An assemblage of Frankia Cluster II strains from California contains the canonical nod genes and also the sulfotransferase gene nodH.

BACKGROUND: The ability to establish root nodule symbioses is restricted to four different plant orders. Soil actinobacteria of the genus Frankia can establish a symbiotic relationship with a diverse group of plants within eight different families from three different orders, the Cucurbitales, Fagales and Rosales. Phylogenetically, Frankia strains can be divided into four clusters, three of which (I, II, III) contain symbiotic strains. Members of Cluster II nodulate the broadest range of host plants with species from four families from two different orders, growing on six continents. Two Cluster II genomes were sequenced thus far, both from Asia. RESULTS: In this paper we present the first Frankia cluster II genome from North America (California), Dg2, which represents a metagenome of two major and one minor strains. A phylogenetic analysis of the core genomes of 16 Frankia strains shows that Cluster II the ancestral group in the genus, also ancestral to the non-symbiotic Cluster IV. Dg2 contains the canonical nod genes nodABC for the production of lipochitooligosaccharide Nod factors, but also two copies of the sulfotransferase gene nodH. In rhizobial systems, sulfation of Nod factors affects their host specificity and their stability. CONCLUSIONS: A comparison with the nod gene region of the previously sequenced Dg1 genome from a Cluster II strain from Pakistan shows that the common ancestor of both strains should have contained nodABC and nodH. Phylogenetically, Dg2 NodH proteins are sister to rhizobial NodH proteins. A glnA-based phylogenetic analysis of all Cluster II strains sampled thus far supports the hypothesis that Cluster II Frankia strains came to North America with Datisca glomerata following the Madrean-Tethyan pattern.

Uptake of human pharmaceuticals and personal care products by cabbage (Brassica campestris) from fortified and biosolids-amended soils.

Human pharmaceuticals and personal care products (PPCPs) are routinely found in biosolids from wastewater treatment plants (WWTPs). Once land applied, the PPCPs in biosolids are potentially available for plant uptake and bioaccumulation. This study used a greenhouse model to investigate uptake of PPCPs commonly detected in biosolids by the agricultural plant Chinese cabbage (Brassica campestris). Two series of greenhouse experiments were conducted as part of this project. In the first set of experiments, four pharmaceuticals were added to an organic matter-rich soil in environmentally relevant concentrations based on typical biosolids application rates, resulting in final soil concentrations of 2.6 ng g(-1) carbamazepine, 3.1 ng g(-1) sulfamethoxazole, 5.4 ng g(-1) salbutamol, and 0.5 ng g(-1) trimethoprim. In the second set of experiments, the cabbage was grown in soil amended with an agronomic rate of biosolids from a local WWTP. The ambient concentration of PPCPs in the biosolids resulted in final soil concentrations of 93.1 ng g(-1) carbamazepine, 67.4 ng g(-1) sulfamethoxazole, 30.3 ng g(-1) salbutamol, 433.7 ng g(-1) triclosan, and 24.7 ng g(-1) trimethoprim. After growing to maturity, the aerials of the plants were separated from roots and the two tissue types were analyzed separately. All four human pharmaceuticals were detected in both tissues in the cabbage grown in the soil fortified with the four pharmaceuticals with median concentrations of 255.4 ng g(-1) aerials and 272.9 ng g(-1) roots carbamazepine; 222.8 ng g(-1) aerials and 260.3 ng g(-1) roots sulfamethoxazole; 108.3 ng g(-1) aerials and 140.6 ng g(-1) roots salbutamol; and 20.6 ng g(-1) aerials and 53.7 ng g(-1) roots trimethoprim. Although all study compounds were present in the biosolids-amended planting soil, only carbamazepine (317.6 ng g(-1) aerials and 416.2 ng g(-1) roots), salbutamol (21.2 ng g(-1) aerials and 187.6 ng g(-1) roots), and triclosan (22.9 ng g(-1) aerials and 1220.1 ng g(-1) roots) were detected in the aerials of the cabbage. In addition to the study compounds detected in the aerials, sulfamethoxazole was detected in the roots of one of the plants in the biosolid-amended soil. In comparison to many previous studies that have utilized PPCP concentration that exceed environmentally relevant concentrations, plants in this study were exposed to environmentally relevant concentrations of the PPCPs, yet resulted in uptake concentrations similar to or greater than those reported in comparable studies. We suggest that rhizosphere conditions, particularly the presence of dissolved organic matter in the planting matrix, might be one of the critical factors determining mobilization and bioavailability of xenobiotic compounds such as PPCPs.

Effects of short-term environmental stresses on the onset of cannabinoid production in young immature flowers of industrial hemp (Cannabis sativa L.).

