycle and contribute to bacterial diversity via the killing mechanisms described above. Virtually nothing is known about the prevalence and importance of temperate phages in plant-associated microbial communities. Here, I use a community passaging approach to disentangle the effects of lytic and lysogenic bacteriophages on epiphytic bacterial communities in the phyllosphere. Phyllosphere microbiomes were allowed to colonize plants in succession in the presence and absence of their “free” phage fraction , and under conditions that did or did not allow co-evolution of the bacterial and phage in these communities. Specifically, the experiment consisted of the following passage lines: 1) bacteria only, 2) both bacteria and phage, 3) bacteria passaged between plants but only in the presence of ancestral phage, and 4) ancestral bacteria introduced at each plant passage but with phage that had evolved during previous passages . Passaging occurred weekly for three weeks on a total of three cohorts of plant hosts; the communities were passaged from one cohort of plants to the next every week for a total of three passages. At the end of three weeks, microbiomes were collected, and 16S rRNA amplicon sequencing was used to describe the bacterial communities.Additionally, bacteria were cultured from each plant cohort at the final time point, and attempts were made to culture lytic phages. Attempts were almost made to induce lysogenic phages from the bacterial isolates.By analyzing bacterial communities via 16S amplicon sequencing, it was apparent that by three weeks after initial inoculation that the treatments resulted in distinct bacterial communities. After calculating Bray-Curtis distances between all samples, a measure of bacterial community dissimilarity, samples are plotted on a PCoA plot . The plot indicates separation amongst treatment. Furthermore, the original inoculum sample that was sequenced was dissimilar to all experimental samples, indicating that the community that was originally sprayed onto the plants changed over the course of the three weeks. Indeed, sample type explains 14% of the dissimilarity amongst all samples . When the inoculum sample is removed and the effect of treatment is analyzed using the same ANOSIM test, 42% of variation amongst the remaining samples can be explained by treatment . Because treatment BaP seems distinct from the rest, those samples were removed and the test was re-run. In this case, treatment remains significant , but with only 26% of variation explained . The data can also be analyzed by distinguishing between which types of phages were passaged during the experiment . In B only lines, only lysogenic phages would have been passaged. In the BP line, both lytic and lysogenic would have been passaged. In the BaP line , only lytic phages would have been passaged. Lastly, in the BPa line , only lysogenic phages would have been passaged: although ancestral lytic phages were applied at each passage, only those that were already in or had been incorporated into the bacterial genome were passaged onward. Therefore, lines can be classified as “lytic passage ”; “lytic and lysogenic passaged ”; and “lysogenic passaged ” respectively. When visualized in this manner , samples do appear to be distinguished by that of the phages that were passaged. Statistically, 22% of the dissimilarity in bacterial community composition is explained by this variable , although this is, of course, reflective of the overall treatment effect observed in Figure 5-1. Visually, it appears as though there may be a continuum in bacterial community composition in which lytic and lysogenic only lines are the least similar, while communities associated with lytic/lysogenic show intermediate changes between these two extremes. Statistically however, this is not the case. Lyt/Lys samples are just as similar to Lyt samples as they are to Lys samples . A similar analysis can be conducted in which the bacteria-onlylines are excluded, and only treatments in which lytic phages were passaged and/or applied are compared. In this analysis, treatment explains 39% of variation amongst samples .From each of the six replicates of the 4 treatments, bacteria were isolated, totaling 80 bacterial strains. These bacteria isolates were used to “fish” for lytic phages in both the resultant phage fractions obtained after passaging in each treatment and also from the phage fraction from the original inoculum. Then, for the B and BP lines, lysogeny of temperate phages was induced via UV treatment.

When inoculum and control samples are removed from analysis, there remains a significant effect of passage number and a significant overall effect of plant genotype on community composition . When variance is partitioned, passage can explain 51% of dissimilarity, whereas genotype explains only 4%. Replicate lines from accession 2934 were lost after P3 due to a stem rot fungal pathogen present in the original inoculum that seemingly only infected this genotype. However, the observed overall genotype effect was not driven by this accession, as there remains a significant effect of genotype after its removal , and passage number remains highly significant . To better understand how the original, diverse, field inoculum changed over four passages on plants in the greenhouse, we calculated the percentage of OTUs in the original inoculum that were detectable over the course of the experiment . At the end of P1, 92% of the field inoculum OTUs were still present on the plants, but by P4, this was reduced to 29%. We then calculated if the decrease in original community member diversity was the result of replacement by non-inoculum taxa . In this case, we observed that the proportion of sequencing reads representing the original inoculum OTUs remains above 78% . This suggests that a relatively small percentage of the community was made up of OTUs that colonized plants from the greenhouse environment.

Of note, some OTUs considered “non-inoculum” were likely present in the initial inoculum, but in too low of abundance to detect. In particular, there were 27 OTUs with reads in the spray inoculum sample in the non-rarefied dataset, but this was number was reduced to zero after data rarefaction. To account for the impact of the small percentage of arriving species on community composition, we re-analyzed the dataset using only those OTUs that were observed to be present in the initial inoculum . In this case, passage number remains a significant driver of community dissimilarity , as does genotype . We next measured changes in community diversity over the course of passaging and across lines. We found a significant decrease in both OTU richness and alpha diversity over time across all plant genotypes , including when only original spray inoculum OTUs are considered . Importantly, this drop in diversity from the start of the experiment does not correspond to a decrease in overall bacterial abundance on plants . Note that our measures of bacterial growth likely largely overestimate the starting densities and do not account for population turnover , and are therefore highly conservative. In P1, we also estimated fold change of bacterial abundance on control plants that were sprayed with heatkilled inoculum, and found an average change of 0.76, which is significantly lower than the averaged 11-fold change for experimental plants which received live inoculum .

