Lactococcus lactis MA5 is a potential autochthonous probiotic for nutrient digestibility enhancement and bacterial pathogen inhibition in hybrid catfish (Ictalurus punctatus × I. furcatus).

With the emergence of diseases, the U.S. catfish industry is under challenge. Current trends prefer autochthonous bacteria as potential probiotic candidates owing to their adaptability and capacity to effectively colonize the host's intestine, which can enhance production performance and bolster disease resistance. The objective of this study was to isolate an autochthonous bacterium as probiotic for hybrid catfish. Initially, an analysis of the intestinal microbiota of hybrid catfish reared in earthen ponds was conducted for subsequent probiotic development. Twenty lactic acid bacteria were isolated from the digesta of overperforming catfish, and most of the candidates demonstrated probiotic traits, including proteolytic and lipolytic abilities; antagonistic inhibition of catfish enteric bacterial pathogens, negative haemolytic activity and antibiotic susceptibility. Subsequent to this screening process, an isolate of Lactococcus lactis (MA5) was deemed the most promising probiotic candidate. In silico analyses were conducted, and several potential probiotic functions were predicted, including essential amino acids and vitamin synthesis. Moreover, genes for three bacteriocins, lactococcin A, enterolysin A and sactipeptide BmbF, were identified. Lastly, various protectant media for lyophilization of MA5 were assessed. These findings suggest that Lactococcus lactis MA5 can be an autochthonous probiotic from hybrid catfish, holding promise to be further tested in feeding trials.

Bringing COVID-19 vaccine to medically underserved populations utilizing a community-guided mobile vaccine program.

Between March 23, 2021, and December 31, 2022, the Mobile Vaccine Program (MVP) vaccinated 5044 individuals from medically-underserved communities in Middle Tennessee identified through and guided by a collaboration of local community agencies. The primary objective of the MVP was to vaccinate individuals for COVID-19 who had barriers to traditional mass vaccine strategies through community-guided strategies and partnerships. Three strategies were developed and implemented with community partners and their affiliated community health workers (CHWs). The strategies included pop-up vaccination clinics at community partner events, CHW-guided door-to-door in-home vaccination, and community partner-initiated homebound referrals for vaccination.

Microbial succession from a subsequent secondary death event following mass mortality.

BACKGROUND: Each death event can be characterized by its associated microbes - a living community of bacteria composed of carcass, soil, and insect-introduced bacterial species - a necrobiome. With the possibility for close succession of these death events, it may be beneficial to characterize how the magnitude of an initial death event may impact the decomposition and necrobiomes of subsequent death events in close proximity. In this paper we hope to characterize the microbial communities associated with a proximate subsequent death event, and distinguish any changes within those communities based on the magnitude of an initial death event and the biomass of preexisting carcass (es) undergoing decomposition. For this experiment, 6 feral swine carcasses in containers were placed in the vicinity of preexisting and ongoing carcass decomposition at sites of three different scales of decomposing carcass biomass. Swab samples were collected from the skin and eye sockets of the container pigs and subjected to 16 s rRNA sequencing and OTU assignment. RESULTS: PERMANOVA analysis of the bacterial taxa showed that there was no significant difference in the bacterial communities based on initial mortality event biomass size, but we did see a change in the bacterial communities over time, and slight differences between the skin and ocular cavity communities. Even without soil input, necrobiome communities can change rapidly. Further characterization of the bacterial necrobiome included utilization of the Random Forest algorithm to identify the most important predictors for time of decomposition. Sample sets were also scanned for notable human and swine-associated pathogens. CONCLUSIONS: The applications from this study are many, ranging from establishing the environmental impacts of mass mortality events to understanding the importance of scavenger, and scavenger microbial community input on decomposition.

Results of a Doravirine-Atorvastatin Drug-Drug Interaction Study.