BACKGROUNDS: Cannabis sativa L. produces at least 120 cannabinoids. Although genetic variation is the main factor in cannabinoid production, the effects of short-term environmental stresses in the early flowering stage remains largely unknown. METHODS: To investigate the effects of short-term environmental stresses on the onset of cannabinoid production in young immature flowers, a hemp variety, Green-Thunder (5-8% CBD/mg of dry weight), was treated with mechanical damage, insect herbivory, extreme heat, or drought stress for 5-7 days during the first 2 weeks of flowering. Three hemp tissues, including flowers, leaves, and stems, were collected from hemp grown under these stress conditions at multiple time points during the first 2 weeks after transition to the short photoperiod and analyzed using high pressure liquid chromatography to quantify phytocannabinoids including cannabigerolic acid (CBGA), cannabigerol (CBG), cannabidiolic acid (CBDA), cannabidiol (CBD), Δ-tetrahydrocannabinolic acid (THCA), Δ-tetrahydrocannabinol (THC), and cannabinol (CBN). RESULTS: The 5 days of mechanical wounding did not affect the production of any of the cannabinoids during the initial stage of flowering. However, after 5 days of herbivore treatment, there was a significant difference in concentration between day 1 and day 6 of CBGA (control: 308 µg/g; treatment - 24 µg/g), CBG (control: 69 µg/g; treatment: 52 µg/g), and CBD (control: 755 µg/g; treatment: 194 µg/g) between the control and treatment plants. The 7 days of heat treatment at 45-50 oC significantly reduced the production of CBGA during this observed window (control: 206 µg/g; treatment: 182 µg/g) and CBG (control: 21 µg/g; treatment: - 112 µg/g). Notably, the largest change was observed after 7 days of drought stress, when plants showed a 40% greater accumulation of CBG (control: 336 µg/g; treatment: 622 µg/g), and a significant decrease (70-80%) in CBD (control: 1182 µg/g; treatment: 297 µg/g) and THC amounts (control: 3927 µg/g; treatment: 580 µg/g). CONCLUSIONS: Although this observation is limited in the early flowering stage, the common field stresses are adequate to induce changes in the cannabinoid profiles, particularly drought stress being the most impactful stress for hemp flower initiation with the altering the cannabinoid production by decreasing CBD and THC accumulation while increasing CBG by 40%.

Genome sequence of "Candidatus Frankia datiscae" Dg1, the uncultured microsymbiont from nitrogen-fixing root nodules of the dicot Datisca glomerata.

Members of the noncultured clade of Frankia enter into root nodule symbioses with actinorhizal species from the orders Cucurbitales and Rosales. We report the genome sequence of a member of this clade originally from Pakistan but obtained from root nodules of the American plant Datisca glomerata without isolation in culture.

Cannabis Contaminants Limit Pharmacological Use of Cannabidiol.

For nearly a century, Cannabis has been stigmatized and criminalized across the globe, but in recent years, there has been a growing interest in Cannabis due to the therapeutic potential of phytocannabinoids. With this emerging interest in Cannabis, concerns have arisen about the possible contaminations of hemp with pesticides, heavy metals, microbial pathogens, and carcinogenic compounds during the cultivation, manufacturing, and packaging processes. This is of particular concern for those turning to Cannabis for medicinal purposes, especially those with compromised immune systems. This review aims to provide types of contaminants and examples of Cannabis contamination using case studies that elucidate the medical consequences consumers risk when using adulterated Cannabis products. Thus, it is imperative to develop universal standards for cultivation and testing of products to protect those who consume Cannabis.

Cannabidiol Drugs Clinical Trial Outcomes and Adverse Effects.

This review aims to present completed clinical trial data surrounding the medicinal benefits and potential side effects of the increasingly popular cannabidiol (CBD)-based drug products, specifically Epidiolex. The article is divided into two sections based on if the ailment being treated by this cannabinoid is classified as either physiological or neurological conditions. In addition to describing the current status, we also examined the different primary and secondary outcomes recorded for each study, which varies greatly depending on the funding source of the clinical trial. With the recent FDA-approval of Epidiolex, this review mainly focused on trials involving this specific formulation since it is the only CBD-based drug currently available to clinicians, although all other clinically trialed CBD(A) drugs were also examined. We hope this review will help guide future research and clinical trials by providing the various outcomes measured in a single review.

Candidatus Frankia Datiscae Dg1, the Actinobacterial Microsymbiont of Datisca glomerata, Expresses the Canonical nod Genes nodABC in Symbiosis with Its Host Plant.