Finally, although passaging was performed in a control temperature greenhouse, outside high and low temperatures and humidity all varied significantly across passages , which may have impacted the observed differences in both abundance and growth across passages. With the knowledge that communities were drastically changing over time, we sought to determine if the rate at which the communities were changing was consistent. To do this, we calculated Bray-Curtis distances of microbiomes in each passage to P1 microbiomes . As we similarly observed through ordination plots in Figure 1, the communities become less similar to P1 over time. We then fit both a linear and quadratic regression to these data, and we found a better fit of a quadratic model than linear. We determined this through comparing both R2 values and calculating Akaike information criterion values . Both models were highly significant . Taken together, this suggests that community change may be slowing down, although it appears to have not entirely stopped. We next observed changes in relative abundance of specific taxa within lines over time . At each passage, there are numerous taxa that are differentially abundant compared to other passages. In some cases, there was evidence for replacement of OTUs within taxonomic groups. Specifically, in the top 10 most differentially abundant taxa as determined by using a Kruskal-Wallis test, three of them are in the family Pseudomonadaceae. OTUs 0010 and 0004 are in significantly higher relative abundance in P1 than in P4 , and gradually decreased in relative abundance, whereas OTU0002, an unclassified Pseudomonadaceae, is significantly more abundant in P4 as compared to other passages . All three OTUs are present in the initial spray inoculum, although OTU0002 represents only 0.03% of rarified spray inoculum reads whereas OTU0004 represents 27% and OTU0010 represents 21%. To better understand how bacterial community dynamics were changing over the course of the four passages, we utilized a recently developed cohesion metric to quantify connectivity of microbial community. In brief, community cohesion is a computational method used to predict within-microbiome dynamics by quantifying connectivity of microbial communities based on pairwise correlations and relative abundance of taxa. Changes in community cohesion over time are suggestive of biotic interactions, where connectivity can arise from either, or both, positive and negative interactions resulting from cross-feeding or competition as well as environmental co-filtering. When applied to our dataset , we find a mild but significant increase in positive cohesion values from P1 to P4 . Consistent with positive cohesion values showing increased biotic interactions, there are also increasingly negative cohesion values from P1 to P4, which again is mild but significant . To test if bacterial communities were changing due to neutral processes alone, we first applied the Sloan neutral community model and found that a neutral model is less correlated with observed communities on the plants over time . However our data violate a key assumption of the neutral model in that dispersal was experimentally constrained within lines, and thus we took the approach of generating a null prediction based on the known community composition of inocula applied at each passage and comparing our observed communities to the predicted neutral community using a recently developed approached. We found that Bray Curtis distances between predicted and observed communities moderately increases over time , suggesting that community change over the course of the passaging experimentis likely the result of deterministic rather than neutral processes. Further evidence for a shift away from neutrality can be observed using occupancy- abundance curves in which the occupancy, or proportion of individuals in which an OTU is found, is plotted against its relative abundance. A positive correlation between the two is expected to occur by chance, curing marijuana as in a neutrally assembled community, but a change in distribution of individuals may indicate a community shaped by deterministic processes. When our data are visualized in this manner , we see that in P1, the most abundant taxa also occupy the highest proportion of plants, as you would expect in a neutral community not undergoing niche selection.

However, this trend collapses by P4 with many abundant taxa occupying far fewer individuals than would be expected under an assumption of neutrality. We next designed an experiment to which we could apply Sloan’s model of neutral theory . All lines from the end of P4 were pooled together and re-inoculated onto tomato plants, mimicking the inoculation procedure from the first passage. Plants that received the P4-pooled inoculum had significantly different bacterial community composition than the P4 plants themselves . We did not observe an effect of genotype on the communities assembled from this combined inoculum . We also found that the majority of the variation between samples was driven by an exceptional situation of introduction of a greenhouse taxon to the plants . To test if neutral processes were driving community structure in this experiment, we again examined fit to a neutral model using the Sloan model approach. In this case, as with P1, the assumption of equal dispersal potential among plants is met. In 200 iterative predictions, the fit of the neutral model is significantly higher in P1 than P4 Combined , suggesting that neutral processes are dictating the community structure after the first passage, but not in the P4 Combined experiment . We also see the occupancy-abundance relationship breakdown in P4-Combined when compared to P1 directly .The similarity of changes in community structure both across replicates and genotypes over the course of the passaging experiment led us to predict that these microbiomes were adapting to the local plant and greenhouse environment. To further determine if the community changes we observed from P1 to P4 were due to habitat selection rather than neutral processes, we employed a community coalescence competition experiment. In this experiment , phyllosphere communities from the end of P1 and the end of P4 were inoculated onto a new cohort of plants, either on their own or in an approximately 50:50 mixture of live cells . To ensure that our method for the mixed inoculum was effective, we sequenced multiple replicates of the P1, P4, and Mix inoculums and found that source of inoculum explains 88% of dissimilarity amongst samples . To ensure that the Mix inoculum was significantly different than both P1 and P4 separately, we compared P1 and Mix inocula directly and found that 75% of difference between samples can be explained by this variable . Similarly, when P4 and Mix are compared directly, 74% of variation in the community is explained . This consistent difference among the three inocula allowed us to compare the communities colonizing plants from each treatment. We first measured final bacterial abundance and found that colonization was lower on these plants than in previous experiments, but does not significantly differ among treatments , apart from control plants, where bacterial colonization was greatly reduced . We then compared bacterial communities again using 16S amplicon sequencing and ordinated samples on a PCoA based on Bray-Curtis distances. Plants that received P1 inoculum had distinctly different communities than those that received either P4 or the Mixed inoculum. Plants that received the Mixed inoculum clustered together with those receiving P4 and were relatively indistinguishable. Using ADONIS tests, we determined that inoculum source can explain 45% of Bray-Curtis dissimilarity amongst samples , and there was no effect of plant genotype . In a pairwise analysis between P1 and Mixed, inoculum source explains 31% of the community dissimilarity . In contrast, inoculum source does not explain any significant variation in dissimilarity amongst P4 and Mixed inoculum plants . Together, these results suggest that the plants receiving the 50:50 mixed inoculum were indistinguishable in community composition from those receiving the pooled, P4 adapted microbiomes, and that these selected communities were not invadable by the microbial communities from the start of the experiment. Consistent with our results from the passaging experiment itself, alpha diversity was highest in P1 plants compared to both P4 and Mixed plants . Alpha diversity did not differ amongst communities colonizing plants from the P4 and Mixed inoculums, despite being different between the two inocula themselves. We also examined compositional makeup of the communities , and consistent with P1 to P4 passaging results, we see differentially abundant taxa between groups . Again, two Pseudomonas OTUs are more abundant in P1 plants as compared to P4 and Mix, in which there was an unclassified Pseudomonaceae that was higher in relative abundance.