Doravirine is a novel, highly potent, nonnucleoside reverse transcriptase inhibitor that is administered once daily and that is in development for the treatment of HIV-1 infection. In vitro and clinical data suggest that doravirine is unlikely to cause significant drug-drug interactions via major drug-metabolizing enzymes or transporters. As a common HIV-1 infection comorbidity, hypercholesterolemia is often treated with statins, including the commonly prescribed atorvastatin. Atorvastatin is subject to drug-drug interactions with cytochrome P450 3A4 (CYP3A4) inhibitors. Increased exposure due to CYP3A4 inhibition may lead to serious adverse events (AEs), including rhabdomyolysis. Furthermore, atorvastatin is a substrate for breast cancer resistance protein (BCRP), of which doravirine may be a weak inhibitor; this may increase atorvastatin exposure. The potential of doravirine to affect atorvastatin pharmacokinetics was investigated in a two-period, fixed-sequence study in healthy individuals. In period 1, a single dose of atorvastatin at 20 mg was administered followed by a 72-h washout. In period 2, doravirine at 100 mg was administered once daily for 8 days, with a single dose of atorvastatin at 20 mg concomitantly being administered on day 5. Sixteen subjects were enrolled, and 14 completed the trial; 2 discontinued due to AEs unrelated to the treatment. The atorvastatin area under the curve from time zero to infinity was similar with and without doravirine (geometric mean ratio [GMR] for doravirine-atorvastatin/atorvastatin, 0.98; 90% confidence interval [CI], 0.90 to 1.06), while the maximum concentration decreased by 33% (GMR for doravirine-atorvastatin/atorvastatin, 0.67; 90% CI, 0.52 to 0.85). These changes were deemed not to be clinically meaningful. Both of the study drugs were generally well tolerated. Doravirine had no clinically relevant effect on atorvastatin pharmacokinetics in healthy subjects, providing support for the coadministration of doravirine and atorvastatin.

Geographic Variation of Amyotrophic Lateral Sclerosis Incidence in New Jersey, 2009-2011.

Few analyses in the United States have examined geographic variation and socioeconomic disparities in amyotrophic lateral sclerosis (ALS) incidence, because of lack of population-based incidence data. In this analysis, we used population-based ALS data to identify whether ALS incidence clusters geographically and to determine whether ALS risk varies by area-based socioeconomic status (SES). This study included 493 incident ALS cases diagnosed (via El Escorial criteria) in New Jersey between 2009 and 2011. Geographic variation and clustering of ALS incidence was assessed using a spatial scan statistic and Bayesian geoadditive models. Poisson regression was used to estimate the associations between ALS risk and SES based on census-tract median income while controlling for age, sex, and race. ALS incidence varied across and within counties, but there were no statistically significant geographic clusters. SES was associated with ALS incidence. After adjustment for age, sex, and race, the relative risk of ALS was significantly higher (relative risk (RR) = 1.37, 95% confidence interval (CI): 1.02, 1.82) in the highest income quartile than in the lowest. The relative risk of ALS was significantly lower among blacks (RR = 0.57, 95% CI: 0.39, 0.83) and Asians (RR = 0.63, 95% CI: 0.41, 0.97) than among whites. Our findings suggest that ALS incidence in New Jersey appears to be associated with SES and race.

Effects of demographic factors on survival time after a diagnosis of amyotrophic lateral sclerosis.

BACKGROUND: The Agency for Toxic Substances and Disease Registry established surveillance projects to determine the incidence, prevalence, and demographic characteristics of persons with Amyotrophic Lateral Sclerosis (ALS) in defined geographic areas. There is a need to characterize and account for the survival and prognostic factors among a population-based cohort of ALS cases in the United States. METHODS: A cohort of incident cases diagnosed from 2009-2011 in New Jersey was followed until death or December 31, 2013, whichever happened first. Survival was assessed using Kaplan-Meier curves and Cox proportional hazards regression was used to identify prognostic factors. RESULTS: Sixty-four percent of incident cases died between 2009 and 2013, 93.7% specifically from ALS. Among the 456 cases studied in the survival analysis, the median survival from diagnosis was 21 months; 46% of cases survived longer than two years from diagnosis. Older age predicted shorter survival. While there is some indication of differences because of sex, race, and ethnicity, these differences were not statistically significant when accounting for age. CONCLUSIONS: New Jersey mortality data were queried to determine the vital status of a cohort of incident ALS cases and used to investigate relationships between demographic factors and survival. Results are consistent with other population-based studies. Older age was a strong predictor of shorter survival time. Additional follow-up time is needed to characterize longer-term survival.

