Analysis of 2'-hydroxyflavanone (2HF) in mouse whole blood by HPLC-MS/MS for the determination of pharmacokinetic parameters.

Given the lack of investments, structure, and difficulty of metabolite isolation, promising natural product studies do not progress to preclinical studies, such as pharmacokinetics. 2'-Hydroxyflavanone (2HF) is a flavonoid that has shown promising results in different types of cancer and leishmaniasis. For accurate quantification of 2HF in BALB/c mouse blood, a validated HPLC-MS/MS method was developed. Chromatographic analysis was performed using C18 (5µm, 150 mm × 4.6 mm). The mobile phase consisted of water containing 0.1% formic acid, acetonitrile, and methanol (35/52/13 v/v/v) at a flow rate and total running time of 0.8 mL/min and 5.50 min, respectively, with an injection volume of 20 µL. 2HF was detected by electrospray ionization in negative mode (ESI-) using multiple reaction monitoring (MRM). The validated bioanalytical method showed satisfactory selectivity without significant interference for the 2HF and IS. In addition, the concentration range between 1 and 250 ng/mL showed good linearity (r = 0.9969). The method showed satisfactory results for the matrix effect. Precision and accuracy intervals varied between 1.89% and 6.76% and 95.27% and 100.77%, respectively, fitting the criteria. No degradation of 2HF in the biological matrix was observed since stability under freezing and thawing conditions, short duration, postprocessing, and long duration showed deviations less than 15%. Once validated, the method was successfully applied in a 2HF oral pharmacokinetic study with mouse blood, and the pharmacokinetic parameters were determined. 2HF demonstrated a Cmax of 185.86 ng/mL, a Tmax of 5 min, and a half-life (T1/2) of 97.52 min.

The use of photobiomodulation therapy or LED and mineral trioxide aggregate improves the repair of complete tibial fractures treated with wire osteosynthesis in rodents.

The repair of large bone defects is lengthy and complex. Both biomaterials and phototherapy have been used to improve bone repair. We aimed to describe histologically the repair of tibial fractures treated by wiring (W), irradiated or not, with laser (λ780 nm, 70 mW, CW, spot area of 0.5 cm2, 20.4 J/cm2 (4 × 5.1 J/cm2, Twin Flex Evolution®, MM Optics, Sao Carlos, SP, Brazil) per session, 300 s, 142.8 J/cm2 per treatment) or LED (λ850 ± 10 nm, 150 mW, spot area of 0.5 cm2, 20.4 J/cm2 per session, 64 s, 142.8 J/cm2 per treatment, Fisioled®, MM Optics, Sao Carlos, Sao Paulo, Brazil) and associated or not to the use of mineral trioxide aggregate (MTA, Angelus®, Londrina, PR, Brazil). Inflammation was discrete on groups W and W + LEDPT and absent on the others. Phototherapy protocols started immediately before suturing and repeated at every other day for 15 days. Collagen deposition intense on groups W + LEDPT, W + BIO-MTA + LaserPT and W + BIO-MTA + LEDPT and discrete or moderate on the other groups. Reabsorption was discrete on groups W and W + LEDPT and absent on the other groups. Neoformation varied greatly between groups. Most groups were partial and moderately filed with new-formed bone (W, W + LaserPT, W + LEDPT, W + BIO-MTA + LEDPT). On groups W + BIO-MTA and W + BIO-MTA + LaserPT bone, neoformation was intense and complete. Our results are indicative that the association of MTA and PBMT (λ = 780 nm) improves the repair of complete tibial fracture treated with wire osteosynthesis in a rodent model more efficiently than LED (λ = 850 ± 10 nm).

Pharmacological profiling of JME-173, a novel mexiletine derivative combining dual anti-inflammatory/anti-spasmodic functions and limited action in Na+ channels.