On the other hand, the field upstream facility showed a significant increase in COGS at lower production levels due to the minimum ownership costs of field equipment regardless of the small acreage size. DSP followed the expected behavior that economy of scale dictates, weed dryer with sharp decrease in COGS at lower production levels and diminishing returns at higher production levels. The deviation from linear trend at 150 MT/year in field upstream and DSP is likely due to the model’s specified equipment maximum rating, which allows for the inclusion of a new equipment in parallel beyond this rating.The impact of varying the highest cost drivers in each of the facility’s category by 25% on COGS is portrayed as a tornado diagram in Figure 5c. Field labor was the most sensitive cost variable, having the highest impact on the COGS, followed by the ultrafiltration membrane, which is replaced every 30 cycles. In this model, we assume a relatively high downstream recovery of the protein from harvest to formulation. The reason for this assumption is that spinach, being edible crop, allows for a lower target product purity and a consequently fewer DSP steps. It is particularly important to focus resources on maximizing downstream recovery during process development because it ultimately affects plant biomass and spray volume requirement upstream to appropriately compensate for these losses, which in turn affects equipment sizing in DSP based on the amount of plant material to be processed.

The unit operations were resized according to the scenario design requirement for downstream recovery ranging from 50 to 95% while scheduling parameters were left unchanged. This effect of downstream recovery on theAlthough our analysis indicates a relatively high COGS range for a sugar substitute, there are unrealized costs savings from thaumatin use due to its unique sweetness intensity. Thaumatin’s use in extremely small quantities is essentially why it is considered a noncaloric sweetener, as it provides only 4 calories per gram. Sensory evaluation studies have found that a sample with 5% sucrose +4.6 ppm thaumatin II had similar sweetness as a 10% sucrose control with minimal lingering aftertaste, suggesting that up to one-half of the sugar could be replaced by thaumatin II . SSBs including sodas, fruit drinks, and sport drinks account for 50% of the total added sugar in Western diets, and therefore provide an attractive avenue for thaumatin emergence as a sugar substitute. The incorporation of thaumatin by the industry not only offers a tool to help decelerate the obesity epidemic caused by increased childhood sugar intake decades ago, but also provides itself with a more economically viable solution. Firstly, as sugar taxations emerge, sugar reduction becomes a financial incentive. Secondly, the reduction of sugar and the addition of thaumatin to retain the same level of sweetness has the potential to save millions of dollars per day on the cost of sweetening beverages.

Assuming that the average “standard” sucrose concentration in SSBs is 35.5 g per 12 fl oz. drink ~10% , and a $0.30/kg sugar price, Figure 6 shows the potential savings from using thaumatin to reduce sugar content by 20%, 30%, and 50%, while maintaining the same sweetness as the standard for a range of thaumatin purchase prices. The amount of thaumatin needed to obtain the same sweetness as a 10% solution in each sugar reduction scenario was calculated using the sensory regression analysis included in a published GRAS notice . Table 3 shows the daily and annual amount of thaumatin needed for each sugar reductions scenario, assuming that one billion 12 fl oz drinks are to be sweetened per day. Successful implementation of thaumatin in this avenue can liberate R&D resources to improve expression levels and increase production volumes, both of which have a substantial impact on COGS reduction, as we have demonstrated.Our preliminary engineering facility design indicates the feasibility of thaumatin manufacturing by various molecular farming platforms. The most economic method is the field grown ethanol-inducible, transgenic N. tabacum, assuming a downstream facility without chromatography . It remains unclear whether heat incubation is sufficient to achieve the desired purity for a safe product without the inclusion of chromatography on a large-scale. In a previous plant-made food safety product techno-economic analysis, a chromatography unit was included for protein purification from N. benthamiana; however, heat precipitation of host cell proteins was not included as a purification step. We also demonstrate the importance of resin selection and thorough chromatography operation optimization by evaluating the cost benefit of maximizing resin binding capacity to target product. Of course, further work is needed to verify whether the use of column chromatography is needed. Transient production of thaumatin in the edible crop Spinacia oleracea was also economically competitive and captures the benefits of obviating the need for an intensive DSP. According to this analysis, the cost to produce a kg of fresh weight of spinach is $0.10, as opposed to a cheaper price for tobacco . This is attributed to the higher cost of the seeds of spinach, the longer turnaround time assumed for spinach, and the higher plant density assumed for tobacco. It is evident that field operation is very labor intensive, due to the low recipe cycle time of 2 days, which is different than the traditional time frame of growing those crops. The potential for high intra-batch variations in product yield and quality due to meteorological factors is one of the concerns of using field grown plant material for this application. These variations in turns cause inconsistency in key facility performance parameters that should be quantified using a probabilistic approach and communicated to stakeholders and will be addressed in a follow-up communication. The cost of obtaining a more controlled supply of product is reflected in the indoor upstream facilities CAPEX and COGS. This should facilitate decision making when assessing the risk and reward of each scenario. The large-scale recombinant production of thaumatin can address the growing market need for natural, safe, non-caloric sweeteners. Like stevia, the advent of thaumatin as a sugar substitute is contingent on the feasibility of its large-scale manufacturing which was addressed in this work. However, there are also social, cultural, cannabis curing and behavioral factors impacting sugar consumption habits that were not considered. Consumer’s preference of such products will open the door for more plant-made biologics for food and beverage applications, which could drive the adoption of cost-effective solutions to rising challenges through environmentally friendly and sustainable processes.As the demand for agricultural production grows and additional challenges arise as our climate changes, there is increasing interest in harnessing beneficial plant-microbe interactions to improve promote plant growth and health. Furthermore, though plants and microbes have been co-evolving for millions of years, human agriculture has only been in practice for 12,000 years. Plant breeding and agricultural processes such as chemical usage, monoculture, and annual tillage are far removed from how plants naturally grow and thrive, and as such, there is a need to understand how agricultural and breeding practices impact plant-microbe interactions. There remain key open questions regarding the effects of plant breeding for agriculturally beneficial traits, transmission of plant microbiota, and overall importance of microbial community living on the above ground surfaces of plants, a habitat known as the phyllosphere.The plant host study system in this work is the modern tomato plant, Solanum lycopersicum. Tomatoes have an extensive and interesting history of domestication.