Amyotrophic lateral sclerosis surveillance in Baltimore and Philadelphia.

INTRODUCTION: Limited epidemiological data on amyotrophic lateral sclerosis (ALS) exist in defined geographic areas in the United States. METHODS: Neurologists submitted case reports for patients under their care between January 1, 2009, and December 31, 2011, who met the El Escorial criteria. Diagnosis was confirmed for a sample of cases by the consulting neurologist. Death certificate data were used for supplemental case identification. RESULTS: The 248 reported cases were most likely to be 50-69 years old, men, white, and non-Hispanic. The total crude average annual incidence rate was 1.46 per 100,000 person-years. CONCLUSIONS: The reported demographic characteristics were consistent with previously published findings. The crude annual incidence was slightly lower than the expected rate of 1.6 but was within the range reported previously (0.7-2.5). These findings help quantify the burden of ALS in the United States.

Population-based surveillance of amyotrophic lateral sclerosis in New Jersey, 2009-2011.

BACKGROUND: Limited epidemiological data exist about amyotrophic lateral sclerosis (ALS) in the United States (US). The Agency for Toxic Substances and Disease Registry maintains the National ALS Registry and funded state and metropolitan surveillance projects to obtain reliable, timely information about ALS in defined geographic areas. METHODS: Neurologists submitted case reports for ALS patients under their care between January 1, 2009 and December 31, 2011 who were New Jersey residents. A medical record verification form and electromyogram (EMG) report were requested for a sample of case reports. Incidence rates were standardized to the 2000 US Standard Population. RESULTS: The average crude annual incidence rate was 1.87 per 100,000 person-years, the average age-adjusted annual incidence rate was 1.67 per 100,000 person-years, and the point prevalence rate on December 31, 2011 was 4.40 per 100,000 persons. Average annual incidence rates and point prevalence rates were statistically higher for men compared with women; Whites compared with Blacks/African Americans and Asians; and non-Hispanics compared with Hispanics. CONCLUSIONS: The project findings contribute new, population-based, state-specific information to epidemiological data regarding ALS. The findings are generally consistent with previously published surveillance studies conducted in the US and abroad.

Identifying a distractor produces object-based inhibition in an allocentric reference frame for saccade planning.

We investigated whether distractor inhibition occurs relative to the target or fixation in a perceptual decision-making task using a purely saccadic response. Previous research has shown that during the process of discriminating a target from distractor, saccades made to a target deviate towards the distractor. Once discriminated, the distractor is inhibited, and trajectories deviate away from the distractor. Saccade deviation magnitudes provide a sensitive measure of target-distractor competition dependent on the distance between them. While saccades are planned in an egocentric reference frame (locations represented relative to fixation), object-based inhibition has been shown to occur in an allocentric reference frame (objects represented relative to each other independent of fixation). By varying the egocentric and allocentric distances of the target and distractor, we found that only egocentric distances contributed to saccade trajectories shifts towards the distractor during active decision-making. When the perceptual decision-making process was complete, and the distractor was inhibited, both ego- and allocentric distances independently contributed to saccade trajectory shifts away from the distractor. This is consistent with independent spatial and object-based inhibitory mechanisms. Therefore, we suggest that distractor inhibition is maintained in cortical visual areas with allocentric maps which then feeds into oculomotor areas for saccade planning.

Target-Distractor Competition Modulates Saccade Trajectories in Space and Object Space.