Existing evidence suggests that the local anaesthetic mexiletine can be beneficial for patients with asthma. However, caution is required since anaesthesia of the airways inhibits protective bronchodilator neuronal reflexes, limiting applications in conditions of hyperirritable airways. Here, we describe the synthesis of a new series of mexiletine analogues, which were screened for reduced activity in Na+ channels and improved smooth muscle relaxant effects, that were evaluated using the patch-clamp technique and an isolated tracheal organ bath, respectively. JME-173 (1-(4-bromo-3,5-dimethylphenoxy)propan-2-amine) was the most effective among the four mexiletine analogues investigated. JME-173 was then studied in vivo using a murine model of lung inflammation induced by cigarette smoke (CS) and in vitro using neutrophil chemotaxis and mast cell degranulation assays. Finally, the JME-173 pharmacokinetic profile was assessed using HPLC-MS/MS bioanalytical method. JME-173 directly inhibited IL-8 (CXCL8)- and FMLP-induced human neutrophil chemotaxis and allergen-induced mast cell degranulation. After oral administration 1 h before CS exposure, JME-173 (50 mg/kg) strongly reduced the increased number of macrophages and neutrophils recovered in the bronchoalveolar effluent without altering lymphocyte counts. Pharmacokinetic experiments of JME-173 (10 mg/kg, orally) showed values of maximum concentration (Cmax), maximum time (Tmax), area under the blood concentration-time curve (AUC0-t) and area under the blood concentration-time curve from 0-Inf (AUC0-inf) of 163.3 ± 38.3 ng/mL, 1.2 ± 0.3 h, 729.4 ± 118.3 ng*h/ml and 868.9 ± 117.1 ng*h/ml (means ± S.E.M.), respectively. Collectively, these findings suggest that JME-173 has the potential to be an effective oral treatment for diseases associated with bronchoconstriction and inflammation.

Sofosbuvir shows a protective effect against vertical transmission of Zika virus and the associated congenital syndrome in rhesus monkeys.

The outbreaks of Zika virus (ZIKV) infection in Brazil, 2015-2016, were associated with severe congenital malformations. Our translational study aimed to test the efficacy of the antiviral agent sofosbuvir (SOF) against vertical transmission of ZIKV and the associated congenital syndrome (CZS), using a rhesus monkey model. Eight pregnant macaques were successfully infected during the organogenesis phase with a Brazilian ZIKV strain; five of them received SOF from two to fifteen days post-infection. Both groups of dams showed ZIKV-associated clinical signals, detectable ZIKV RNA in several specimens, specific anti-ZIKV IgM and IgG antibodies, and maternal neutralizing antibodies. However, malformations occurred only among non-treated dam offspring. Compared to non-treated animals, all SOF-treated dams had a shorter ZIKV viremia and four of five neonates had undetectable ZIKV RNA in blood and tissue samples. These results support further clinical evaluations aiming for the prevention of CZS.

Raman spectroscopic study of the effect of the use of laser/LED phototherapy on the repair of complete tibial fracture treated with internal rigid fixation.

This study aimed to assess the repair of complete surgical tibial fractures fixed with internal rigid fixation (IRF) associated or not to the use of mineral trioxide aggregate (MTA) cement and treated or not with laser (λ = 780 nm, infrared) or LED (λ = 850 ±â€¯10 nm, infrared) lights, 142.8 J/cm2 per treatment, by means of Raman spectroscopy. Open surgical tibial fractures were created on 18 rabbits (6 groups of 3 animals per group, ∼8 months old) and fractures were fixed with IRF. Three groups were grafted with MTA. The groups of IRF and IRF + MTA that received laser or LED were irradiated every other day during 15 days. Animals were sacrificed after 30 days, being the tibia surgically removed. Raman spectra were collected via the probe at the defect site in five points, resulting in 15 spectra per group (90 spectra in the dataset). Spectra were collected at the same day to avoid changes in laser power and experimental setup. The ANOVA general linear model showed that the laser irradiation of tibial bone fractures fixed with IRF and grafted with MTA had significant influence in the content of phosphate (peak ∼960 cm-1) and carbonated (peak ∼1,070 cm-1) hydroxyapatites as well as collagen (peak 1,452 cm-1). Also, peaks of calcium carbonate (1,088 cm-1) were found in the groups grafted with MTA. Based on the Raman spectroscopic data collected in this study, MTA has been shown to improve the repair of complete tibial fractures treated with IRF, with an evident increase of collagen matrix synthesis, and development of a scaffold of hydroxyapatite-like calcium carbonate with subsequent deposition of phosphate hydroxyapatite.