Their wild ancestors, Solanum pimpinellifolium, originate from the Andean region of South America. They are thought to have been domesticated initially in Mexico and then brought to Europe in the 1500s. Though wild tomatoes have a large genetic diversity, domesticated cultivars are estimated to contain less than 5% of this genetic diversity due to a history of inbreeding and back crossing of domesticated lines. It was not until the early 1900s that breeders began to cross S. lycopersicum with wild lines to reintroduce genetic diversity and select for desirable traits such as disease resistance. The tomato plant is an ideal host system for various reasons. First, researchers and breeders have access to thousands of tomato accessions through the UC Davis Tomato Genetics Resource Center , allowing for consistent and reliable control over host genetics. Furthermore, tomato plants can be easily propagated in controlled-temperature growth chambers, the greenhouse, or the field. Finally, there is an applied interest in work such as this, as the tomato industry in the US produces 2.6 billion dollars worth of fresh and market tomatoes , and locally, is one of California’s top ten most valued agricultural products .Many of the ways in which microbial communities interact with their host is through the host immune system. As such, a cursory understanding of the plant immune system, and how this compares to immunity in other systems, is necessary to fully understand how microbes influence host health. The adaptive immune system is thought to have arisen in jawed fish ≈500 million years ago, whereas the innate immune system likely dates back to early eukaryotic cells themselves. As microbial communities greatly pre-date the existence of multicellular eukaryotes, both branches of the immune system evolved in the presence of microbes, and it follows that tolerance for commensal microbiota must have been a key factor in shaping the evolution of immunity. Innate immunity, found across all kingdoms of life, is non-specific and responds broadly to ‘non-self’ invaders. Its hallmarks include protective physical barriers and general pattern recognition receptors that sense non-self signals and elicit host responses. Adaptive immunity is unique to vertebrates and responds to specific pathogens through detection of antigens via somatically-generated receptors and specialized white blood cells , resulting in immunological memory, in which the organism is protected from future infection . Broadly speaking, the adaptive immune response is highly specific to particular pathogens and can change over the course of a host’s lifetime whereas the innate immune response is a general resistance that can only respond to selection across host generations. Plant innate immunity consists of two primary responses to microbes. The first branch of the immune system recognizes microbial- or pathogen-associated molecular patterns through the use of transmembrane pattern recognition receptors and results in pattern-triggered immunity. However, many plant pathogens have evolved to overcome these defenses through the use of effectors. Plants with resistance genes for specific pathogens can detect the effectors through NB-LRR proteins, which represent the second response to microbes: effector triggered immunity. Additionally, plants have physical barriers to infection such as cell wall defenses, and they can also secrete antimicrobial peptides to ward off infection. Although vertebrate adaptive immunity is typically considered more specific, plant immunity is also the result of specific recognition, either of pathogen effectors or molecular patterns, and immune ‘priming’ in plants has been shown repeatedly. As is becoming increasingly evident, in addition to its well-studied role in preventing pathogen establishment, the immune system also influences both the composition and abundance of non-pathogenic microbiota. In mammals, this is best studied in the gut microbiome, where differentiating between these diverse commensals and colonizing pathogens is clearly a highly complex problem.

In order to adjust their calendar, vertical growing weed the Mexica would have needed to know the position of the sun on particular dates of the solar year, a feat that could have been accomplished only by marking the sunrise bearing relative to a geographic landmark. Many studies have analyzed the alignment of temples and ceremonial centers with the sun’s azimuthal bearing at sunrise or sunset on culturally relevant dates, so the architectural orientation of major buildings such as the Templo Mayor, and in general the urban design, would reflect important calendar dates . This architectural design would have had a great symbolic, ritual, and cultural importance but would have not been the most accurate way of measuring the annual march of time because of parallax error: small shifts in the position of the observer relative to a building or ceremonial structure can project large errors in the distant horizon. Because Mesoamerican settlements were all located inside the tropics, i.e., south of latitude 23.5°N, the zenithal transition of the sun occurs here twice every year; first, in late May as the sun’s trajectory in the celestial sphere moves northward toward the summer solstice, and then back in late July, as the sun’s trajectory returns southward .

Two settlements south of the Basin of Mexico—Xochicalco and Monte Albán—had specially constructed “zenith tubes;” vertical shafts perforated on the rock or constructed inside large ceremonial buildings that would project direct sunlight onto an observation chamber below ground . The projection of these solar flecks on the ground would have allowed observers to keep track of exact solar dates and, potentially, to use them for calendric adjustments . However, no evidence exists that zenith tubes were ever used in the Basin of Mexico, and many scholars believe that the calendar-keepers of Tenochtitlan directly used the sun’s position at sunrise against the prominent peaks on the basin’s mountainous horizon as calendrical landmarks, a “horizon calendar” that provided accurate indicators of specific dates along the solar year .The use of the landscape as a calendric tool is based on the fact that because of the tilting of the Earth, the point in the horizon from where the sun rises shifts day-to-day along the year . In reality, the sun rises due east only during the equinoxes .

