Disruption of cellular plasticity by repeat RNAs in human pancreatic cancer.

Aberrant expression of repeat RNAs in pancreatic ductal adenocarcinoma (PDAC) mimics viral-like responses with implications on tumor cell state and the response of the surrounding microenvironment. To better understand the relationship of repeat RNAs in human PDAC, we performed spatial molecular imaging at single-cell resolution in 46 primary tumors, revealing correlations of high repeat RNA expression with alterations in epithelial state in PDAC cells and myofibroblast phenotype in cancer-associated fibroblasts (CAFs). This loss of cellular identity is observed with dosing of extracellular vesicles (EVs) and individual repeat RNAs of PDAC and CAF cell culture models pointing to cell-cell intercommunication of these viral-like elements. Differences in PDAC and CAF responses are driven by distinct innate immune signaling through interferon regulatory factor 3 (IRF3). The cell-context-specific viral-like responses to repeat RNAs provide a mechanism for modulation of cellular plasticity in diverse cell types in the PDAC microenvironment.

Cohort profile: The Bariatric Experience Long Term (BELONG): a long-term prospective study to understand the psychosocial, environmental, health and behavioural predictors of weight loss and regain in patients who have bariatric surgery.

PURPOSE: The Bariatric Experience Long Term (BELONG) prospective study cohort was created to address limitations in the literature regarding the relationship between surgical weight loss and psychosocial, health, behaviour and environmental factors. The BELONG cohort is unique because it contains 70% gastric sleeve and 64% patients with non-white race/ethnicity and was developed with strong stakeholder engagement including patients and providers. PARTICIPANTS: The BELONG cohort study included 1975 patients preparing to have bariatric surgery who completed a baseline survey in a large integrated health system in Southern California. Patients were primarily women (84%), either black or Hispanic (59%), with a body mass index (BMI) of 45.1±7.4 kg/m2, age 43.3±11.5 years old, and 32% had at least one comorbidity. FINDINGS TO DATE: A total of 5552 patients were approached before surgery between February 2016 and May 2017, and 1975 (42%) completed a baseline survey. A total of 1203 (73%) patients completed the year 1 and 1033 (74%) patients completed the year 3 postoperative survey. Of these survey respondents, 1341 at baseline, 999 at year 1, and 951 at year 3 were included in the analyses of all survey and weight outcome data. A total of 803 (60% of eligible patients) had survey data for all time points. Data collected were self-reported constructs to support the proposed theoretical model. Height, weight and BMI were abstracted from the electronic medical record to obtain the main outcomes of the study: weight loss and regain. FUTURE PLANS: We will collect self-reported constructs and obtain height, weight and BMI from the electronic medical record 5 years after bariatric surgery between April 2022 and January 2023. We will also collect patient experiences using focus groups of 8-12 patients each throughout 2022.

Symptom burden profiles in men with advanced prostate cancer undergoing androgen deprivation therapy.

To identify symptom burden profiles among men with advanced prostate cancer undergoing androgen-deprivation therapy and examine their association with baseline sociodemographic and medical characteristics and psychosocial outcomes over time. Latent profile analysis was employed to identify distinct groups based on the Expanded Prostate Index Composite and the McGill Pain Questionnaire at baseline. Psychosocial outcomes were assessed at baseline, 6- and 12-month follow-ups. Three profiles emerged: "high symptom burden," "high sexual bother," and "low symptom burden." Men with "high symptom burden" were younger and exhibited higher baseline levels of depression, stress, cancer-specific distress, and anxiety than men in the other two groups. However, men with "high symptom burden" also demonstrated improvement in these psychosocial outcomes over time. Men with advanced prostate cancer who experience multiple co-occurring symptoms demonstrate worse psychosocial adjustment. Patients with substantial symptom burden, and specifically young men, may benefit from prompt referral to supportive care services.

Minimum Threshold of Bariatric Surgical Weight Loss for Initial Diabetes Remission.