The use of computed tomography images as a prognostic marker in critically ill cancer patients.

BACKGROUND & AIMS: In the intensive care unit (ICU) setting, body composition is typically estimated through anthropometry, which does not specifically quantify skeletal muscle (SM). Recent findings have shown that computed tomography (CT) is a useful tool to identify low SM in critically ill patients, which in turn is associated with poor clinical outcomes. So, the present study aims at comparing low SM assessed by CT to BMI anthropometric data and its association with outcomes in critically ill patients. METHODS: Observational study was used, which included >18 year-old patients, with over 72 h of ICU length of stay, who had an abdominal CT at ICU admission. Demographic, body mass index (BMI), hospital outcomes and abdominal CT data (SM and adipose tissue at the 3rd lumbar vertebrae) were collected for analysis. ROC curve optimal stratification analysis for hospital mortality was applied to classify people into low SM (sarcopenic) versus normal SM (non-sarcopenic). A Cox regression was applied to find independent associations between sarcopenia and 30-day survival. RESULTS: The study involved 99 patients, 56% male, mean-age of 61.6 years old, BMI 24.19 ± 4.49 kg/m2; hospital mortality was 26%. According to BMI, 19.4% of the patients were underweight. However, a poor correlation was observed between BMI and SM index by CT: R2 = 0.39, P < 0.001. The cutoff point for determining sarcopenia by CT was 41.2 cm2/m2 (sensitivity 70%, specificity 69.5%, AUC 70.3) for both sexes. The sarcopenia diagnosis by CT as nutritional evaluation parameters was correlated with malnutrition BMI diagnosis in only 35.5%. When compared to non-sarcopenic patients, those with sarcopenia presented worse 30-day survival adjusted by age and SAPS 3 (HR = 2.74, 95%CI = 1.02-7.35, P = 0.04), higher hospital mortality (41.9% vs 14.6%, P = 0.006) and ICU non-infectious complications (76.7% vs 52.1%, P = 0.016). CONCLUSION: Sarcopenia assessed by abdominal CT demonstrated low correlation with BMI and was a risk factor for lower 30-day survival, higher hospital mortality and higher complications in critically ill patients.

Laser/LED phototherapy on the repair of tibial fracture treated with wire osteosynthesis evaluated by Raman spectroscopy.

The aim of the present study was to assess, by means of Raman spectroscopy, the repair of complete surgical tibial fractures fixed with wire osteosynthesis (WO) treated or not with infrared laser (λ780 nm) or infrared light emitting diode (LED) (λ850 ± 10 nm) lights, 142.8 J/cm2 per treatment, associated or not to the use of mineral trioxide aggregate (MTA) cement. Surgical tibial fractures were created on 18 rabbits, and all fractures were fixed with WO and some groups were grafted with MTA. Irradiated groups received lights at every other day during 15 days, and all animals were sacrificed after 30 days, being the tibia removed. The results showed that only irradiation with either laser or LED influenced the peaks of phosphate hydroxyapatite (~ 960 cm-1). Collagen (~ 1450 cm-1) and carbonated hydroxyapatite (~ 1070 cm-1) peaks were influenced by both the use of MTA and the irradiation with either laser or LED. It is concluded that the use of either laser or LED phototherapy associated to MTA cement was efficacious on improving the repair of complete tibial fractures treated with wire osteosynthesis by increasing the synthesis of collagen matrix and creating a scaffold of calcium carbonate (carbonated hydroxyapatite-like) and the subsequent deposition of phosphate hydroxyapatite.

Diagnostic utility of DREAM gene mRNA levels in thyroid tumours.