In the Northern Hemisphere summer when the North Pole is tilted toward the sun, the sun in the Basin of Mexico will rise north of due east in the celestial horizon, reaching a compass bearing of ca. 65° during the summer solstice. Likewise, in winter, it will rise south of the 90° east bearing, reaching an azimuth of ca. 115° during the winter solstice. The azimuth angle of the sun on the celestial horizon at sunrise is a function of the declination of the Sun from the celestial equator at any given date and the latitude of the observation point . If the observer is on a fixed location, say, the Templo Mayor, then latitude is constant and the solar azimuth at sunrise becomes a direct function of the winter-to-summer declination of the sun, which is in turn a function of the date in the solar year. So, by looking at sunrise against a distant mountainous landscape from a fixed point, an observer can keep tab on the days in a year with minimal parallax error. Landscape features are still often used to mark calendric dates in many traditional villages. Despite modern communications and the standardization of calendric time, many communities still have ceremonies related to planting or harvest that are celebrated when sunrise or sunset is aligned with mountains that act as reference points . Using landscape silhouettes in the horizon as a means for keeping count of time and seasons has been so important in the past that some cultures have even built their own calendric reference points to use as a sunrise observatory in flat terrains, like the towers of Chankillo in north coastal Peru, built in the 4th century BCE . In the Basin of Mexico, as in many other parts of the Americas, the alignment of the rising sun with mountains seems to have been a common calendric and navigational tool. For example, the Florentine Codex describes how mineral experts used the alignment of sunrise against surrounding mountains to mark and relocate mineral deposits .The central hypothesis of this study is that the eastern mountain landscape of the Basin of Mexico played a central role as a tool for adjusting the calendar system to the solar year. In contrast, we will concentrate on the ecological and agricultural importance of the Basin’s landscape as a tool for accurate calendric adjustments. Our study concentrates on the xiuhpohualli or “year count,” the solar calendar that regulated the cycles of agriculture . We explore four hypotheses: a) The eastern horizon landmarks could have provided a series of reference points that would have allowed the adjustment of the agricultural calendar to the solar year. b) There is evidence in the texts of early chroniclers and codices that the Mexica people were indeed using horizon landmarks to follow the dates of the solar year. c) An ancient construction in the summit of Mount Tlaloc seems to have been used as a fixed solar observatory built for the purpose of calendric adjustments. d) Before the foundation of Tenochtitlan and the establishment of the Aztec dominion in the Basin of Mexico, other large agricultural civilizations that preceded them were also using the Basin’s mountainous horizon for calendric purposes.

For example, viewed from the top of the Templo Mayor, sunrise during the winter solstice would have broken at an azimuth of 116°22′ and an angular elevation of 3°20′ behind Mount Tehuicocone, a prominence on the northern slope the great Iztaccíhuatl volcano and very close to the “head” of the volcano’s “sleeping woman” silhouette . During the summer solstice, the sun would have been seen rising behind the settlement of Tepetlaoxtoc, pipp shelving in the foothills of the low Sierra de Patlachique, north-east of Texcoco, with an azimuth of 65°04′ and an angular elevation of 10′. On March 16 and September 30, the sun would have been seen rising behind the peak of Mount Tlaloc, close to due east, with an azimuth of 93°17′ and an angular elevation of 2°51′. Finally, on March 1 and October 15, the sun would have been seen rising behind the peak of Mount Telapón with an azimuth of 98°55′ and an angular elevation of 2°45′ . Not all these landmark points, however, have the same resolution for the purpose of keeping count of the days in the solar year. The solar azimuth at sunrise will vary at the latitude of the Basin of Mexico from summer to winter forming a wave-like function along the year. The changes in azimuth from one day to the next— i.e., the azimuthal shifts, or difference between the sunrise azimuth one day and that of the previous day—form a similarly shaped function displaced ca. 91 d . During the spring equinox the daily azimuthal shift at sunrise will be −0.418° . During the fall equinox the azimuthal shift will be numerically similar but opposite in sign . Near the summer and winter solstices, sunrise will seem to stand still for some 10 d, shifting its sunrise azimuth in 2′ or less per day and appearing to rise in the horizon from the same spot . The angular size of the sun’s disk is ca. 31′, a value only slightly larger than the daily shift in sunrise azimuth during the equinox . This means that an observer seeing the sun rise behind a horizon landmark near the equinox—like, for example, Mount Tlaloc—would see sunrise azimuths shift daily by a value that is 81% as large as the sun’s apparent diameter. In simple terms, there is only 1 d in the spring and 1 d in the fall in which the sun can be seen rising exactly behind Mount Tlaloc. In contrast, during the solstices, when the sun reaches its maximum declination, the sunrise azimuth varies in less than 2′, that is, one-fifteenth of the angular size of the solar disk, an amount that is not perceptible to the naked eye. For a Mexica astronomer following the calendric horizon, during late December and late June, the point of sunrise would seem to stand unchanging for at least 10 d. It seems, then, reasonable to assume that Mexica astronomers keeping a record of the calendric horizon would have used Mount Tlaloc, the horizontal point of reference nearest to the equinox, as their fundamental tool for calendar counts and calendric adjustments, because this landmark would have given them maximum calendric precision. Following this approach, the number of days in a year could be counted from the day that the sun rises behind Mount Tlaloc in spring to the next springtime sunrise behind Mount Tlaloc. The count would be 365 d: the 18 Mexica “months” of 20 d each plus the 5 nemontemi, or “useless” days, at the end of the year. But anyone keeping count of days this way would observe a gradual displacement of the sun’s sunrise position because the true length of the solar year is closer to 365.2422 d. After four years, adding an extra nemontemi day to adjust the day count to the horizon calendar would have been necessary in order to keep the sun rising behind the peak of Mount Tlaloc for the same day of the year. This hypothesized adjustment using the distant horizon, however, is only possible, as discussed above, if a landmark point near the equinox is used for the calendric adjustments, so that the precise day of the year can be identified without error.ccording to Sahagún’s 1575 description , the new year in the Basin of Mexico started on February 2 of the Julian Calendar in use at that time, which translates to February 12 of our current Gregorian calendar. However, comparing dates of historical events between 1519 and 1521 that were recorded both in Mexica codices and in Spanish chronicles, Tena estimated that at the time of the arrival of the first Europeans to Mexico, the first month of the Mexica calendar, Atlcahualo, began on February 13 of the Julian calendar or February 23 of the Gregorian calendar. In later papers , Tena added three more days to compensate for the suppression of leap days in the secular years of 1700, 1800, and1900, arguing that the Mexica calendar year started on February 26. Although his recent correction might be relevant to convert modern dates into the Mexica calendar, it seems clear that in the 16th Century, when the Gregorian calendar was established, the Mexica year started on February 23. Following Tena’s chronography, viewed from the Templo Mayor sunrise aligned with Mount Tlaloc 5 d before the equinox, at the end of Atlcahualo. In his description of the Mexica calendar, Sahagún noted that on Atlcahualo the Mexica “celebrated a feast […] to the Tlaloc gods, whom they held to be gods of rain.” The tributes to Tlaloc continued during the dry fore-summer: Sahagún also narrates that at the beginning of the third month—Tozoztontli ca. April 4—“a feast was made to a god called Tlaloc, who is the god of rains.” The clear association between Tlaloc, the equinox, and the dry springtime supports the assumption that sunrise behind Mount Tlaloc, viewed from Tenochtitlan, marked the highpoint of Mesoamerica’s premonsoon dry spring and led the people of the Basin to plead to Tlaloc for the arrival of the rains.