Saccade planning and execution can be affected by a multitude of factors present in a target selection task. Recent studies have shown that the similarity between a target and nearby distractors affects the curvature of saccade trajectories, because of target-distractor competition. To further understand the nature of this competition, we varied the distance between and the similarity of complex target and distractor objects in a delayed match-to-sample task to examine their effects on human saccade trajectories and better understand the underlying neural circuitry. For trials with short saccadic reaction times (SRTs) when target-distractor competition is still active, the distractor is attractive and saccade trajectories are deviated toward the distractor. We found a robust effect of distance consistent with saccade vector averaging, whereas the effect of similarity suggested the existence of an object-based suppressive surround. At longer SRTs, there was sufficient time for competition between the objects to complete and the distractor to be repulsive, which resulted in saccade trajectory deviations away from the distractor exhibiting the effects of a spatial suppressive surround. In terms of similarity, as the target-distractor similarity decreased, the initial saccade angle shifted toward the target, reflecting stronger distractor inhibition. There were no interactions between distance and similarity at any point in the time course of target-distractor competition. Together, saccade trajectories reflect target-distractor competition that is affected independently by both spatial and object space suppressive surrounds. The differences in saccade trajectories at short and long SRTs distinguish between active and completed decision-making processes.

Quorum sensing gene regulation in Staphylococcus epidermidis reduces the attraction of Aedes aegypti (L.) (Diptera: Culicidae).

Introduction: Identifying mechanisms regulating mosquito attraction to hosts is key to suppressing pathogen transmission. Historically, the ecology of the host microbial community and its influence on mosquito attraction, specifically, whether bacterial communication through quorum sensing (QS) modulates VOC production that affects mosquito behavior have not been extensively considered. Methods: Behavioral choice assays were applied along with volatile collection, followed by GC-MS and RNA transcriptome analyses of bacteria with and without a quorum-sensing inhibitor, furanone C-30. Results: Utilizing the quorum-sensing inhibitor on a skin-inhabiting bacterium, Staphylococcus epidermidis, we disrupted its interkingdom communication with adult Aedes aegypti and mitigated their attraction to a blood-meal by 55.1%. Discussion: One potential mechanism suppressing mosquito attraction could be the reduction (31.6% in our study) of bacterial volatiles and their associated concentrations by shifting S. epidermidis metabolic (12 of 29 up regulated genes) and stress (5 of 36 down regulated genes) responses. Manipulating the quorum-sensing pathways could serve as a mechanism to reduce mosquito attraction to a host. Such manipulations could be developed into novel control methods for pathogen-transmitting mosquitoes and other arthropods.

Impact of Temperature and Oxygen Availability on Gene Expression Patterns of Mycobacterium ulcerans.

Buruli ulcer disease is a neglected tropical disease caused by the environmental pathogen Mycobacterium ulcerans. The M. ulcerans major virulence factor is mycolactone, a lipid cytotoxic compound whose genes are carried on a plasmid. Although an exact reservoir and mode(s) of transmission are unknown, data provide evidence of both. First, Buruli ulcer incidence and M. ulcerans presence have been linked to slow-moving water with low oxygen. M. ulcerans has also been suggested to be sensitive to UV due to termination in crtI, encoding a phytoene dehydrogenase, required for carotenoid production. Further, M. ulcerans has been shown to cause disease following puncture but not when introduced to open abrasion sites, suggesting that puncture is necessary for transmission and pathology. Despite these findings, the function and modulation of mycolactone and other genes in response to dynamic abiotic conditions such as UV, temperature, and oxygen have not been shown. In this study, we investigated modulation of mycolactone and other genes on exposure to changing UV and oxygen microenvironmental conditions. Mycolactone expression was downregulated on exposure to the single stress high temperature and did not change significantly with exposure to UV; however, it was upregulated when exposed to microaerophilic conditions. Mycolactone expression was downregulated under combined stresses of high temperature and low oxygen, but there was upregulation of several stress response genes. Taken together, results suggest that temperature shapes M. ulcerans metabolic response more so than UV exposure or oxygen requirements. These data help to define the environmental niche of M. ulcerans and metabolic responses during initial human infection. IMPORTANCE Buruli ulcer is a debilitating skin disease caused by the environmental pathogen Mycobacterium ulcerans. M. ulcerans produces a toxic compound, mycolactone, which leads to tissue necrosis and ulceration. Barriers to preventing Buruli ulcer include an incomplete understanding of M. ulcerans reservoirs, how the pathogen is transmitted, and under what circumstances mycolactone and other M. ulcerans genes are expressed and produced in its natural environment and in the host. We conducted a study to investigate M. ulcerans gene expression under several individual or combined abiotic conditions. Our data showed that mycolactone expression was downregulated under combined stresses of high temperature and low oxygen but there was upregulation of several stress response genes. These data are among only a few studies measuring modulation of mycolactone and other M. ulcerans genes that could be involved in pathogen fitness in its natural environment and virulence while within the host.

