The soil that remained adhered to the roots after removal from the ground was used to produce the rhizosphere soil samples.The rhizosphere soil was removed from the roots by shaking the root into a whirlpak bag.All samples were immediately transferred to storage at 4uC for shipping back to the laboratory for processing.All root samples were rinsed with alcohol and sterile water before the extraction.DNA was isolated from 0.25 g of soil or root per extraction using standard protocol for PowerSoil DNA Isolation Kit , with the modification of heating the extraction at 65uC for 10 minutes prior to the initial vortex step.The soil physicochemical data was generated by Fruit Growers Laboratory , including total carbon and nitrogen concentrations, pH, salinity, and water content for all samples.Endorhiza, rhizosphere, and bulk soil samples for the second experiment were taken from 6 organically-grown Cannabis plants of two different strains from two locations in August, 2012: Vista and Orange County, California.Triplicate samples were taken from each of the six plants and surrounding rhizosphere , as well as from each of the two bulk soils used in the different locations , totaling 42 samples.In contrast to the first experiment, all samples were taken two weeks prior to harvest.Additionally, triplicate samples from the second experiment were taken from different roots on the same plant.Cannabinoid data was taken from the buds of three White Widow plants and one Mauie Wowie plant.All cannabinoid data was processed at Delta-9-Technologies, LLC.Otherwise, sampling procedure matched the first experiment.Recent literature has suggested a two-step selection model for the endorhiza, where bulk-soil microbial communities are filtered by increased concentration of rhizodeposits, followed by convergent host genotype-dependent selection on endophytic communities.Results from both experiments support many of the expectations produced by this model.Most importantly,dry rack cannabis the principal coordinate analysis plots for the second experiment demonstrate highly significant clustering patterns.
First, soil type is the main determinant of PC1 for the unweighted analysis of the second experiment, revealing that soil is undoubtedly the most important factor in all samples for determining what microbes are present.Second, communities within both soil types demonstrate a similar community shift from bulk soil to endorhiza samples along PC2 , which is dominated by differentiation between sample types.Specifically, endorhiza samples have high, positive values along PC2, rhizosphere samples have intermediate values, and bulk soil samples have more negative values.Third, Cannabis strain is the main determinant of PC1 for the weighted analysis of all samples in the second experiment, suggesting that convergent host genotype-dependent selection acts through controlling community structure more than composition.PCoA results exhibit how all sample types form significantly differentiated clusters in weighted analyses but that only rhizosphere and endorhiza samples form significantly differentiated clusters in unweighted analyses, suggesting niche-filtering of microbes in rhizosphere and endorhiza samples from bulk soil.Furthermore, there were no significant segregating OTUs based on unweighted analysis between cultivars in endorhiza and rhizosphere samples in the second experiment, however there were 71 when abundance was accounted for.This differs greatly from the 657 OTUs that significantly differ between soil types in the same dataset.Testing of the two-step selection model with pairwise comparisons of shared OTUs between endorhiza and bulk soil samples also validated the hypothesis that a portion of the endophytic microbes are inherited and selected from the surrounding soil, showing significantly more OTU overlap between endorhiza and their own bulk soil compared to endorhiza and foreign bulk soil.Given the results from the second experiment strongly suggesting that Cannabis cultivars have important structuring effects on both rhizosphere and endorhiza samples, it may seem troubling that results from the first experiment do not suggest this for the rhizosphere samples.However, differences in Cellvibrio abundance between experiments show that root decay could have diminished the rhizosphere effect, thus diminishing this potential signal.Sampling for the first experiment was done post-harvest, when plant tissues were undergoing senescence and decay, while samples for the second experiment were taken from actively growing plants.Considering the extensive work demonstrating the importance of plant growth stage on the microbiota, as well as the plant-soil feed backs identified in structuring below ground microbial communities, the differences between the first and second experiments are unsurprising.
The similarities, however, are surprising.In particular, that cultivar-specificity could be identified in the microbiota within the endorhiza samples in the first experiment without any input of cultivar-specific metabolites from the living plant for weeks.Although we have presented several highly significant findings supporting expectations of the two-step selection model, some expectations remain to be validated.Specifically, although the mean beta-diversity distances indicate that rhizosphere and endorhiza samples are closer than bulk soil and endorhiza samples, this difference was not significant and thus provides little evidence for the first differentiation step of the two-step selection model.Future work with the Cannabis micro-biome should focus on elucidating the role of cultivar on rhizosphere, as well as what aspects of host genotype are producing the structure observed across Cannabis strains.Increased testing of cannabinoids and decoupling this variation from edaphic factors will improve our understanding of the importance of cannabinoid production in structuring endorhiza communities.Sampling a time series of endorhiza communities across several plants may help us to understand natural variation in the endorhiza during the reproductive cycles of Cannabis.Understanding this natural variation will help direct future mechanistic studies aimed at using microbial communities to increase plant fitness, suppress disease, or augment desired metabolite production.Legalization of cannabis use is increasingly widespread across the United States, but the ramifications are unknown.In 2016 California approved Proposition 64 legalizing recreational cannabis.Unintentional pediatric ingestion is one possible ramification, as occurred in Colorado after legalization in 2009.Regional poison center cases involving marijuana increased by an average 34% per year from 2009 to 2015 in Colorado.During that time, 34% of cases involved self-reported cases of poor product storage.Children who unintentionally ingest cannabis can present with lethargy, ataxia, tachycardia, mydriasis, and hypotonia, which can lead to preventable emergency department visits, invasive workups, and hospital admissions.Despite the institution of safe storage guidelines in Colorado, a recent study found continuing sub-optimal storage practices in that state.5 This trend was mirrored in the use of medical marijuana, as oncology patients and their caregivers reported sub-optimal storage practices and had received little storage education from healthcare providers.