OBJECTIVE: There are few studies testing the amount of weight loss necessary to achieve initial remission of type 2 diabetes mellitus (T2DM) following bariatric surgery and no published studies with use of weight loss to predict initial T2DM remission in sleeve gastrectomy (SG) patients. RESEARCH DESIGN AND METHODS: With Cox proportional hazards models we examined the relationship between initial T2DM remission and percent total weight loss (%TWL) after bariatric surgery. Categories of %TWL were included in the model as time-varying covariates. RESULTS: Of patients (N = 5,928), 73% were female; mean age was 49.8 ± 10.3 years and BMI 43.8 ± 6.92 kg/m2, and 57% had Roux-en-Y gastric bypass (RYGB). Over an average follow-up of 5.9 years, 71% of patients experienced initial remission of T2DM (mean time to remission 1.0 year). With 0-5% TWL used as the reference group in Cox proportional hazards models, patients were more likely to remit with each 5% increase in TWL until 20% TWL (hazard ratio range 1.97-2.92). When categories >25% TWL were examined, all patients had a likelihood of initial remission similar to that of 20-25% TWL. Patients who achieved >20% TWL were more likely to achieve initial T2DM remission than patients with 0-5% TWL, even if they were using insulin at the time of surgery. CONCLUSIONS: Weight loss after bariatric surgery is strongly associated with initial T2DM remission; however, above a threshold of 20% TWL, rates of initial T2DM remission did not increase substantially. Achieving this threshold is also associated with initial remission even in patients who traditionally experience lower rates of remission, such as patients taking insulin.

Bariatric Surgery and Risk of Death in Persons With Chronic Kidney Disease.

OBJECTIVE: A retrospective cohort study investigated the association between having surgery and risk of mortality for up to 5 years and if this association was modified by incident ESRD during the follow-up period. Summary of Background Data: Mortality risk in individuals with pre-dialysis CKD is high and few effective treatment options are available. Whether bariatric surgery can improve survival in people with CKD is unclear. METHODS: Patients with class II and III obesity and pre-dialysis CKD stages 3-5 who underwent bariatric surgery between January 1, 2006 and September 30, 2015 (n = 802) were matched to patients who did not have surgery (n = 4933). Mortality was obtained from state death records and ESRD was identified through state-based or healthcare system-based registries. Cox regression models were used to investigate the association between bariatric surgery and risk of mortality and if this was moderated by incident ESRD during the follow-up period. RESULTS: Patients were primarily women (79%), non-Hispanic White (72%), under 65 years old (64%), who had a body mass index > 40kg/m 2 (59%), diabetes (67%), and hypertension (89%). After adjusting for incident ESRD, bariatric surgery was associated with a 79% lower 5-year risk of mortality compared to matched controls (hazard ratio = 0.21; 95% confidence interval: 0.14-0.32; P < 0.001). Incident ESRD did not moderate the observed association between surgery and mortality (hazard ratio = 1.59; 95% confidence interval: 0.31-8.23; P =0.58). CONCLUSIONS: Bariatric surgery is associated with a reduction in mortality in pre-dialysis patients regardless of developing ESRD. These findings are significant because patients with CKD are at relatively high risk for death with few efficacious interventions available to improve survival.

Bariatric Surgical Alterations in Tolerability, Enjoyment and Cravings in the Diet (BSATED) instrument: A new scale to measure food preferences following bariatric surgery.

Bariatric surgery is associated with changing food preferences, but it is not known whether these changes differ by type of operation or are associated with weight loss. The current study presents validation results for a new 27-item scale, Bariatric Surgical Alterations in Tolerability, Enjoyment and Cravings in the Diet (BSATED). This scale measured enjoyment, craving, and intolerance changes for nine food and beverage categories common to dietary habits in the Southern California region of the U.S. one year following bariatric surgery in the Bariatric Experience Long Term (BELONG) study. Validation of BSATED was done using exploratory factor analyses, construct validity with other conceptually related survey instruments, and criterion validity using hypothesized differences for operation type and percent total weight loss (%TWL) at 12-18 months after surgery. Participants (n = 999) were 86% female, 41% non-Hispanic White, with a mean age of 43.1 ± 11.3 years and a body mass index (BMI) of 43.4 ± 6.8 kg/m2 at the time of surgery. Participants reported less enjoyment and craving for high-fat meats (62%), grains (54%), candy and other desserts (e.g. candy bars, chocolate, ice cream) (52%), and sweet baked goods (48%) 12 months after surgery. These changes were more common among participants undergoing Roux-en-Y gastric bypass (RYGB) compared to those receiving sleeve gastrectomy (SG). Participants who reported decreased enjoyment and craving for foods and beverages that post-bariatric patients are counseled to reduce or avoid had greater %TWL at 12-18 months following surgery (p < .001 and p = .003 respectively). The foods and beverages in BSATED that post-bariatric patients are counseled to reduce or avoid could be used to understand how changes in enjoyment, craving and tolerability of these foods/beverages contribute to weight loss following surgery.