Behavioral interplay between mosquito and mycolactone produced by Mycobacterium ulcerans and bacterial gene expression induced by mosquito proximity.

Mycolactone is a cytotoxic lipid metabolite produced by Mycobacterium ulcerans, the environmental pathogen responsible for Buruli ulcer, a neglected tropical disease. Mycobacterium ulcerans is prevalent in West Africa, particularly found in lentic environments, where mosquitoes also occur. Researchers hypothesize mosquitoes could serve as a transmission mechanism resulting in infection by M. ulcerans when mosquitoes pierce skin contaminated with M. ulcerans. The interplay between the pathogen, mycolactone, and mosquito is only just beginning to be explored. A triple-choice assay was conducted to determine the host-seeking preference of Aedes aegypti between M. ulcerans wildtype (MU, mycolactone active) and mutant (MUlac-, mycolactone inactive). Both qualitative and quantitative differences in volatile organic compounds' (VOCs) profiles of MU and MUlac- were determined by GC-MS. Additionally, we evaluated the interplay between Ae. aegypti proximity and M. ulcerans mRNA expression. The results showed that mosquito attraction was significantly greater (126.0%) to an artificial host treated with MU than MUlac-. We found that MU and MUlac produced differential profiles of VOCs associated with a wide range of biological importance from quorum sensing (QS) to human odor components. RT-qPCR assays showed that mycolactone upregulation was 24-fold greater for MU exposed to Ae. aegypti in direct proximity. Transcriptome data indicated significant induction of ten chromosomal genes of MU involved in stress responses and membrane protein, compared to MUlac- when directly having access to or in near mosquito proximity. Our study provides evidence of possible interkingdom interactions between unicellular and multicellular species that MU present on human skin is capable of interreacting with unrelated species (i.e., mosquitoes), altering its gene expression when mosquitoes are in direct contact or proximity, potentially impacting the production of its VOCs, and consequently leading to the stronger attraction of mosquitoes toward human hosts. This study elucidates interkingdom interactions between viable M. ulcerans bacteria and Ae. aegypti mosquitoes, which rarely have been explored in the past. Our finding opens new doors for future research in terms of disease ecology, prevalence, and pathogen dispersal outside of the M. ulcerans system.

Bioinformatic Surveillance Leads to Discovery of Two Novel Putative Bunyaviruses Associated with Black Soldier Fly.

