Whole-Body PET Imaging in Humans Shows That 11C-PS13 Is Selective for Cyclooxygenase-1 and Can Measure the In Vivo Potency of Nonsteroidal Antiinflammatory Drugs.

Both cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) convert arachidonic acid to prostaglandin H2, which has proinflammatory effects. The recently developed PET radioligand 11C-PS13 has excellent in vivo selectivity for COX-1 over COX-2 in nonhuman primates. This study sought to evaluate the selectivity of 11C-PS13 binding to COX-1 in humans and assess the utility of 11C-PS13 to measure the in vivo potency of nonsteroidal antiinflammatory drugs. Methods: Baseline 11C-PS13 whole-body PET scans were obtained for 26 healthy volunteers, followed by blocked scans with ketoprofen (n = 8), celecoxib (n = 8), or aspirin (n = 8). Ketoprofen is a highly potent and selective COX-1 inhibitor, celecoxib is a preferential COX-2 inhibitor, and aspirin is a selective COX-1 inhibitor with a distinct mechanism that irreversibly inhibits substrate binding. Because blood cells, including platelets and white blood cells, also contain COX-1, 11C-PS13 uptake inhibition from blood cells was measured in vitro and ex vivo (i.e., using blood obtained during PET scanning). Results: High 11C-PS13 uptake was observed in major organs with high COX-1 density, including the spleen, lungs, kidneys, and gastrointestinal tract. Ketoprofen (1-75 mg orally) blocked uptake in these organs far more effectively than did celecoxib (100-400 mg orally). On the basis of the plasma concentration to inhibit 50% of the maximum radioligand binding in the spleen (in vivo IC 50), ketoprofen (<0.24 µM) was more than 10-fold more potent than celecoxib (>2.5 µM) as a COX-1 inhibitor, consistent with the in vitro potencies of these drugs for inhibiting COX-1. Blockade of 11C-PS13 uptake from blood cells acquired during the PET scans mirrored that in organs of the body. Aspirin (972-1,950 mg orally) blocked such a small percentage of uptake that its in vivo IC 50 could not be determined. Conclusion: 11C-PS13 selectively binds to COX-1 in humans and can measure the in vivo potency of nonsteroidal antiinflammatory drugs that competitively inhibit arachidonic acid binding to COX-1. These in vivo studies, which reflect the net effect of drug absorption and metabolism in all organs of the body, demonstrated that ketoprofen had unexpectedly high potency, that celecoxib substantially inhibited COX-1, and that aspirin acetylation of COX-1 did not block binding of the representative nonsteroidal inhibitor 11C-PS13.

Tandem Mass Spectrometry as an Independent Method for Corroborating Fluorine-18 Radioactivity Measurements in Positron Emission Tomography.

Positron emission tomography (PET) uses many tracers labeled with fluorine-18 (t 1/2 = 109.8 min; ß+ 97%) for quantitative imaging of biochemical and physiological processes in animal and human subjects. In PET methodology, the radioactivity in a dose of an 18F-labeled tracer to be administered to a living subject is measured with a calibrated ionization chamber. This type of detector measures the radioactivity of a sample relative to those of certified amounts of longer-lived surrogate isotopes that are recommended for detector calibration. No alternative means for corroborating widely varying fluorine-18 radioactivity measurements from calibrated ionization chambers has been available. Here, we describe an independent nonradiometric method for this purpose. In this method, highly sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) is used to quantify the relative masses of the radioactive isotopologue ([18F]1) and the accompanying nonradioactive counterpart (carrier 1) in an 18F-labeled tracer preparation to give the mole ratio of [18F]1. High-performance liquid chromatography (HPLC) with a mass-calibrated absorbance detection is used alongside to provide a separate measurement of the aggregate mass of all isotopologues. The radioactivity of the radiotracer is then derived in becquerels (Bq) from these two measurements, plus Avogadro's number and the decay constant of fluorine-18. For the chosen example [18F]LSN3316612, the radioactivity values determined nonradiometrically and with a selected ionization chamber were in fair agreement. In addition, LC-MS/MS alone was found to provide an accurate measure of the half-life of fluorine-18.

Ethnic inequities in multimorbidity among people with psychosis: a retrospective cohort study.