The Bariatric Experience Long Term (BELONG): Factors Related to Having Bariatric Surgery in a Large Integrated Healthcare System.

PURPOSE: Bariatric surgery is the most effective treatment for severe obesity, but currently, only 1-2% of all eligible patients undergo surgery each year. This study examined which factors were associated with a patient receiving bariatric surgery after referral in a real-world healthcare setting. MATERIALS AND METHODS: The current study used the baseline survey and electronic medical record (EMR) data from the Bariatric Experience Long Term (BELONG) study (n = 1975). Predictors of who did (n = 1680) and who did not (n = 295) have surgery were analyzed using multivariate logistic regression. RESULTS: Participants (n = 1975; 42.4% response rate) were primarily women (84%) and either non-Hispanic Black or Hispanic (60%). In the fully adjusted multivariate model, the strongest predictors of having surgery were being a woman (OR = 3.17; 95% CI = 2.15, 4.68; p < .001) and losing at least 5% of their body weight in the year before surgery (OR = 3.16; 95% CI = 2.28, 4.38; p < .001). The strongest predictors of not having surgery were a ≥ BMI 50 kg/m2 (OR = .39; 95% CI = .27, .56; p < .001) and having a higher physical comorbidity burden (OR = .84; 95% CI = .75, .94; p = .004). CONCLUSIONS: Practices such as 5-10% total weight loss before surgery and selection of patients with safer operative risk profiles (younger with lower comorbidity burden) may inadvertently contribute to under-utilization of bariatric surgery among some demographic subpopulations who could most benefit from this intervention.

Five-year Longitudinal Cohort Study of Reinterventions After Sleeve Gastrectomy and Roux-en-Y Gastric Bypass.

OBJECTIVE: To compare the long-term risks of reintervention following sleeve gastrectomy (SG) and Roux-en-Y gastric bypass (RYGB) in a large surgical cohort. BACKGROUND: The use of SG has increased dramatically relative to RYGB for the treatment of obesity. However, long-term risks following SG compared with RYGB have not been adequately defined in a large population-based study. METHODS: A retrospective longitudinal cohort study of all adult health-plan members undergoing SG or RYGB for obesity in a multistate integrated health care system from January 2005 through September 2015. The risks of nutritional, endoscopic, radiologic, and surgical reintervention as well as the overall risk of any reinterventions at 1, 3, and 5 years were identified using diagnosis and procedure codes from comprehensive electronic medical records. RESULTS: The study included 15,319 patients who underwent SG and 19,954 patients who underwent RYGB with a follow-up of 79.2%. The overall risk of any reintervention at 5 years was 21.3% for SG and 28.3% for RYGB (P < 0.0001). After adjustment, SG was associated with fewer reinterventions through 5 years than RYGB (hazard ratio, 0.78; 95% confidence interval, 0.74-0.84). When comparing subcategories, SG also had a lower risk of nutritional, endoscopic, radiologic, and surgical reinterventions when examined versus RYGB. The findings for risks of reinterventions were consistent across clinical subgroups. CONCLUSION: SG has significantly lower risk of reintervention in all categories studied when compared with RYGB at 5-year follow-up. The long-term safety profile of LSG compared with RYGB should be an essential part of the discussion in patient-centered decision making when choosing between bariatric procedure options.

Effectiveness of Gastric Bypass Versus Gastric Sleeve for Cardiovascular Disease: Protocol and Baseline Results for a Comparative Effectiveness Study.