The black soldier fly (Hermetia illucens, BSF) has emerged as an industrial insect of high promise because of its ability to convert organic waste into nutritious feedstock, making it an environmentally sustainable alternative protein source. As global interest rises, rearing efforts have also been upscaled, which is highly conducive to pathogen transmission. Viral epidemics have stifled mass-rearing efforts of other insects of economic importance, such as crickets, silkworms, and honeybees, but little is known about the viruses that associate with BSF. Although BSFs are thought to be unusually resistant to pathogens because of their expansive antimicrobial gene repertoire, surveillance techniques could be useful in identifying emerging pathogens and common BSF microbes. In this study, we used high-throughput sequencing data to survey BSF larvae and frass samples, and we identified two novel bunyavirus-like sequences. Our phylogenetic analysis grouped one in the family Nairoviridae and the other with two unclassified bunyaviruses. We describe these putative novel viruses as BSF Nairovirus-like 1 and BSF uncharacterized bunyavirus-like 1. We identified candidate segments for the full BSF Nairovirus-like 1 genome using a technique based on transcript co-occurrence and only a partial genome for BSF uncharacterized bunyavirus-like 1. These results emphasize the value of routine BSF colony surveillance and add to the number of viruses associated with BSF.

Electronic Health Record Prompt to Improve Lung Cancer Screening in Primary Care.

INTRODUCTION: Lung cancer is the leading cause of cancer death in the U.S. Combusted tobacco use, the primary risk factor, accounts for 90% of all lung cancers. Early detection of lung cancer improves survival, yet lung cancer screening rates are much lower than those of other cancer screening tests. Electronic health record (EHR) systems are an underutilized tool that could improve screening rates. METHODS: This study was conducted in the Rutgers Robert Wood Johnson Medical Group, a university-affiliated network in New Brunswick, NJ. Two novel EHR workflow prompts were implemented on July 1, 2018. These prompts included fields to determine tobacco use and lung cancer screening eligibility and facilitated low-dose computed tomography ordering for eligible patients. The prompts were designed to improve tobacco use data entry, allowing for better lung cancer screening eligibility identification. Data were analyzed in 2022 retrospectively for the period July 1, 2017 to June 30, 2019. The analyses represented 48,704 total patient visits. RESULTS: The adjusted odds of patient record completeness to determine eligibility for low-dose computed tomography (AOR=1.19, 95% CI=1.15, 1.23), eligibility for low-dose computed tomography (AOR=1.59, 95% CI=1.38, 1.82), and whether low-dose computed tomography was ordered (AOR=1.04, 95% CI=1.01, 1.07) all significantly increased after the electronic medical record prompts were implemented. CONCLUSIONS: These findings show the utility and benefit of EHR prompts in primary care settings to increase identification for lung cancer screening eligibility as well as increased low-dose computed tomography ordering.

Inhibition of Zoonotic Pathogens Naturally Found in Pig Manure by Black Soldier Fly Larvae and Their Intestine Bacteria.

Black soldier fly (BSF) larvae are often exposed to organic waste which harbors abundant zoonotic pathogens. We investigated the ability of BSF larvae to inhibit the zoonotic pathogens naturally found in pig manure. The zoonotic pathogens populations were detected by using selective medium during the conversion. Results showed that the viability of the zoonotic pathogens in pig manure was significantly affected. After eight days of conversion, the Coliform populations were undetected, and Staphylococcus aureus and Salmonella spp. decreased significantly on the eighth day. Antimicrobial assays of the purified recombinant defensin-like peptide 4 (DLP4) showed that this peptide exhibits inhibitory activity against S. aureus, Salmonella enterica serovar typhimurium, and Escherichia coli in vitro. Bacteria BSF-CL and BSF-F were isolated from the larvae gut, and both inhibited the growth of S. aureus and E. coli, but Salmonella spp. was sensitive to the BSF-CL strain (but not to the BSF-F strain). The results from our experiments indicate that BSF larvae are capable of functionally inhibiting potential zoonotic pathogens in pig manure through a variety of mechanisms including antimicrobial peptides expression and the gut associate microorganisms. This study provides a theoretical basis for further study on the combined mechanism of BSF larvae immunity and its gut microbes against the zoonotic pathogens in pig manure.

Hermetia illucens L. larvae-associated intestinal microbes reduce the transmission risk of zoonotic pathogens in pig manure.