AIMS: Research shows persistent ethnic inequities in mental health experiences and outcomes, with a higher incidence of illnesses among minoritised ethnic groups. People with psychosis have an increased risk of multiple long-term conditions (MLTC; multimorbidity). However, there is limited research regarding ethnic inequities in multimorbidity in people with psychosis. This study investigates ethnic inequities in physical health multimorbidity in a cohort of people with psychosis. METHODS: In this retrospective cohort study, using the Clinical Records Interactive Search (CRIS) system, we identified service-users of the South London and Maudsley NHS Trust with a schizophrenia spectrum disorder, and then additional diagnoses of diabetes, hypertension, low blood pressure, overweight or obesity and rheumatoid arthritis. Logistic and multinomial logistic regressions were used to investigate ethnic inequities in odds of multimorbidity (psychosis plus one physical health condition), and multimorbidity severity (having one or two physical health conditions, or three or more conditions), compared with no additional health conditions (no multimorbidity), respectively. The regression models adjusted for age and duration of care and investigated the influence of gender and area-level deprivation. RESULTS: On a sample of 20 800 service-users with psychosis, aged 13-65, ethnic differences were observed in the odds for multimorbidity. Controlling for sociodemographic factors and duration of care, compared to White British people, higher odds of multimorbidity were found for people of Black African [adjusted Odds Ratio = 1.41, 95% Confidence Intervals (1.23-1.56)], Black Caribbean [aOR = 1.79, 95% CI (1.58-2.03)] and Black British [aOR = 1.64, 95% CI (1.49-1.81)] ethnicity. Reduced odds were observed among people of Chinese [aOR = 0.61, 95% CI (0.43-0.88)] and Other ethnic [aOR = 0.67, 95% CI (0.59-0.76)] backgrounds. Increased odds of severe multimorbidity (three or more physical health conditions) were also observed for people of any Black background. CONCLUSIONS: Ethnic inequities are observed for multimorbidity among people with psychosis. Further research is needed to understand the aetiology and impact of these inequities. These findings support the provision of integrated health care interventions and public health preventive policies and actions.

Translation of 11C-labeled tracer synthesis to a CGMP environment as exemplified by [11C]ER176 for PET imaging of human TSPO.

Radiotracers labeled with carbon-11 (t1/2 = 20.4 min) are widely used with positron emission tomography for biomedical research. Radiotracers must be produced for positron emission tomography studies in humans according to prescribed time schedules while also meeting current good manufacturing practice. Translation of an experimental radiosynthesis to a current good manufacturing practice environment is challenging. Here we exemplify such translation with a protocol for the production of an emerging radiotracer for imaging brain translocator protein 18 kDa, namely [11C]ER176. This radiotracer is produced by rapid conversion of cyclotron-produced [11C]carbon dioxide into [11C]iodomethane, which is then used to treat N-desmethyl-ER176 in the presence of base (tBuOK) at room temperature for 5 min. [11C]ER176 is separated in high purity by reversed-phase HPLC and formulated for intravenous injection in sterile ethanol-saline. The radiosynthesis is reliable and takes 50 min. Quality control takes another 20 min. All aspects of the protocol, including quality control, are discussed.

Development of a non-radiometric method for measuring the arterial input function of a 11C-labeled PET radiotracer.

Positron emission tomography (PET) uses radiotracers to quantify important biochemical parameters in human subjects. A radiotracer arterial input function (AIF) is often essential for converting brain PET data into robust output measures. For radiotracers labeled with carbon-11 (t1/2 = 20.4 min), AIF is routinely determined with radio-HPLC of blood sampled frequently during the PET experiment. There has been no alternative to this logistically demanding method, neither for regular use nor validation. A 11C-labeled tracer is always accompanied by a large excess of non-radioactive tracer known as carrier. In principle, AIF might be obtained by measuring the molar activity (Am; ratio of radioactivity to total mass; Bq/mol) of a radiotracer dose and the time-course of carrier concentration in plasma after radiotracer injection. Here, we implement this principle in a new method for determining AIF, as shown by using [11C]PBR28 as a representative tracer. The method uses liquid chromatography-tandem mass spectrometry for measuring radiotracer Am and then the carrier in plasma sampled regularly over the course of a PET experiment. Am and AIF were determined radiometrically for comparison. The new non-radiometric method is not constrained by the short half-life of carbon-11 and is an attractive alternative to conventional AIF measurement.

Influence of apical preparation size and irrigation technique on root canal debridement: a histological analysis of round and oval root canals.