OBJECTIVE: The transcriptional repressor DREAM is involved in thyroid-specific gene expression, thyroid enlargement and nodular development, but its clinical utility is still uncertain. In this study we aimed to investigate whether DREAM mRNA levels differ in different thyroid tumors and how this possible difference would allow the use of DREAM gene expression as molecular marker for diagnostic and/or prognosis purpose. MATERIALS AND METHODS: We quantified DREAM gene mRNA levels and investigated its mutational status, relating its expression and genetic changes to diagnostic and prognostic features of 200 thyroid tumors, being 101 malignant [99 papillary thyroid carcinomas (PTC) and 2 anaplastic thyroid carcinomas] and 99 benign thyroid lesions [49 goiter and 50 follicular adenomas (FA)]. RESULTS: Levels of mRNA of DREAM gene were higher in benign (0.7909 ± 0.6274 AU) than in malignant (0.3373 ± 0.6274 AU) thyroid lesions (p < 0.0001). DREAM gene expression was able to identify malignancy with 66.7% sensitivity, 85.4% specificity, 84.2% positive predictive value (PPV), 68.7% negative predictive value (NPV), and 75.3% accuracy. DREAM mRNA levels were also useful distinguishing the follicular lesions FA and FVPTC with 70.2% sensitivity, 73.5% specificity, 78.5% PPV, 64.1% NPV, and 71.6% accuracy. However, DREAM gene expression was neither associated with clinical features of tumor aggressiveness, nor with recurrence or survival. Six different genetic changes in non-coding regions of DREAM gene were also found, not related to DREAM gene expression or tumor features. CONCLUSION: We suggest that DREAM gene expression may help diagnose thyroid nodules, identifying malignancy and characterizing follicular-patterned thyroid lesions; however, it is not useful as a prognostic marker.

Diagnostic utility of DREAM gene mRNA levels in thyroid tumours

ABSTRACT Objective The transcriptional repressor DREAM is involved in thyroid-specific gene expression, thyroid enlargement and nodular development, but its clinical utility is still uncertain. In this study we aimed to investigate whether DREAM mRNA levels differ in different thyroid tumors and how this possible difference would allow the use of DREAM gene expression as molecular marker for diagnostic and/or prognosis purpose. Materials and methods We quantified DREAM gene mRNA levels and investigated its mutational status, relating its expression and genetic changes to diagnostic and prognostic features of 200 thyroid tumors, being 101 malignant [99 papillary thyroid carcinomas (PTC) and 2 anaplastic thyroid carcinomas] and 99 benign thyroid lesions [49 goiter and 50 follicular adenomas (FA)]. Results Levels of mRNA of DREAM gene were higher in benign (0.7909 ± 0.6274 AU) than in malignant (0.3373 ± 0.6274 AU) thyroid lesions (p < 0.0001). DREAM gene expression was able to identify malignancy with 66.7% sensitivity, 85.4% specificity, 84.2% positive predictive value (PPV), 68.7% negative predictive value (NPV), and 75.3% accuracy. DREAM mRNA levels were also useful distinguishing the follicular lesions FA and FVPTC with 70.2% sensitivity, 73.5% specificity, 78.5% PPV, 64.1% NPV, and 71.6% accuracy. However, DREAM gene expression was neither associated with clinical features of tumor aggressiveness, nor with recurrence or survival. Six different genetic changes in non-coding regions of DREAM gene were also found, not related to DREAM gene expression or tumor features. Conclusion We suggest that DREAM gene expression may help diagnose thyroid nodules, identifying malignancy and characterizing follicular-patterned thyroid lesions; however, it is not useful as a prognostic marker.

Computed tomography-derived skeletal muscle index: A novel predictor of frailty and hospital length of stay after transcatheter aortic valve replacement.