Black soldier fly (BSF) larvae are considered a promising biological reactor to convert organic waste and reduce the impact of zoonotic pathogens on the environment. We analysed the effects of BSF larvae on Staphylococcus aureus and Salmonella spp. populations in pig manure (PM), which showed that BSF larvae can significantly reduce the counts of the associated S. aureus and Salmonella spp. Then, using a sterile BSF larval system, we validated the function of BSF larval intestinal microbiota in vivo to suppress pathogens, and lastly, we isolated eight bacterial strains from the BSF larval gut that inhibit S. aureus. Results indicated that functional microbes are essential for BSF larvae to antagonise S. aureus. Moreover, the analysis results of the relationship between the intestinal microbiota and S. aureus and Salmonella spp. showed that Myroides, Tissierella, Oblitimonas, Paenalcalignes, Terrisporobacter, Clostridium, Fastidiosipila, Pseudomonas, Ignatzschineria, Savagea, Moheibacter and Sphingobacterium were negatively correlated with S. aureus and Salmonella. Overall, these results suggested that the potential ability of BSF larvae to inhibit S. aureus and Salmonella spp. present in PM is accomplished primarily by gut-associated microorganisms.

A need for null models in understanding disease transmission: the example of Mycobacterium ulcerans (Buruli ulcer disease).

Understanding the interactions of ecosystems, humans and pathogens is important for disease risk estimation. This is particularly true for neglected and newly emerging diseases where modes and efficiencies of transmission leading to epidemics are not well understood. Using a model for other emerging diseases, the neglected tropical skin disease Buruli ulcer (BU), we systematically review the literature on transmission of the etiologic agent, Mycobacterium ulcerans (MU), within a One Health/EcoHealth framework and against Hill's nine criteria and Koch's postulates for making strong inference in disease systems. Using this strong inference approach, we advocate a null hypothesis for MU transmission and other understudied disease systems. The null should be tested against alternative vector or host roles in pathogen transmission to better inform disease management. We propose a re-evaluation of what is necessary to identify and confirm hosts, reservoirs and vectors associated with environmental pathogen replication, dispersal and transmission; critically review alternative environmental sources of MU that may be important for transmission, including invertebrate and vertebrate species, plants and biofilms on aquatic substrates; and conclude with placing BU within the context of other neglected and emerging infectious diseases with intricate ecological relationships that lead to disease in humans, wildlife and domestic animals.

Alternative splicing of the G protein-coupled receptor superfamily in human airway smooth muscle diversifies the complement of receptors.

G protein-coupled receptors (GPCRs) are the largest signaling family in the genome, serve an expansive array of functions, and are targets for approximately 50% of current therapeutics. In many tissues, such as airway smooth muscle (ASM), complex, unexpected, or paradoxical responses to agonists/antagonists occur without known mechanisms. We hypothesized that ASM express many more GPCRs than predicted, and that these undergo substantial alternative splicing, creating a highly diversified receptor milieu. Transcript arrays were designed detecting 434 GPCRs and their predicted splice variants. In this cell type, 353 GPCRs were detected (including 111 orphans), with expression levels varying by approximately 900-fold. Receptors used for treating airway disease were expressed lower than others with similar signaling properties, indicating potentially more effective targets. A disproportionate number of Class-A peptide-group receptors, and those coupling to G(q)/(11) or G(s) (vs. G(i)), was found. Importantly, 192 GPCRs had, on average, five different expressed receptor isoforms because of splicing events, including alternative splice donors and acceptors, novel introns, intron retentions, exon(s) skips, and novel exons, with the latter two events being most prevalent. The consequences of splicing were further investigated with the leukotriene B4 receptor, known for its aberrant responsiveness in lung. We found transcript expression of three variants because of alternative donor and acceptor splice sites, representing in-frame deletions of 38 and 100 aa, with protein expression of all three isoforms. Thus, alternative splicing, subject to conditional, temporal, and cell-type regulation, is a major mechanism that diversifies the GPCR superfamily, creating local recepteromes with specialized environments.