AIM: To examine the debridement of round and oval root canals prepared to two apical sizes with and without ultrasonically activated irrigation. METHODOLOGY: Mandibular premolars with round (n = 48) and oval (n = 48) root canals were pair-matched after microCT scanning and randomly divided into two experimental groups (n = 20): group 1, rotary NiTi to size 20, .04 taper; group 2, rotary NiTi to size 40, .04 taper. Specimens were subdivided into two subgroups (n = 10): subgroup A, syringe and needle (SNI); subgroup B, ultrasonically activated irrigation (UAI). Untreated canals (eight oval and eight round) served as controls. Specimens were processed for histological evaluation for measurement of the remaining pulp tissue and debris (RPT), and the perimeter percentage of root canal area untouched by the instruments (PRAU). Following assessment of normality, multiple-way anova models were used to study the effects of preparation size, irrigation technique and canal cross-sectional shape, and their interactions on the RPT and PRAU (α = 0.05). RESULTS: All experimental groups had significantly less RPT than the control (P < 0.05). Both the preparation size (20 vs. 40) and the irrigation technique (SNI vs. UAI) had a significant effect on RPT (P = 0.006 and P < 0.001, respectively). Groups irrigated with SNI always had significantly greater RPT than those irrigated with UAI, irrespective of the preparation size (P < 0.001). Canals prepared to size 20 had significantly greater RPT than those with size 40 in the SNI subgroup (P < 0.001), but there was no significant difference in the UAI subgroup (P = 0.481). CONCLUSIONS: Root canals prepared to a larger size (40) were cleaner than those prepared to a smaller size (20), when irrigation was performed with a syringe and needle. When the irrigant was ultrasonically activated, smaller preparations resulted in canals that were as clean as larger preparations. This finding was common to both round and oval canals of freshly extracted premolars with vital pulp tissue.

Clozapine and all-cause mortality in treatment-resistant schizophrenia: a historical cohort study.

OBJECTIVE: Large-scale epidemiological studies have demonstrated a protective effect of clozapine on mortality in people with schizophrenia. Clozapine is reserved for use in patients with treatment-resistant schizophrenia (TRS), but evidence of clozapine's effect on mortality exclusively within TRS samples is inconclusive. Hence, we aimed to investigate the effect of clozapine use on all-cause mortality in TRS patients. METHODS: A historical patient cohort sample of 2837 patients, who met criteria for TRS between 1 Jan 2008 and 1 Jan 2016, were selected from the South London and Maudsley NHS Foundation Trust (SLAM) electronic health records (EHR). The national Zaponex Treatment Access System (ZTAS) mandatory monitoring system linked to the SLAM EHR was used to distinguish which patients were initiated on clozapine (n = 1025). Cox proportional hazard models were used, adjusting for sociodemographics, clinical monitoring, mental and physical illness severity and functional status. RESULTS: After controlling for potential confounders, the protective effect of clozapine on all-cause mortality was significant (adjusted hazard ratio 0.61; 95% confidence interval 0.38-0.97; P = 0.04). CONCLUSIONS: Clozapine reduces the risk of mortality in patients who meet criteria for TRS. We provide further evidence that improving access to clozapine in TRS is likely to reduce the mortality gap in schizophrenia.

Long-term antipsychotic polypharmacy prescribing in secondary mental health care and the risk of mortality.

OBJECTIVES: To investigate the association between long-term antipsychotic polypharmacy use and mortality; and determine whether this risk varies by cause of death and antipsychotic dose. METHODS: Using data from a large anonymised mental healthcare database, we identified all adult patients with serious mental illness (SMI) who had been prescribed a single antipsychotic or polypharmacy, for six or more months between 2007 and 2014. Multivariable Cox regression models were constructed, adjusting for sociodemographic, socioeconomic, clinical factors and smoking, to examine the association between APP use and the risk of death. RESULTS: We identified 10 945 adults with SMI who had been prescribed long-term antipsychotic monotherapy (76.9%) or APP (23.1%). Patients on long-term APP had a small elevated risk of mortality, which was significant in some but not all models. The adjusted hazard ratios for death from natural and unnatural causes associated with APP were 1.2 (0.9-1.4, P = 0.111) and 1.1 (0.7-1.9, P = 0.619) respectively. The strengths of the associations between APP and mortality outcomes were similar after further adjusting for % BNF antipsychotic dose (P = 0.031) or olanzapine equivalence (P = 0.088). CONCLUSION: The findings suggest that the effect of long-term APP on mortality is not clear-cut, with limited evidence to indicate an association, even after controlling for the effect of dose.

