Increasing lifetime and past 30-day marijuana use among middle school students regardless of recreational marijuana sales.

PURPOSE: This study investigated whether adult use marijuana sales were associated with changes in lifetime and past 30-day (P30D) marijuana use among middle school students in Nevada (NV), which had adult-use marijuana sales during the study period, compared to New Mexico (NM), which did not have adult-use marijuana sales during the study period. METHODS: Data were drawn from the middle school 2017 and 2019 NV Youth Risk Behavior and NM Youth Risk and Resiliency Surveys. Difference-in-difference analyses compare changes in lifetime and P30D marijuana use in NV (adult-use sales implemented July 2017) vs. NM (no adult-use sales during the study period). RESULTS: There was no difference in lifetime (aOR 1.11; 95% CI 0.91,1.36) and P30D (aOR 1.17; 95% CI 0.91,1.51) marijuana use by adult-use sales status. The odds of lifetime and P30D marijuana use increased in both states, particularly among students who were female, older, non-White, or attending a Title 1 school. DISCUSSION: Adult-use sales were not associated with an increase in lifetime or P30D marijuana use. State-level prevention efforts should focus on sub-populations with increasing lifetime and P30D use regardless of adult-use sales status.

Diagnostic colonoscopy completion after abnormal fecal immunochemical testing and quality of tests used at 8 Federally Qualified Health Centers in Southern California: Opportunities for improving screening outcomes.

BACKGROUND: The effectiveness of colorectal cancer screening with fecal immunochemical tests (FITs) of stool blood depends on high rates of colonoscopy follow-up for abnormal FITs and the use of high-quality tests. This study characterized colonoscopy referral and completion among patients with abnormal FITs and the types of FITs implemented in a sample of Southern California Federally Qualified Health Centers (FQHCs). METHODS: FQHCs in San Diego, Imperial, and Los Angeles Counties were invited to define a cohort of ≥150 consecutive patients with abnormal FITs in 2015-2016 and to provide data on sex, insurance status, diagnostic colonoscopy referrals and completion within 6 months of abnormal FITs, and the types (brands) of FITs implemented. The primary outcomes were the proportions with colonoscopy referrals and completion for all patients at each FQHC and in aggregate. RESULTS: Eight FQHCs provided data for 1229 patients with abnormal FITs; 46% were male, and 20% were uninsured. Among patients with abnormal FITs, 89% (1091 of 1229; 95% confidence interval [CI], 0.87-0.91) had a colonoscopy referral, and 44% (539 of 1229; 95% CI, 0.41-0.47) had colonoscopy completion. Across FQHCs, the range for colonoscopy referral was 73% to 96%, and the range for completion was 18% to 57%. Six of the 8 FQHCs (75%) reported FIT brands with limited data to support their effectiveness. CONCLUSIONS: In a sample of Southern California FQHCs, diagnostic colonoscopy completion after abnormal FITs was substantially below the nationally recommended benchmark to achieve 80% completion, and the use of FIT brands with limited data to support their effectiveness was high. These findings suggest a need for policies and multilevel interventions to promote diagnostic colonoscopy among individuals with abnormal FITs and the use of higher quality FITs.

Non-excitable fluorescent protein orthologs found in ctenophores.

BACKGROUND: Fluorescent proteins are optically active proteins found across many clades in metazoans. A fluorescent protein was recently identified in a ctenophore, but this has been suggested to derive from a cnidarian, raising again the question of origins of this group of proteins. RESULTS: Through analysis of transcriptome data from 30 ctenophores, we identified a member of an orthologous group of proteins similar to fluorescent proteins in each of them, as well as in the genome of Mnemiopsis leidyi. These orthologs lack canonical residues involved in chromophore formation, suggesting another function. CONCLUSIONS: The phylogenetic position of the ctenophore protein family among fluorescent proteins suggests that this gene was present in the common ancestor of all ctenophores and that the fluorescent protein previously found in a ctenophore actually derives from a siphonophore.

Repeated mild traumatic brain injury causes focal response in lateral septum and hippocampus.

AIM: To advance our understanding of regional and temporal cellular responses to repeated mild traumatic brain injury (rmTBI), we used a mouse model of rmTBI that incorporated acceleration, deceleration and rotational forces. MATERIALS & METHODS: A modified weight-drop method was used to compare two inter-injury intervals, rmTBI-short (five hits delivered over 3 days) and rmTBI-long (five hits delivered over 15 days). Regional investigations of forebrain and midbrain histological alterations were performed at three post-injury time points (immediate, 2 weeks and 6 weeks). RESULTS: The rmTBI-short protocol generated an immediate, localized microglial and astroglial response in the dorsolateral septum and hippocampus, with the astroglial response persisting in the dorsolateral septum. The rmTBI-long protocol showed only a transitory astroglial response in the dorsolateral septum. CONCLUSION: Our results indicate that the lateral septum and hippocampus are particularly vulnerable regions in rmTBI, possibly contributing to memory and emotional impairments associated with repeated concussions.