A relatively high rate of initial volitional emigration was seen in the first week across all fields, followed by a much lower rate of emigration in the second week , and steadily increasing emigration in weeks three through five . Manual fish recovery with a seine at the end of field drainage in the trenchless fields ranged between 5 to 20% of the total surviving fish compared to less than 0.5% of survivors from the trenched fields which indicated a more efficient drainage procedure in trenched fields. Again there appeared to be functional equivalence between the 0.5m and 1.0m trench treatments.Rearing of juvenile Chinook Salmon within winter-flooded rice fields shows strong potential for reconciling agricultural floodplain land use with habitat needs of an imperiled and economically important fish. Winter-flooded rice fields demonstrated high production of naturally occurring fish food leading to high growth rates of salmon reared in these environments. As with past fish conservation studies in altered environments, our adaptive research approach enabled us to successfully answer experimental questions despite unpredictable winter hydrologic and temperature regimes in the Central Valley. In our studies, post-harvest field substrate did not have a statistically significant effect on the composition or abundance of zooplankton species, nor on growth rates of rearing juvenile Chinook Salmon.

Overall, fish growth across all treatments was extremely fast and much greater than previously documented in the Sacramento River channel environments over the last century. Accordingly, we do not recommend a specific post-harvest straw management practice. Instead, we feel that field preparation should be left to the farmer. However, we encourage future research that explores other approaches for enhancing in-field habitats to decrease predation risk for rearing fish. There is currently limited means of cost effectively providing avian predation refugia for fish on winter-flooded rice fields. We investigated the potential of in-field trenches to provide depth refuge from avian predation, but direct benefits to survival were found to be insignificant in this study. Fields containing perimeter trenches connecting the inlet and outlet structures did however, show higher rates of volitional emigration of salmon and reduced rates of stranding following draining. We speculate that fish used the trenches as migration corridors when emigrating from the fields. Increased rates of volitional egress would further diversify timing of emigration which has been identified as a key component of population stability via the portfolio effect. Additionally, the trenches buffered water temperatures from the daily maximums observed in the middle of the fields, expedited field drainage, and reduced the number of fish stranded during field draw down.

In floodplain river ecosystems, fishes often respond strongly to hydrological dynamics of ascending and descending flood conditions. Juvenile Chinook Salmon in the Central Valley have evolved physiological and behavioral strategies for the use and egress from winter flooded floodplain habitats. Accordingly, the rate of field drainage and inflow conditions in winter-flooded rice fields may provide important cues for rearing juvenile Chinook Salmon. In our study, extending the drainage period and manipulating inflow conditions in the slow drain treatments had a detrimental effect on survival and the best method was a fast drain where fields were drained in a single day. This was likely the result of increased vulnerability to predation and reduced thermal buffering due to a prolonged period with shallower water depths in the slow drain treatments. Again, these results provide a relatively simple management recommendation for farmers in that simple opening of outlet water control structures with rapid drainage appears to be the best method. We encourage exploration of other drainage methods, and production of other species in winter-flooded rice fields may require different draining practices. An initial mortality of approximately one third was observed in the first week of the 2016 salmon survival through time experiment. The cause of this initial mortality is unknown and could have resulted from a combination of factors, including a stressful transport and acclimation stress under new physical water conditions. Additionally, because we know of no other experiments that have been able to track and assess post-release mortality of hatchery fish through time, we cannot rule out the possibility that the high rate of initial mortality observed immediately after release into the “wild” is a potentially common phenomenon. Transport is a known stressor on many fishes, including juvenile Chinook Salmon. In our study, fish were captured from hatchery raceways, coded wire tagged, allowed to recover for several days and then placed in a fish hauling tank at high densities and delivered to the fields in early February. Exposing naïve hatchery salmon to a new environment in the flooded agricultural fields may have increased stress as it necessitated behavioral adaptations of prey switching and predator avoidance as well as rapid acclimation to the new physical water quality parameters. After the high rate of initial mortality, survival stabilized in week two and remained high for the remainder of the experiment. Without accurate assessment and accounting of initial post-release mortality there is potential for fishery managers to be chronically overestimating habitat-specific mortality rates determined by recapture of hatchery fish transported and released into natural habitats. We therefore recommend that future research examine effects on initial post-release mortality of transporting, acclimatizing, and releasing hatchery fish into the wild.While farmers have incentives to prepare their fields for a new rice crop as early in the spring as possible, fish conservationists may theoretically prefer to keep the rice fields inundated as late as possible to maximize fish growth and survival before release into the river. However, in practice, when weather conditions on the floodplain are good for fish in the late winter and early spring, they are generally not conducive to agricultural field preparation. The inverse is also true, as spring conditions become dry and hot and generally suitable for agricultural field preparation, water quality conditions rapidly become unsuitable for juvenile Chinook Salmon . Thus, given proper timing and coordination within an adaptive management framework, farmers and fish conservationists can collaborate to promote threatened fisheries without impacting crop yields.