Effectiveness of supplementary irrigant agitation with the Finisher GF Brush on the debridement of oval root canals instrumented with the Gentlefile or nickel titanium rotary instruments.

AIM: To examine the efficacy of a novel supplementary irrigant agitating brush (Finisher GF Brush, MedicNRG, Kibbutz Afikim, Israel) on the debridement of root canals prepared with a novel stainless steel rotary instrumentation system (Gentlefile; MedicNRG), or nickel titanium rotary instruments in oval root canals. METHODOLOGY: Mandibular premolars (n = 72) were selected and divided randomly into three experimental groups (n = 24) after microCT scanning: group 1, canal preparation to rotary NiTi size 20, .04 taper (R20); group 2, rotary NiTi to size 25, .04 taper (R25) and group 3, Gentlefile size 23, .04 taper (GF). Specimens were subdivided into two subgroups: subgroup A, syringe-and-needle irrigation (SNI); subgroup B, Finisher GF Brush (GB). Ten untreated canals served as controls. Specimens were processed for histological evaluation, and the remaining pulp tissue (RPT) was measured. Data were analysed using Mann-Whitney and Kruskal-Wallis tests (P = 0.05). RESULTS: All experimental groups had significantly less RPT than the control (P < 0.05). Group 3B (GF-GB) had significantly less RPT than groups 1B (R20-GB) and 2B (R25-GF; P < 0.05). When irrigated with SNI, there was no significant difference in the RPT between the three groups (P > 0.05). When instrumented with R20, there was no significant difference between SNI and GF (P < 0.05) whilst GB had significantly less RPT than SNI for R25 (P < 0.05). CONCLUSIONS: Supplementary irrigant agitation with the Finisher GF Brush improved the debridement of canals prepared with Gentlefile and size 25, .04 taper rotary NiTi. Root canal debridement did not significantly differ between the instruments when syringe irrigation was used.

Draft genome sequence of Sclerospora graminicola, the pearl millet downy mildew pathogen.

Sclerospora graminicola pathogen is the most important biotic production constraints of pearl millet in India, Africa and other parts of the world. We report a de novo whole genome assembly and analysis of pathotype 1, one of the most virulent pathotypes of S. graminicola from India. The draft genome assembly contained 299,901,251 bp with 65,404 genes. This study may help understand the evolutionary pattern of pathogen and aid elucidation of effector evolution for devising effective durable resistance breeding strategies in pearl millet.

Contributions of specific causes of death to lost life expectancy in severe mental illness.

The life expectancy gap between people with severe mental illness (SMI) and the general population persists and may even be widening. This study aimed to estimate contributions of specific causes of death to the gap. Age of death and primary cause of death were used to estimate life expectancy at birth for people with SMI from a large mental healthcare case register during 2007-2012. Using data for England and Wales in 2010, death rates in the SMI cohort for each primary cause of death category were replaced with gender- and age-specific norms for that cause. Life expectancy in SMI was then re-calculated and, thus, the contribution of that specific cause of death estimated. Natural causes accounted for 79.2% of lost life-years in women with SMI and 78.6% in men. Deaths from circulatory disorders accounted for more life-years lost in women than men (22.0% versus 17.4%, respectively), as did deaths from cancer (8.1% versus 0%), but the contribution from respiratory disorders was lower in women than men (13.7% versus 16.5%). For women, cancer contributed more in those with non-affective than affective disorders, while suicide, respiratory and digestive disorders contributed more in those with affective disorders. In men, respiratory disorders contributed more in non-affective disorders. Other contributions were similar between gender and affective/non-affective groups. Loss of life expectancy in people with SMI is accounted for by a broad range of causes of death, varying by gender and diagnosis. Interventions focused on multiple rather than individual causes of death should be prioritised accordingly.

Novel psychoactive substances: An investigation of temporal trends in social media and electronic health records.