The 2016 California legislation did not include regulations on the safe storage and disposal of cannabis products, creating a potential for similarly unsafe storage practices.The purpose of this study, which was based on a community presenting to a pediatric ED, was to assess the prevalence of cannabis and how it is stored in the home and, secondly, to assess attitudes regarding use of cannabis and storage education among Californians who live in households with children.We conducted a cross-sectional survey with a goal enrollment of 400 adult visitors in an academic pediatric ED in California from June 8–August 16, 2018.During this time, a convenience sample between the hours of 8 am -10 pm was conducted daily in which all adult visitors were screened for eligibility and subsequently approached.The survey was generated and finalized by the investigators and research assistants based on similar studies found during literature review.The survey contained 42 yes-no or Likert-scale questions regarding cannabis use and storage, and education on cannabis storage.Eligible participants were >18 years old and lived in a household with children <18 years old.Participants were excluded if they did not speak English or Spanish, or if the patient was critically ill.Only one survey was administered per household.All participants were notified that their responses were not shared with law enforcement or their care team, and they completed the survey in the absence of a RA.English-speaking participants filled out the survey electronically and submitted their responses directly into Research Electronic Data Capture.Spanish-speaking participants filled out a Spanish-language survey on paper, which was subsequently placed in a lockbox after which these de-identified surveys were uploaded to REDCap weekly.We used descriptive statistics to analyze data.Subjects who were screened but excluded were not tracked during this study.The UC Davis Institutional Review Board approved this study.A 2017 national survey indicated that as many as 11.5% of California adults reported regular cannabis use.Of adults surveyed, 14.5% reported cannabis use in a home with children.Since the legalization of cannabis in California, there has been a steady rise in prevalence of use,7 likely due to increased availability.In our sample, users perceived cannabis to be significantly safer for both adult use and possession inside a home with children, as compared with non-users.Further study is warranted to investigate how the public perceives the risk of cannabis as more time passes since legalization.Currently, little research exists on cannabis storage in homes with children,roll bench and there is no research that describes sources of storage information.Both users and non-users strongly felt that safe storage was important despite poor compliance with safe storage practices.Our results suggest a lack of educational sources regarding safe storage despite 23 years of medical cannabis use in California.The Public Health Department of Colorado set guidelines including locking, hiding, and using child-resistant packaging, yet California does not currently define safe storage.Providing guidelines at a local or state level may provide a reference for cannabis users as well as healthcare providers.
Based on participant responses, cannabis dispensaries may also serve as another point for the distribution of safe storage information.With the increasing prevalence of cannabis use in California,downstream effects on the pediatric population should be further investigated.Healthcare providers in primary care, pediatrics, and the ED should be prepared to screen and educate families on cannabis use and the importance of safe storage in homes with children.Nonpharmaceutical interventions were developed in response to the 2009 H1N1 pandemic and included a protocol to slow the spread of future novel respiratory influenza A virus pandemics.NPIs are strategies for disease control when no pharmaceutical alternative exists and include actions at the personal, environmental, and community level.Specifically, NPIs put in place during the pandemic included travel restriction, restriction of mass gatherings and recommendations for transition to virtual events, social distancing measures and stay-athome orders, closure of non-essential work spaces and schools, and cloth face covering guidance.3,8 However, it is unclear what effect, both intended and unintended, these policy implications have on populations.It is also unclear what effect the pandemic itself has had and will have on populations.A study from the Centers for Disease Control and Prevention examined differences among stay-at-home orders across US states from March 1st to May 31st, 2020, on population movement.Stay-at-home orders were associated with decreased population movement; however, movement increased significantly as states began lifting restrictions.Kaufman et al.reported the initial effect of state variation in social distancing policies and non-essential business closures on COVID-19 rates.Social distancing and closure of non-essential businesses and public schools were shown to reduce daily COVID-19 cases by 15.4% with effects varying across states.10 Finally, Pan et al.showed that there was heterogeneity in NPI domains across the US census region and concluded that states with the most aggressive policies had the highest mitigation of COVID-19 infection.11 While heterogeneity in intensity and duration of state policies on COVID-19 mitigation were demonstrated, all such studies have been restricted to the initial wave of the pandemic and have only assessed the associations of policies on COVID-19 infection spread at the population level.The COVID-19 pandemic and policy interventions such as social distancing, closure of spaces for gathering, and stay-at-home orders may have had varying economic, health, and social effects across different populations.Of particular concern, are negative implications on lesbian, gay, bisexual, transgender, and queer and other sexual minority populations, an already vulnerable, marginalized, and stigmatized group with even larger disparities among racially marginalized communites.The COVID-19 pandemic has highlighted and exacerbated challenges for the LGBTQ community which include rising food and shelter insecurity, economic fallout, job loss, disruption in health care, elevated risk of domestic and family violence, social isolation, increased anxiety, scapegoating/discrimination/stigma, abuse of state power, and concerns about organizational survival.There is an estimated 16 million LGBTQ adults and youth in the United States of which, 5 million work in jobs that are more likely impacted due to the COVID- 19 pandemic.For instance, 15% work in restaurants, 7.5% in hospitals, 7% in K-12 education, 7% in colleges/universities, and 4% in retail, all industries that have been impacted by the pandemic.Moreover, LGBTQ communities before and during the pandemic were more likely to be unemployed, at increased risk of poverty, have issues affording health care, and experience greater workplace discrimination compared to cisgender and straight people.