BACKGROUND: When compared with conventional weight loss strategies, bariatric surgery results in substantially greater durable weight loss and rates of disease remission. OBJECTIVE: The ENGAGE CVD (Effectiveness of Gastric Bypass versus Gastric Sleeve for Cardiovascular Disease) cohort study aimed to provide population-based, comprehensive, rigorous evidence for clinical and policy decision making regarding the choice between gastric bypass and gastric sleeve for overall cardiovascular disease (CVD) risk reduction, risk factor remission, and safety. METHODS: The cohort had 22,095 weight loss surgery patients from a large integrated health care system in Southern California assembled from 2009 to 2016 who were followed up through 2018. Bariatric surgery patients were followed up for the length of their membership in the health care system. Of the patients who had at least five years of follow-up (surgery between 2009 and 2013), 85.86% (13,774/16,043) could contribute to the outcome analyses for the ENGAGE CVD cohort. RESULTS: Patients in the ENGAGE CVD cohort were 44.6 (SD 11.4) years old, mostly women (17,718/22,095; 80.19%), with 18.94% (4185/22,095) non-Hispanic black and 41.80% (9235/22,095) Hispanic, and had an average BMI of 44.3 (SD 6.9) kg/m2 at the time of surgery. When compared with patients who did not contribute data to the 5-year outcome analysis for the ENGAGE CVD cohort (2269/16,043; 14.14%), patients who contributed data (13,774/16,043; 85.86%) were older (P=.002), more likely to be women (P=.02), more likely to be non-Hispanic white (P<.001), more likely to have had an emergency department visit in the year before surgery (P=.006), less likely to have a mental illness before surgery (P<.001), and more likely to have had a CVD event at any time before surgery (P<.001). CONCLUSIONS: This study had one of the largest populations of gastric sleeve patients (n=13,459). The 5-year follow-up for those patients who had surgery between 2009 and 2013 was excellent for a retrospective cohort study at 85.86% (13,774/16,043). Unlike almost any study in the literature, the majority of the ENGAGE CVD cohort was racial and ethnic minority, providing a rare opportunity to study the effects of bariatric surgery for different racial and ethnic groups, some of whom have the highest rates of severe obesity in the United States. Finally, it also used state-of-the-art statistical and econometric comparative effectiveness methods to mimic the effect of random assignment and control for sources of confounding inherent in large observational studies. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): RR1-10.2196/14936.

MRI-Based Upper Abdominal Organs-at-Risk Atlas for Radiation Oncology.

PURPOSE: The purpose of our study was to provide a guide for identification and contouring of upper abdominal organs-at-risk (OARs) in the setting of online magnetic resonance imaging (MRI)-guided radiation treatment planning and delivery. METHODS AND MATERIALS: After a needs assessment survey, it was determined that an upper abdominal MRI-based atlas of normal OARs would be of benefit to radiation oncologists and radiation therapists. An anonymized diagnostic 1.5T MRI from a patient with typical upper abdominal anatomy was used for atlas development. Two MRI sequences were selected for contouring, a T1-weighted gadoxetic acid contrast-enhanced MRI acquired in the hepatobiliary phase and axial fast imaging with balanced steady-state precession. Two additional clinical MRI sequences from commercial online MRI-guided radiation therapy systems were selected for contouring and were included in the final atlas. Contours from each data set were completed and reviewed by radiation oncologists, along with a radiologist who specializes in upper abdominal imaging, to generate a consensus upper abdominal MRI-based OAR atlas. RESULTS: A normal OAR atlas was developed, including recommendations for contouring. The atlas and contouring guidance are described, and high-resolution MRI images and contours are displayed. OARs, such as the bile duct and biliary tree, which may be better seen on MRI than on computed tomography, are highlighted. The full DICOM/DICOM-RT MRI images from both the diagnostic and clinical online MRI-guided radiation therapy systems data sets have been made freely available, for educational purposes, at econtour.org. CONCLUSIONS: This MRI contouring atlas for upper abdominal OARs should provide a useful reference for contouring and education. Its routine use may help to improve uniformity in contouring in radiation oncology planning and OAR dose calculation. Full DICOM/DICOM-RT images are available online and provide a valuable educational resource for upper abdominal MRI-based radiation therapy planning and delivery.