We therefore encourage the continued development of adaptive frameworks for the integration of floodplain fish habitat into farm operations that reconcile the needs and timing for both fish and farm operations. Incentive programs for farmers may be needed to promote these activities to their fullest potential. Land manager and farmer involvement has generally exceeded expectations in our projects, and we are optimistic about continued stakeholder involvement. Given issues with water scarcity in the Central Valley, pipp rack the dual-use of rice fields for agriculture and rearing juvenile salmon could establish stronger water security for farmers. Additionally, off-season inundation of rice fields promotes rice straw decomposition while approximating the natural long-duration inundation patterns that fuel a productive aquatic food web. When compared to concurrent samples in the adjacent Sacramento River channel habitat, the winter-flooded rice fields had 150x zooplankton abundance in 2013 and approximately 53x zooplankton abundance in 2016. Resultantly, the juvenile salmon growth rates we observed in winter-flooded rice fields were 2-5x higher than previously or concurrently observed in the adjacent Sacramento River. By creating high quality habitat on their fields, farmers can help bolster fish populations by rapidly turning small fry into large, healthy smolts during mid-winter when water temperatures are low, river flows are high and when predators are less active, thus improving salmon survival rates during outmigration. Managed inundation of rice fields in winter and early spring appears to mimic historical Sacramento Valley floodplain processes, re-exposing salmon to an approximated version of the hydrologic selection regime under which they evolved and to which they are adapted. The exceptional productivity and resulting rapid rates of salmon growth documented on the managed agricultural floodplain lead us to conclude that winter inundation of rice fields creates high-quality rearing opportunities for juvenile Chinook Salmon. Although these studies suggest that agricultural landscapes can function as high-quality rearing habitat for juvenile Chinook Salmon, our results should not be interpreted to diminish the conservation need for restoring naturally functioning floodplains where feasible or to suggest that suitable natural habitats are not essential to establishing self-sustaining runs of naturally produced Central Valley Chinook Salmon. Rather, these data demonstrate the potential to reconcile management of agricultural floodplain landscapes with the conservation of wild Chinook Salmon populations through slight modification and reoperation of existing agricultural infrastructure. Managed agricultural floodplains are likely to become another important means for fishery managers to produce ecologically functioning off-channel habits for imperiled native fish, especially during times of low water when remaining natural floodplain habitats do not inundate and are therefore inaccessible to salmon populations confined to leveed stream channels.The Special Feature on Diversified Farming Systems is motivated by a desire to understand how agriculture designed according to whole-systems, agroecological principles can contribute to creating a more sustainable, socially just, and secure global food system. “How to feed the world” is an increasingly urgent and looming concern voiced by many people, from local community groups to national and international governing bodies. By 2050, the world population is projected to rise to 9+ billion and food demands to double from current levels. At the same time, climate change, interacting with increasingly uneven access to declining oil, water, and phosphorus supplies, will greatly exacerbate the year-to-year unpredictability of agricultural production, potentially undermining the entire agricultural enterprise . Meanwhile, industrialized agricultural techniques are exacting a huge toll on surrounding environments, polluting waterways, creating dead zones in the oceans, destroying biodiverse habitats, releasing toxins into food chains, endangering public health via disease outbreaks and pesticide exposures, and contributing to climate warming . Moreover, industrial agricultural methods are inherently unsustainable in mining soils and aquifers far more quickly than they can be replenished, and in their high use of fossil fuels . These numerous environmental and social externalities create a huge economic cost that industrialized food producers seldom pay. For instance, pesticide use alone causes up to $10 billion in damage to humans and ecosystems in the United States every year . Finally, although the agricultural sector currently produces more than enough calories to feed humanity, one billion people remain hungry and an additional one billion have micronutrient deficiencies . This paradoxical situation occurs because many people still lack access to sufficiently diverse and healthy food, or the means to produce it, which is primarily a problem of distribution rather than production . As further evidence of this paradox, global obesity rates have more than doubled since 1980 , reflecting an overproduction of food in industrialized countries that creates strong incentives for agrifood companies to absorb excess food production into processed foods and to market and distribute them to customers in supersized portions . This series of articles examines the proposition that diversified farming systems, with their focus on local production, local and agroecological knowledge, and whole systems approachesreduce negative environmental externalities and decrease social costs associated with industrialized monocultures, enhance the sustainability and resilience of agriculture, and contribute significantly to global food security and health.We refer to a farming system as “diversified” when it intentionally includes functional biodiversity at multiple spatial and/or temporal scales, through practices developed via traditional and/or agroecological scientific knowledge. At the plot scale, diversified farming systems may include multiple genetic varieties of a given crop and/or multiple crops grown together as polycultures, and may stimulate biodiversity within the soil through addition of compost or manure .

Their results focus mostly on statistics of characteristics of overall U.S. female-run farms and female farm operators. They find that 58% of all female operators have no reported off-farm labor, and that female operators of dairy farms tend to be younger than the U.S. female operators’ average age. Griffin et al. utilize the COA data over five Census rounds and discuss the impact of farm operators’ demographics on farm exit rates. They find that larger farms are less likely to exit, and those female operators are more likely to exit than male operators. However, their study includes all farms with no industry limitations. Furthermore, research on female operators’ impact and representation within the dairy industry is a point of interest because, historically, it was not uncommon for dairy farms to be run by spouses and because off-farm employment is less likely on a dairy farm than it is on other farms. Sander finds that women working on dairy farms tend to have less off-farm employment than other farm types. He outlines the role of income variability on farms run by spouses’ decision to be both spouses’ main income with off-farm work as a possible risk mitigation strategy for farms run by spouses when farm revenue is highly variable. Schultz detailed some economic theories related to women focusing mainly on developing nations. Specifically, the role of family dynamics in economic choices on farms and female influence on such outcomes.

Rather than taking a theoretical approach, Zeuli and King provide detailed statistics of the characteristics of farmers and their commercial farms in 13 states. They find that in 1991 the average age of females relative to males is insignificant, but that the women in their sample tended to have a higher level of schooling. Interestingly, they found contradicting results, at least based on acreage, to other studies stating that women tend to manage smaller farms, with women operating more acreage on average, but this could be heavily influenced by what they grow and location. Sociology and anthropology research on female farm labor and agriculture tends to report findings based on case studies of specific regions and industries . These papers tend to discuss social incentives, norms, or barriers that influence the gender demographics of the industries of interest and, therefore, influence female representation and the impact of management decisions on the farm. Brasier et al. discuss the history of how women identify their labor on farms. Historically, female participation in farming communities was accessed through family or marriage. Typically, women involved in agriculture were either born into a family that farmed or married a farmer. In the past women often viewed their role on the farm as farm homemakers or farm helpers, following gender norms of the times, and often because they had off-farm income or only participated in farm labor seasonally .