BACKGROUND: Public health monitoring is commonly undertaken in social media but has never been combined with data analysis from electronic health records. This study aimed to investigate the relationship between the emergence of novel psychoactive substances (NPS) in social media and their appearance in a large mental health database. METHODS: Insufficient numbers of mentions of other NPS in case records meant that the study focused on mephedrone. Data were extracted on the number of mephedrone (i) references in the clinical record at the South London and Maudsley NHS Trust, London, UK, (ii) mentions in Twitter, (iii) related searches in Google and (iv) visits in Wikipedia. The characteristics of current mephedrone users in the clinical record were also established. RESULTS: Increased activity related to mephedrone searches in Google and visits in Wikipedia preceded a peak in mephedrone-related references in the clinical record followed by a spike in the other 3 data sources in early 2010, when mephedrone was assigned a 'class B' status. Features of current mephedrone users widely matched those from community studies. CONCLUSIONS: Combined analysis of information from social media and data from mental health records may assist public health and clinical surveillance for certain substance-related events of interest. There exists potential for early warning systems for health-care practitioners.

(18)F-FCWAY, a serotonin 1A receptor radioligand, is a substrate for efflux transport at the human blood-brain barrier.

Efflux transporters at the blood-brain barrier can decrease the entry of drugs and increase the removal of those molecules able to bypass the transporter. We previously hypothesized that (18)F-FCWAY, a radioligand for the serotonin 5-HT1A receptor, is a weak substrate for permeability glycoprotein (P-gp) based on its very early peak and rapid washout from human brain. To determine whether (18)F-FCWAY is a substrate for P-gp, breast cancer resistance protein (BCRP), and multidrug resistance protein (MRP1) - the three most prevalent efflux transporters at the blood-brain barrier - we performed three sets of experiments. In vitro, we conducted fluorescence-activated cell sorting (FACS) flow cytometry studies in cells over-expressing P-gp, BCRP, and MRP1 treated with inhibitors specific to each transporter and with FCWAY. Ex vivo, we measured (18)F-FCWAY concentration in plasma and brain homogenate of transporter knockout mice using γ-counter and radio-HPLC. In vivo, we conducted positron emission tomography (PET) studies to assess changes in humans who received (18)F-FCWAY during an infusion of tariquidar (2-4mg/kg iv), a potent and selective P-gp inhibitor. In vitro studies showed that FCWAY allowed fluorescent substrates to get into the cell by competitive inhibition of all three transporters at the cell membrane. Ex vivo measurements in knockout mice indicate that (18)F-FCWAY is a substrate only for P-gp and not BCRP. In vivo, tariquidar increased (18)F-FCWAY brain uptake in seven of eight subjects by 60-100% compared to each person's baseline. Tariquidar did not increase brain uptake via some peripheral mechanism, given that it did not significantly alter concentrations in plasma of the parent radioligand (18)F-FCWAY or its brain-penetrant radiometabolite (18)F-FC. These results show that (18)F-FCWAY is a weak substrate for efflux transport at the blood-brain barrier; some radioligand can enter brain, but its removal is hastened by P-gp. Although (18)F-FCWAY is not ideal for measuring 5-HT1A receptors, it demonstrates that weak substrate radioligands can be useful for measuring both increased and decreased function of efflux transporters, which is not possible with currently available radioligands such as (11)C-loperamide and (11)C-verapamil that are avid substrates for transporters.

Services for people at high risk improve outcomes in patients with first episode psychosis.

OBJECTIVE: About one-third of patients referred to services for people at high risk for psychosis may have already developed a first episode of psychosis (FEP). We compared clinical outcomes in FEP patients who presented to either high risk or conventional mental health services. METHOD: Retrospective study comparing duration of hospital admission, referral-to-diagnosis time, need for compulsory hospital admission and frequency of admission in patients with FEP who initially presented to a high-risk service (n = 164) to patients with FEP who initially presented to conventional mental health services (n = 2779). Regression models were performed, controlling for several confounders. RESULTS: FEP patients who had presented to a high-risk service spent 17 fewer days in hospital [95% CI: -33.7 to (-0.3)], had a shorter referral-to-diagnosis time [B coefficient -74.5 days, 95% CI: -101.9 to -(47.1)], a lower frequency of admission [IRR: 0.49 (95% CI: 0.39-0.61)] and a lower likelihood of compulsory admission [OR: 0.52 (95% CI: 0.34-0.81)] in the 24 months following referral, as compared to FEP patients who were first diagnosed at conventional services. CONCLUSION: Services for people at high risk for psychosis are associated with better clinical outcomes in patients who are already psychotic.