Ultrasound-guided radial artery cannulation using dynamic needle tip positioning versus conventional long-axis in-plane techniques in cardiac surgery patients: a randomized, controlled trial.

BACKGROUND: A novel ultrasound imaging technique, dynamic needle tip positioning (DNTP), enables continuous visualization of the needle tip during ultrasound-guided cannulation. The purpose of this study was to compare the rate of successful first-attempt radial artery cannulations between DNTP and the conventional long-axis in-plane (LAX-IP) technique. METHODS: Adult patients undergoing cardiac surgery requiring radial artery cannulation were included and randomly allocated into either a DNTP or LAX-IP group. Radial artery cannulation was performed by a single experienced practitioner. The primary outcome was the first-attempt success rate of radial artery cannulation. Secondary outcomes included the length of time needed for cannulation and overall incidence of complications. RESULTS: A total of 136 patients were studied. The first-attempt success rate of cannulation was 94% in the DNTP group (N.=70) and 68% in the LAX-IP group (N.=66; OR 7.70, 95% CI 2.48-24.94, P<0.001). The total procedure time was also shorter in the DNTP group (median [IQR]; 87 [72-108] seconds) versus the LAX-IP group (118 [93-182] seconds; P<0.001). During cannulation, vasospasm occurred less often in the DNTP (4%) group compared to the LAX-IP group (17%; P=0.018). CONCLUSIONS: The DNTP technique had a greater first-attempt success rate of radial artery cannulation compared to the conventional LAX-IP technique. Also, DNTP required significantly less time for cannulation and had fewer complications.

The velocity curve of the clotting waveform of rotational thromboelastometry predicts bleeding after cardiac surgery but conventional rotational thromboelastometric parameters do not.

BACKGROUND: Early detection of a risk of postoperative bleeding is essential in cardiac surgery patients. The aim of the present study was to evaluate the utility of the first derivative curve (the V-curve) of the clotting waveform of rotational thromboelastometry (ROTEM) in terms of predicting bleeding after cardiac surgery. METHODS: We retrospectively analysed 534 cardiac surgery patients. We used the chest tube output during the stay in the intensive care unit to divide patients into a higher blood loss group (HBL group; the fourth quartile) and a lower blood loss group (LBL group; the lower quartiles). We performed multivariable logistic regression using the V-curve parameters and potential confounders including conventional ROTEM parameters. RESULTS: In the multivariable model, the adjusted odds ratios for HBL of patients with a lower maximum clotting velocity (MaxVel ≤9 mm*100 s-1) as revealed by extrinsically activated ROTEM (EXTEM), and the area under the velocity curve (AUC ≤988 mm*100) of the fibrin-based extrinsically activated ROTEM (FIBTEM), both measured at skin closure in the end of surgery, were 1.78 (95% CI 1.03 to 3.07) and 2.14 (95% CI 1.20 to 3.82), respectively. However, conventional ROTEM parameters were not included in the final model. Additionally, lower EXTEM MaxVel and FIBTEM AUC values were associated with the need for a higher transfusion volume, longer postoperative intensive care unit and hospital stays, and more frequent re-exploration to control bleeding. CONCLUSIONS: The ROTEM V-curve parameters can predict postoperative bleeding and clinical outcomes after cardiac surgery.

The role of psychological factors in the development of burning mouth syndrome.

The psychiatric profiles of 50 patients diagnosed with burning mouth syndrome (BMS) were compared to those of 50 age- and sex-matched individuals as the control group. The Symptom Checklist-90-Revised (SCL-90-R) questionnaire was used to evaluate the role of psychological factors in the development of BMS. Somatization, obsessive-compulsive, depression, anxiety, hostility, phobic anxiety, psychoticism, global severity index (GSI), positive symptom total (PST), and positive symptom distress index (PSDI) scores were significantly higher in the patients with BMS than in the control group. In a subgroup analysis according to sex, women with BMS had higher T-scores for somatization, obsessive-compulsive, paranoid ideation, GSI, PST, and PSDI than women in the control group. In contrast, only the PSDI score was significantly higher in men with BMS compared to men in the control group. There was a significant difference in the T-scores for somatization, psychoticism, and GSI between the three age subgroups (≤50, 51-65, and ≥66 years). The obsessive-compulsive and PSDI scores were significantly higher in patients with BMS who also had at least one chronic disease than in patients with BMS who had no chronic disease. In conclusion, psychological factors are correlated with BMS.