This way of thinking about farm labor could have influenced the representation of female operators of farms. Trauger finds that women are more likely to adopt sustainable agriculture. Trauger limits itsscope to a few farms in Pennsylvania, finding that there may be a trend of female-operated farms to adopt socially minded practices, i.e., community education. This research helps build evidence that supports our claim that the presence of female operators can be considered a proxy variable for being adaptable to change. It seems like a basic assumption, but there was, and remains, a large share of women that participate in farm labor that were/are married to principal operators; this trend continues today. Therefore, the research on the relationship between gender and agriculture would not be complete without mentioning research done on agricultural spouses. A large share of female operators are the spouses of a farm operators. Barlett details the typical marriage models of agricultural spousal relationships, characterizing how farm labor related to agricultural spousal relationships is defined from a social perspective and may have influenced how women viewed their labor on the farm and subsequently the data representing farm labor, historically. The role of identity for female farmers and the professional connections can be a pivotal part of female farmer participation. This research provides evidence of the change in gender demographics based on farm size for the dairy industry. It adds to the literature detailed agricultural economic analysis on the intersection of women and agriculture for the dairy industry and discusses the change in data collection and availability by one of the most prevalent data sources for agricultural data, the COA.The survey questions asked of farmers and ranchers by the COA change slightly every Census round, although most remain the same across time. Below are descriptions of questions changes for relevant variables to the analysis. First, in 2002 and 2007, farms were asked for the total amount of dairy sales in that year, but in 2012 and 2017, this question was dropped and replaced with the total amount of milk sales. Furthermore, whether the dairy farm had any level of organic production was only asked 2007, 2012, and 2017. Second, operator characteristic questions have become more detailed over the years and allowed more operators’ data to be collected. In 2002, 2007, and 2012, the COA asked detailed operator characteristic questions about up to three operators, and only one operator was able to be identified as the principal operator. However, in 2017, the COA expanded its detailed operator questions to include up to four operators and now allows for up to four operators to be identified as a principal operator. Furthermore, in 2012, ebb and flow system the COA started asking farmers and ranchers if the secondary operators were married to the principal operator. This question was then adapted in 2017 to reflect the increase in possible principal operators identified and asked if the operator was married to a principal operator. The Census collects two categories of operators.

The first category is for which detailed operator characteristics and for which at most three operators are listed per farm in 2002-2012 and at most four operators per farm are listed in 2017. Going forward, the operators for which the number per farm is limited and detailed information is provided will be referred to the “core operators”. The second category has no limit to the number listed per farm and only gender of each operator and the number per farm is provided in the data.This section detail statistics and characteristics of female commercial dairy farm operators and their commercial dairies. The number of commercial dairies with at least one female core operator increased in every state, except New Mexico, which experienced no change from 2002 to 2017 . In 2017, every state, but New Mexico, has more than 40% of the commercial dairies reporting at least one female core operator. Although these states demonstrate significant increases in the representation of female core operators, the addition of a fourth core operator for the 2017 Census could distort these results. Table 5.2 shows the share of commercial dairies with at least one female operator by state and year. This has very interesting results with all commercial dairies reporting at least one female operator in 2017. All six states saw significant increases in the share of commercial dairies with at least one female operator. The actual share of female operators compared to the share of operators gives us a better representation of demographic changes. The share of female core operators increased from 2002 to 2017 in every state but New Mexico, for which the share of female core operators decreased in 2007 and 2012 but was the same in 2002 as in 2017 . California and New York both increased the number of across each Census year. California had a 27% increase in female core operators from 2002 to 2017 and the share of female core operators in New York increased by 33%. Idaho, Texas, and Wisconsin all had a slight decrease in female core operators in 2007 and 2012, but an increase in 2017 relative to all previous years. Interestingly, when we look at the share of female operators it follows a similar pattern. California and New York both increases in the share of female operators across each Census. Wisconsin, Idaho, and Texas all had slight decreases in 2007 and 2012 relative to the 2002 share, but the share of female operators in 2017 was larger than in 2002. However, the share of female operators in New Mexico had a small decrease from 2002 to 2017. This suggests that despite the addition of a fourth core operator in the 2017 COA the pattern is not substantially different from the trend in operators and that the trend was not only facilitated by capturing previously unmeasured management activities by women. From here characterizing the trend could be thought of in two ways: 1) this describes an actual increase in women operators playing a more prominent role and/or 2) an increase in their male associates being more likely to recognize and report female operators. Disentangling exactly what characterizes these trends is impossible, but it seems likely that the addition of a fourth core operator and the ability for more than one principal operator may have signaled a conversation about representation on the COA for many commercial dairies. Next, it is important to characterize the management characteristics of commercial dairy operators. These results are only characteristic of core operators as this data was not collected for all operators. The COA asked core operators whether their principal occupation was off farm. Overall, a larger share of female core operators had a principal off-farm occupation than male core operators . In California, less than 10% of the male core operators had an off-farm principal occupation, but about 30% of female core operators had an off-farm principal occupation with little variation over time. In other states, like Idaho and Texas, the share of core operators with off-farm principal occupation followed a similar pattern to California by gender. However, there was an 86.6% increase in male core operators with an off-farm principal occupation and an 18% decrease in female core operators in New Mexico. Along a similar thread, a very small portion of female core operators was labeled as principal operators. Now, the definition of a principal operator did change for the 2017 COA, but even with the 2017 addition of more than one core operator being labeled as a principal operator the share of female core operators that are labeled as a principal operator is relatively small. In California, 5% of female core operators are principal operators from 2002to 2012 with a jump in 2017 to 17% with the addition of the fourth core operator . Idaho, New York, and Wisconsin follow a similar pattern as California with little to no change from 2002 to 2012 and a large jump in 2017. New Mexico and Texas, however, had a decrease from 2002 to 2012 and then a large jump in 2017. In 2017, most states had about 16- 20% of female core operators listed as a principal operator, but New Mexico only had 11%.