Transposition of great arteries with natural partial Senning: A rare case report.

The association of transposition of the great arteries (TGA) and anomalous pulmonary venous connection is extremely rare. Children with transposition of the great arteries improved dramatically with the advent of the atrial repair. In this report, we describe a 40-day old male infant with TGA and associated anomalous pulmonary venous connection who presented with the history of cyanosis and hurried breathing. This patient underwent successful balloon atrial septostomy and discharged with uneventful recovery.

Increased permeability-glycoprotein inhibition at the human blood-brain barrier can be safely achieved by performing PET during peak plasma concentrations of tariquidar.

UNLABELLED: The permeability-glycoprotein (P-gp) efflux transporter is densely expressed at the blood-brain barrier, and its resultant spare capacity requires substantial blockade to increase the uptake of avid substrates, blunting the ability of investigators to measure clinically meaningful alterations in P-gp function. This study, conducted in humans, examined 2 P-gp inhibitors (tariquidar, a known inhibitor, and disulfiram, a putative inhibitor) and 2 routes of administration (intravenous and oral) to maximally increase brain uptake of the avid and selective P-gp substrate (11)C-N-desmethyl-loperamide (dLop) while avoiding side effects associated with high doses of tariquidar. METHODS: Forty-two (11)C-dLop PET scans were obtained from 37 healthy volunteers. PET was performed with (11)C-dLop under the following 5 conditions: injected under baseline conditions without P-gp inhibition, injected 1 h after intravenous tariquidar infusion, injected during intravenous tariquidar infusion, injected after oral tariquidar, and injected after disulfiram. (11)C-dLop uptake was quantified with kinetic modeling using metabolite-corrected arterial input function or by measuring the area under the time-activity curve in the brain from 10 to 30 min. RESULTS: Neither oral tariquidar nor oral disulfiram increased brain uptake of (11)C-dLop. Injecting (11)C-dLop during tariquidar infusion, when plasma tariquidar concentrations reach their peak, resulted in a brain uptake of the radioligand approximately 5-fold greater than baseline. Brain uptake was similar with 2 and 4 mg of intravenous tariquidar per kilogram; however, the lower dose was better tolerated. Injecting (11)C-dLop after tariquidar infusion also increased brain uptake, though higher doses (up to 6 mg/kg) were required. Brain uptake of (11)C-dLop increased fairly linearly with increasing plasma tariquidar concentrations, but we are uncertain whether maximal uptake was achieved. CONCLUSION: We sought to increase the dynamic range of P-gp function measured after blockade. Performing (11)C-dLop PET during peak plasma concentrations of tariquidar, achieved with concurrent administration of intravenous tariquidar, resulted in greater P-gp inhibition at the human blood-brain barrier than delayed administration and allowed the use of a lower, more tolerable dose of tariquidar. On the basis of prior monkey studies, we suspect that plasma concentrations of tariquidar did not fully block P-gp; however, higher doses of tariquidar would likely be associated with unacceptable side effects.

Development of SCAR marker associated with downy mildew disease resistance in pearl millet (Pennisetum glaucum L.).

Pearl Millet is an important crop coarse grain cereal crop in the semi arid tropics which is extremely susceptible to oomycete plant pathogen Sclerospora graminicola causing downy mildew (DM) disease. The aim of the current study is to breed resistance against downy mildew disease into high yielding cultivars of pearl millet. Hence, in the present work a sequence characterized amplified region (SCAR) marker was developed as a molecular screening tool to identify DM resistance source and presented here. Of the 27 inter simple sequence repeats (ISSR) decamer primers used to identify polymorphism amongst pearl millet genotypes ICMR-01007 (P1) and ICMR-01004 (P2) and their populations (F1 and F2), only one primer pair ISSR-22 produced polymorphic bands on ICMR-01004 producing 1.4 kb size. The PCR amplification of 1.4 kb band was found tightly linked to the resistant line of ICMR-01004 and also in F2 segregation population was in the ratio 3:1. This band was cloned, sequenced and candidate SCAR primer (SCAR ISSR 863 ) was designed. Segregant analysis of their F2 progeny revealed that the SCAR ISSR 863 marker was linked to downy mildew resistance linkage group (χ(2) 3:1 = 0.86, P = 0.22) with a genetic distance of 0.72 cM. This SCAR marker was further validated using diverse pearl millet lines of India and Africa. Results indicated that the SCAR ISSR 863 band was amplified in all the seven resistant lines and were absent in five susceptible lines. The confirmation of the ISSR-derived SCAR marker in different genetic backgrounds of pearl millet lines suggests that this marker can be exploited for DM resistance screening in pearl millet breeding programs.