Occurrence of Isopenicillin-N-Synthase Homologs in Bioluminescent Ctenophores and Implications for Coelenterazine Biosynthesis.

The biosynthesis of the luciferin coelenterazine has remained a mystery for decades. While not all organisms that use coelenterazine appear to make it themselves, it is thought that ctenophores are a likely producer. Here we analyze the transcriptome data of 24 species of ctenophores, two of which have published genomes. The natural precursors of coelenterazine have been shown to be the amino acids L-tyrosine and L-phenylalanine, with the most likely biosynthetic pathway involving cyclization and further modification of the tripeptide Phe-Tyr-Tyr ("FYY"). Therefore, we searched the ctenophore transcriptome data for genes with the short peptide "FYY" as part of their coding sequence. We recovered a group of candidate genes for coelenterazine biosynthesis in the luminous species which encode a set of highly conserved non-heme iron oxidases similar to isopenicillin-N-synthase. These genes were absent in the transcriptomes and genome of the two non-luminous species. Pairwise identities and substitution rates reveal an unusually high degree of identity even between the most unrelated species. Additionally, two related groups of non-heme iron oxidases were found across all ctenophores, including those which are non-luminous, arguing against the involvement of these two gene groups in luminescence. Important residues for iron-binding are conserved across all proteins in the three groups, suggesting this function is still present. Given the known functions of other members of this protein superfamily are involved in heterocycle formation, we consider these genes to be top candidates for laboratory characterization or gene knockouts in the investigation of coelenterazine biosynthesis.

Characterization of an anthraquinone fluor from the bioluminescent, pelagic polychaete Tomopteris.

Tomopteris is a cosmopolitan genus of polychaetes. Many species produce yellow luminescence in the parapodia when stimulated. Yellow bioluminescence is rare in the ocean, and the components of this luminescent reaction have not been identified. Only a brief description, half a century ago, noted fluorescence in the parapodia with a remarkably similar spectrum to the bioluminescence, which suggested that it may be the luciferin or terminal light-emitter. Here, we report the isolation of the fluorescent yellow-orange pigment found in the luminous exudate and in the body of the animals. Liquid chromatography-mass spectrometry revealed the mass to be 270 m/z with a molecular formula of C(15)H(10)O(5), which ultimately was shown to be aloe-emodin, an anthraquinone previously found in plants. We speculate that aloe-emodin could be a factor for resonant-energy transfer or the oxyluciferin for Tomopteris bioluminescence.

A comparison across non-model animals suggests an optimal sequencing depth for de novo transcriptome assembly.

BACKGROUND: The lack of genomic resources can present challenges for studies of non-model organisms. Transcriptome sequencing offers an attractive method to gather information about genes and gene expression without the need for a reference genome. However, it is unclear what sequencing depth is adequate to assemble the transcriptome de novo for these purposes. RESULTS: We assembled transcriptomes of animals from six different phyla (Annelids, Arthropods, Chordates, Cnidarians, Ctenophores, and Molluscs) at regular increments of reads using Velvet/Oases and Trinity to determine how read count affects the assembly. This included an assembly of mouse heart reads because we could compare those against the reference genome that is available. We found qualitative differences in the assemblies of whole-animals versus tissues. With increasing reads, whole-animal assemblies show rapid increase of transcripts and discovery of conserved genes, while single-tissue assemblies show a slower discovery of conserved genes though the assembled transcripts were often longer. A deeper examination of the mouse assemblies shows that with more reads, assembly errors become more frequent but such errors can be mitigated with more stringent assembly parameters. CONCLUSIONS: These assembly trends suggest that representative assemblies are generated with as few as 20 million reads for tissue samples and 30 million reads for whole-animals for RNA-level coverage. These depths provide a good balance between coverage and noise. Beyond 60 million reads, the discovery of new genes is low and sequencing errors of highly-expressed genes are likely to accumulate. Finally, siphonophores (polymorphic Cnidarians) are an exception and possibly require alternate assembly strategies.

Expression and characterization of the calcium-activated photoprotein from the ctenophore Bathocyroe fosteri: insights into light-sensitive photoproteins.