Robot-assisted excision of the submandibular gland by a postauricular facelift approach: comparison with the conventional transcervical approach.

Various approaches have been described for excision of a submandibular gland including endoscopic and robot-assisted techniques. We present the outcome of excision by a robot-assisted postauricular facelift approach and compare it with the conventional transcervical approach. We studied 30 cases of excision of the gland for benign disease (16 transcervical and 14 robot-assisted), and collected clinical and personal data, and details of patients' satisfaction with the result. The most common conditions were sialadenitis (n=15) and pleomorphic adenoma (n=12). The robot-assisted operations took significantly longer (p=0.045), had more drainage (p<0.001), and a significantly better cosmetic outcome (p=0.002). Robot-assisted excision of the submandibular gland may prove to be a viable option in the treatment of benign conditions for those patients seeking a better cosmetic outcome.

Feasibility of robot-assisted modified radical neck dissection by post-auricular facelift approach.

The aim of this study was to evaluate the technical feasibility and safety of robot-assisted modified radical neck dissection (MRND) for head and neck cancer patients with a clinically node-positive neck. The cases of 10 head and neck cancer patients who underwent unilateral therapeutic robot-assisted MRND by post-auricular facelift approach were analyzed. The robot-assisted MRND was completed successfully in all patients without any conversion to conventional neck dissection. The mean number of lymph nodes removed was 36.7±8.6. The mean duration of surgery for robot-assisted MRND was 274±65min (range 175-395min). Transient marginal nerve palsy occurred in two patients and partial necrosis of the skin flap occurred in one patient. In terms of cosmetic satisfaction, 70% of patients were very satisfied or satisfied with postoperative cosmesis. In conclusion, robot-assisted MRND by post-auricular facelift approach is technically feasible and safe in selected patients with head and neck cancer, and yields excellent postoperative cosmesis.

Interobserver variability in target definition for hepatocellular carcinoma with and without portal vein thrombus: radiation therapy oncology group consensus guidelines.

PURPOSE: Defining hepatocellular carcinoma (HCC) gross tumor volume (GTV) requires multimodal imaging, acquired in different perfusion phases. The purposes of this study were to evaluate the variability in contouring and to establish guidelines and educational recommendations for reproducible HCC contouring for treatment planning. METHODS AND MATERIALS: Anonymous, multiphasic planning computed tomography scans obtained from 3 patients with HCC were identified and distributed to a panel of 11 gastrointestinal radiation oncologists. Panelists were asked the number of HCC cases they treated in the past year. Case 1 had no vascular involvement, case 2 had extensive portal vein involvement, and case 3 had minor branched portal vein involvement. The agreement between the contoured total GTVs (primary + vascular GTV) was assessed using the generalized kappa statistic. Agreement interpretation was evaluated using Landis and Koch's interpretation of strength of agreement. The S95 contour, defined using the simultaneous truth and performance level estimation (STAPLE) algorithm consensus at the 95% confidence level, was created for each case. RESULTS: Of the 11 panelists, 3 had treated >25 cases in the past year, 2 had treated 10 to 25 cases, 2 had treated 5 to 10 cases, 2 had treated 1 to 5 cases, 1 had treated 0 cases, and 1 did not respond. Near perfect agreement was seen for case 1, and substantial agreement was seen for cases 2 and 3. For case 2, there was significant heterogeneity in the volume identified as tumor thrombus (range 0.58-40.45 cc). For case 3, 2 panelists did not include the branched portal vein thrombus, and 7 panelists contoured thrombus separately from the primary tumor, also showing significant heterogeneity in volume of tumor thrombus (range 4.52-34.27 cc). CONCLUSIONS: In a group of experts, excellent agreement was seen in contouring total GTV. Heterogeneity exists in the definition of portal vein thrombus that may impact treatment planning, especially if differential dosing is contemplated. Guidelines for HCC GTV contouring are recommended.