Characterization of fast-decaying PET radiotracers solely through LC-MS/MS of constituent radioactive and carrier isotopologues.

BACKGROUND: The characterization of fast-decaying radiotracers that are labeled with carbon-11 (t1/2 = 20.38 min), including critical measurement of specific radioactivity (activity per mole at a specific time) before release for use in positron-emission tomography (PET), has relied heavily on chromatographic plus radiometric measurements, each of which may be vulnerable to significant errors. Thus, we aimed to develop a mass-specific detection method using sensitive liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS) for identifying 11C-labeled tracers and for verifying their specific radioactivities. METHODS: The LC-MS/MS was tuned and set up with methods to generate and measure the product ions specific for carbon-11 species and M + 1 carrier (predominantly the carbon-13 isotopologue) in four 11C-labeled tracers. These radiotracers were synthesized and then analyzed before extensive carbon-11 decay. The peak areas of carbon-11 species and M + 1 carrier from the LC-MS/MS measurement and the calculated abundances of carbon-12 carrier and M + 1 radioactive species gave the mole fraction of carbon-11 species in each sample. This value upon multiplication with the theoretical specific radioactivity of carbon-11 gave the specific radioactivity of the radiotracer. RESULTS: LC-MS/MS of each 11C-labeled tracer generated the product ion peaks for carbon-11 species and M + 1 carrier at the expected LC retention time. The intensity of the radioactive peak diminished as time elapsed and was undetectable after six half-lives of carbon-11. Measurements of radiotracer-specific radioactivity determined solely by LC-MS/MS at timed intervals gave a half-life for carbon-11 (20.43 min) in excellent agreement with the value obtained radiometrically. Additionally, the LC-MS/MS measurement gave specific radioactivity values (83 to 505 GBq/µmol) in good agreement with those from conventional radiometric methods. CONCLUSIONS: 11C-Labeled tracers were characterized at a fundamental level involving isolation and mass detection of extremely low-abundance carbon-11 species along with the M + 1 carrier counterpart. This LC-MS/MS method for characterizing fast-decaying radiotracers is valuable in both the development and production of PET radiopharmaceuticals.

Rhizosphere fungus Penicillium chrysogenum promotes growth and induces defence-related genes and downy mildew disease resistance in pearl millet.

Susceptible pearl millet seeds (cv 7042S) were treated with the plant growth promoting fungus Penicillium chrysogenum (PenC-JSB9) at 1 × 10(8) spores·ml(-1) to examine mRNA expression profiles of five defence responsive genes and test its ability to induce resistance to downy mildew caused by Sclerospora graminicola. PenC-JSB9 treatment at 1 × 10(8) CFU·ml(-1) for 6 h significantly enhanced seed germination (9.8- 89%), root length (4.08% to 5.1 cm), shoot length (18.9% to 7.77 cm) and reduced disease incidence (28%) in comparison with untreated controls. In planta colonisation of PenC-JSB9 showed that all three root segments (0-6 cm) and soil dilutions incubated on PDA produced extensive mycelial growth, however colonisation frequency of PenC-JSB9 was significantly higher in soil than in root segments. Spatiotemporal studies revealed that induction of resistance was triggered as early as 24 h and a minimum 2-3 days was optimal for total resistance to build up between inducer treatment and challenge inoculation in both experiments. In Northern blot analysis, transcript accumulation of resistant and PenC-JSB9 induced susceptible cultivars showed higher basal levels of defence gene expression than non-pretreated susceptible controls. Transcript accumulation in resistant seedlings challenge-inoculated with the pathogen showed maximum expression of CHS (3.5-fold increase) and Pr-1a (threefold increase) at 24 and 12 h, respectively. While PenC-JSB9 pretreated susceptible seedlings challenge-inoculated showed rapid and enhanced expression of LOX and POX at 48 h and for CHT at 24 h, whereas non-pretreated susceptible seedlings after pathogen inoculation showed weak expression of hybridised defence genes. Enhanced activation of defence genes by PenC-JSB9 suggests its role in elevated resistance against S. graminicola.