Calcium-binding photoproteins have been discovered in a variety of luminous marine organisms [1]. Recent interest in photoproteins from the phylum Ctenophora has stemmed from cloning and expression of several photoproteins from this group [2-5]. Additional characterization has revealed unique biochemical properties found only in ctenophore photoproteins, such as inactivation by light. Here we report the cloning, expression, and characterization of the photoprotein responsible for luminescence in the deep-sea ctenophore Bathocyroe fosteri. This animal was of particular interest due to the unique broad color spectrum observed in live specimens [6]. Full-length sequences were identified by BLAST searches of known photoprotein sequences against Bathocyroe transcripts obtained from 454 sequencing. Recombinantly expressed Bathocyroe photoprotein (BfosPP) displayed an optimal coelenterazine-loading pH of 8.5, and produced calcium-triggered luminescence with peak wavelengths closely matching the 493 nm peak observed in the spectrum of live B. fosteri specimens. Luminescence from recombinant BfosPP was inactivated most efficiently by UV and blue light. Primary structure alignment of BfosPP with other characterized photoproteins showed very strong sequence similarity to other ctenophore photoproteins and conservation of EF-hand motifs. Both alignment and structural prediction data provide more insight into the formation of the coelenterazine-binding domain and the probable mechanism of photoinactivation.

Genomic organization, evolution, and expression of photoprotein and opsin genes in Mnemiopsis leidyi: a new view of ctenophore photocytes.

BACKGROUND: Calcium-activated photoproteins are luciferase variants found in photocyte cells of bioluminescent jellyfish (Phylum Cnidaria) and comb jellies (Phylum Ctenophora). The complete genomic sequence from the ctenophore Mnemiopsis leidyi, a representative of the earliest branch of animals that emit light, provided an opportunity to examine the genome of an organism that uses this class of luciferase for bioluminescence and to look for genes involved in light reception. To determine when photoprotein genes first arose, we examined the genomic sequence from other early-branching taxa. We combined our genomic survey with gene trees, developmental expression patterns, and functional protein assays of photoproteins and opsins to provide a comprehensive view of light production and light reception in Mnemiopsis. RESULTS: The Mnemiopsis genome has 10 full-length photoprotein genes situated within two genomic clusters with high sequence conservation that are maintained due to strong purifying selection and concerted evolution. Photoprotein-like genes were also identified in the genomes of the non-luminescent sponge Amphimedon queenslandica and the non-luminescent cnidarian Nematostella vectensis, and phylogenomic analysis demonstrated that photoprotein genes arose at the base of all animals. Photoprotein gene expression in Mnemiopsis embryos begins during gastrulation in migrating precursors to photocytes and persists throughout development in the canals where photocytes reside. We identified three putative opsin genes in the Mnemiopsis genome and show that they do not group with well-known bilaterian opsin subfamilies. Interestingly, photoprotein transcripts are co-expressed with two of the putative opsins in developing photocytes. Opsin expression is also seen in the apical sensory organ. We present evidence that one opsin functions as a photopigment in vitro, absorbing light at wavelengths that overlap with peak photoprotein light emission, raising the hypothesis that light production and light reception may be functionally connected in ctenophore photocytes. We also present genomic evidence of a complete ciliary phototransduction cascade in Mnemiopsis. CONCLUSIONS: This study elucidates the genomic organization, evolutionary history, and developmental expression of photoprotein and opsin genes in the ctenophore Mnemiopsis leidyi, introduces a novel dual role for ctenophore photocytes in both bioluminescence and phototransduction, and raises the possibility that light production and light reception are linked in this early-branching non-bilaterian animal.

Changes in reflectin protein phosphorylation are associated with dynamic iridescence in squid.

Many cephalopods exhibit remarkable dermal iridescence, a component of their complex, dynamic camouflage and communication. In the species Euprymna scolopes, the light-organ iridescence is static and is due to reflectin protein-based platelets assembled into lamellar thin-film reflectors called iridosomes, contained within iridescent cells called iridocytes. Squid in the family Loliginidae appear to be unique in which the dermis possesses a dynamic iridescent component with reflective, coloured structures that are assembled and disassembled under the control of the muscarinic cholinergic system and the associated neurotransmitter acetylcholine (ACh). Here we present the sequences and characterization of three new members of the reflectin family associated with the dynamically changeable iridescence in Loligo and not found in static Euprymna iridophores. In addition, we show that application of genistein, a protein tyrosine kinase inhibitor, suppresses ACh- and calcium-induced iridescence in Loligo. We further demonstrate that two of these novel reflectins are extensively phosphorylated in concert with the activation of iridescence by exogenous ACh. This phosphorylation and the correlated iridescence can be blocked with genistein. Our results suggest that tyrosine phosphorylation of reflectin proteins is involved in the regulation of dynamic iridescence in Loligo.