A 24-48 h fed Amblyomma americanum tick saliva immuno-proteome.

BACKGROUND: Multiple tick saliva proteins, the majority of which are unknown, confer tick resistance in repeatedly infested animals. The objective of this study was to identify the 24-48 h fed Amblyomma americanum tick saliva immuno-proteome. The 24-48 h tick-feeding phase is critical to tick parasitism as it precedes important events in tick biology, blood meal feeding and disease agent transmission. Fed male, 24 and 96 h fed female phage display cDNA expression libraries were biopanned using rabbit antibodies to 24 and 48 h fed A. americanum female tick saliva proteins. Biopanned immuno-cDNA libraries were subjected to next generation sequencing, de novo assembly, and bioinformatic analysis. RESULTS: More than 800 transcripts that code for 24-48 h fed A. americanum immuno-proteins are described. Of the 895 immuno-proteins, 52% (464/895) were provisionally identified based on matches in GenBank. Of these, ~19% (86/464) show high level of identity to other tick hypothetical proteins, and the rest include putative proteases (serine, cysteine, leukotriene A-4 hydrolase, carboxypeptidases, and metalloproteases), protease inhibitors (serine and cysteine protease inhibitors, tick carboxypeptidase inhibitor), and transporters and/or ligand binding proteins (histamine binding/lipocalin, fatty acid binding, calreticulin, hemelipoprotein, IgG binding protein, ferritin, insulin-like growth factor binding proteins, and evasin). Others include enzymes (glutathione transferase, cytochrome oxidase, protein disulfide isomerase), ribosomal proteins, and those of miscellaneous functions (histamine release factor, selenoproteins, tetraspanin, defensin, heat shock proteins). CONCLUSIONS: Data here demonstrate that A. americanum secretes a complex cocktail of immunogenic tick saliva proteins during the first 24-48 h of feeding. Of significance, previously validated immunogenic tick saliva proteins including AV422 protein, calreticulin, histamine release factor, histamine binding/lipocalins, selenoproteins, and paramyosin were identified in this screen, supporting the specificity of the approach in this study. While descriptive, this study opens opportunities for in-depth tick feeding physiology studies.

Comparative bioinformatics, temporal and spatial expression analyses of Ixodes scapularis organic anion transporting polypeptides.

Organic anion-transporting polypeptides (Oatps) are an integral part of the detoxification mechanism in vertebrates and invertebrates. These cell surface proteins are involved in mediating the sodium-independent uptake and/or distribution of a broad array of organic amphipathic compounds and xenobiotic drugs. This study describes bioinformatics and biological characterization of 9 Oatp sequences in the Ixodes scapularis genome. These sequences have been annotated on the basis of 12 transmembrane domains, consensus motif D-X-RW-(I,V)-GAWW-X-G-(F,L)-L, and 11 conserved cysteine amino acid residues in the large extracellular loop 5 that characterize the Oatp superfamily. Ixodes scapularis Oatps may regulate non-redundant cross-tick species conserved functions in that they did not cluster as a monolithic group on the phylogeny tree and that they have orthologs in other ticks. Phylogeny clustering patterns also suggest that some tick Oatp sequences transport substrates that are similar to those of body louse, mosquito, eye worm, and filarial worm Oatps. Semi-quantitative RT-PCR analysis demonstrated that all 9 I. scapularis Oatp sequences were expressed during tick feeding. Ixodes scapularis Oatp genes potentially regulate functions during early and/or late-stage tick feeding as revealed by normalized mRNA profiles. Normalized transcript abundance indicates that I. scapularis Oatp genes are strongly expressed in unfed ticks during the first 24h of feeding and/or at the end of the tick feeding process. Except for 2 I. scapularis Oatps, which were expressed in the salivary glands and ovaries, all other genes were expressed in all tested organs, suggesting the significance of I. scapularis Oatps in maintaining tick homeostasis. Different I. scapularis Oatp mRNA expression patterns were detected and discussed with reference to different physiological states of unfed and feeding ticks.