Frankia strains are nitrogen-fixing soil actinobacteria that can form root symbioses with actinorhizal plants. Phylogenetically, symbiotic frankiae can be divided into three clusters, and this division also corresponds to host specificity groups. The strains of cluster II which form symbioses with actinorhizal Rosales and Cucurbitales, thus displaying a broad host range, show suprisingly low genetic diversity and to date can not be cultured. The genome of the first representative of this cluster, Candidatus Frankia datiscae Dg1 (Dg1), a microsymbiont of Datisca glomerata, was recently sequenced. A phylogenetic analysis of 50 different housekeeping genes of Dg1 and three published Frankia genomes showed that cluster II is basal among the symbiotic Frankia clusters. Detailed analysis showed that nodules of D. glomerata, independent of the origin of the inoculum, contain several closely related cluster II Frankia operational taxonomic units. Actinorhizal plants and legumes both belong to the nitrogen-fixing plant clade, and bacterial signaling in both groups involves the common symbiotic pathway also used by arbuscular mycorrhizal fungi. However, so far, no molecules resembling rhizobial Nod factors could be isolated from Frankia cultures. Alone among Frankia genomes available to date, the genome of Dg1 contains the canonical nod genes nodA, nodB and nodC known from rhizobia, and these genes are arranged in two operons which are expressed in D. glomerata nodules. Furthermore, Frankia Dg1 nodC was able to partially complement a Rhizobium leguminosarum A34 nodC::Tn5 mutant. Phylogenetic analysis showed that Dg1 Nod proteins are positioned at the root of both α- and ß-rhizobial NodABC proteins. NodA-like acyl transferases were found across the phylum Actinobacteria, but among Proteobacteria only in nodulators. Taken together, our evidence indicates an Actinobacterial origin of rhizobial Nod factors.

Uptake of human pharmaceuticals by plants grown under hydroponic conditions.

Cabbage (Brassica rapa var. pekinensis) and Wisconsin Fast Plants (Brassica rapa) were chosen for a proof of concept study to determine the potential uptake and accumulation of human pharmaceuticals by plants. These plants were grown hydroponically under high-pressure sodium lamps in one of two groups including a control and test group exposed to pharmaceuticals. The control plants were irrigated with a recirculating Hoagland's nutrient solution while the test plants were irrigated with a Hoagland's nutrient solution fortified with the pharmaceuticals carbamazepine, salbutamol, sulfamethoxazole, and trimethoprim at 232.5 microg L(-1). When plants reached maturity, nine entire plants of each species were separated into components such as roots, leaves, stems, and seedpods where applicable. An analytical method for quantifying pharmaceuticals and personal care products was developed using pressurized liquid extraction and liquid chromatography electrospray ionization mass spectrometry (LC/ESI/MS) in positive and negative ion modes using single ion monitoring. The method detection limits ranged from 3.13 ng g(-1) to 29.78 ng g(-1) with recoveries ranging from 66.83% to 113.62% from plant matrices. All four of the pharmaceuticals were detected in the roots and leaves of the cabbage. The maximum wet weight concentrations of the pharmaceuticals were detected in the root structure of the cabbage plants at 98.87 ng g(-1) carbamazepine, 114.72 ng g(-1) salbutamol, 138.26 ng g(-1) sulfamethoxazole, and 91.33 ng g(-1) trimethoprim. Carbamazepine and salbutamol were detected in the seedpods of the Wisconsin Fast Plants while all four of the pharmaceuticals were detected in the leaf/stem/root of the Wisconsin Fast Plants. Phloroglucinol staining of root cross-sections was used to verify the existence of an intact endodermis, suggesting that pharmaceuticals found in the leaf and seedpods of the plants were transported symplastically.

Low genetic diversity among Frankia spp. strains nodulating sympatric populations of actinorhizal species of Rosaceae, Ceanothus (Rhamnaceae) and Datisca glomerata (Datiscaceae) west of the Sierra Nevada (California).

Frankia spp. strains typically induce N2-fixing root nodules on actinorhizal plants. The majority of host plant taxa associated with the uncultured Group 1 Frankia strains, i.e., Ceanothus of the Rhamnaceae, Datisca glomerata (Datiscaceae), and all actinorhizal members of the Rosaceae except Dryas, are found in California. A study was conducted to determine the distribution of Frankia strains among root nodules collected from both sympatric and solitary stands of hosts. Three DNA regions were examined, the 5' end of the 16S rRNA gene, the internal transcribed spacer region between the 16S and 23S rRNA genes, and a portion of the glutamine synthetase gene (glnA). The results suggest that a narrow range of Group 1 Frankia spp. strains dominate in root nodules collected over a large area of California west of the Sierra Nevada crest with no apparent host-specificity. Comparisons with Group 2 Frankia strain diversity from Alnus and Myrica within the study range suggest that the observed low diversity is peculiar to Group 1 Frankia strains only. Factors that may account for the observed lack of genetic variability and host specificity include strain dominance over a large geographical area, current environmental selection, and (or) a past evolutionary bottleneck.