To determine the prevalence of low skeletal muscle mass in patients undergoing transcatheter aortic valve replacement (TAVR) and whether skeletal muscle mass measured from preoperative computed tomography (CT) images provides value in predicting postoperative length of stay (LOS). BACKGROUND: There are limited data on the use of body composition as a frailty measure in TAVR patients and no studies have determined if this measure predicts LOS. METHODS: We studied 104 consecutive patients who underwent TAVR at Tallahassee Memorial Hospital from 2012 to 2016. Patient demographics, standard frailty measures (hand grip, albumin, and 5-m walk test), clinical comorbidities, echocardiographic data, and Valve Academic Research Consortium II major complications were recorded prospectively. Skeletal muscle index (SMI) [skeletal muscle mass cross-sectional area at L3/height2] was measured from CT images using Slice-O-Matic software (Tomovision, Montreal, Quebec, Canada). Clinical outcomes were assessed and multivariate methods used to determine predictors of LOS. RESULTS: Sarcopenia was prevalent in men (83%) and women (56%). Patients who suffered from a major complication had significantly longer length of stay (13 vs 4.6days, P<.0001). Skeletal muscle index correlated with age, sex, body mass index, handgrip strength, and previous coronary artery bypass graft surgery, but not major complications. A multivariate model including all univariate predictors of LOS showed SMI, major complications, transapical access, atrial fibrillation, and chronic obstructive pulmonary syndrome as independent predictors of LOS. For every 14-cm2/m2 increase in SMI, there was a 1-day reduction in LOS. None of the standard measures of frailty predicted LOS. CONCLUSIONS: Skeletal muscle index, a measure of sarcopenia readily determined from pre-TAVR CT scans, independently predicts TAVR LOS better than standard frailty testing. Further evaluation of SMI as a frailty measure after TAVR and other cardiovascular procedures is warranted.

Preclinical pharmacokinetic evaluation of praziquantel loaded in poly (methyl methacrylate) nanoparticle using a HPLC-MS/MS.

Praziquantel (PZQ) is the drug recommended by the World Health Organization for treatment of schistosomiasis. However, the treatment of children with PZQ tablets is complicated due to difficulties to adapt the dose and the extremely bitter taste of PZQ. For this reason, poly (methyl methacrylate) nanoparticles loaded with Praziquantel (PZQ-NP) were developed for preparation of a new formulation to be used in the suspension form. For this reason, the main aim of the present study was to evaluate the pharmacokinetic (PK) profile of PZQ-NP, through HPLC-MS/MS assays. Analyses were performed with an Omnisphere C18 column (5.0 µm×4.6 mm×150.0 mm), using a mixture of an aqueous solution containing 0.1 wt% of formic acid and methanol (15:85-v/v) as the mobile phase at a flow rate of 0.800mL/min. Detection was performed with a hybrid linear ion-trap triple quadrupole mass spectrometer with multiple reactions monitoring in positive ion mode via electrospray ionization. The monitored transitions were m/z 313.18>203.10 for PZQ and m/z 285.31>193.00 for the Internal Standard. The method was validated with the quantification limit of 1.00 ng/mL, requiring samples of 25 µL for analyses. Analytic responses were calibrated with known concentration data, leading to correlation coefficients (r) higher than 0.99. Validation performed with rat plasma showed that PZQ was stable for at least 10 months when stored below -70 °C (long-term stability), for at least 17 h when stored at room temperature (RT, 22 °C) (short-term stability), for at least 47 h when stored at room temperature in auto-sampler vials (post-preparative stability) and for at least 8 successive freeze/thaw cycles at -70 °C. For PK assays, Wistar rats, weighing between 200 and 300 g were used. Blood samples were collected from 0 to 24 h after oral administration of single doses of 60 mg/kg of PZQ-NP or raw PZQ (for the control group). PZQ was extracted from plasma by liquid-liquid extraction with terc-butyl methyl ether. The values obtained for maximum concentration (C(max)) and area under curve (AUC) for the PZQ-NP group were about 3 times smaller than the respective values obtained for the control group. However, the time for achieving maximum concentration (T(max)), the elimination constant (Ke) and the half-life time of elimination (T(½ß)) were not statistically different. These results suggest that PZQ absorption is probably the rate-limiting step for obtainment of better PK parameters for PZQ-NP. Thus, further studies are needed to understand both the PZQ-NP absorption mechanisms and the drug diffusion process through the polymer matrix in vivo, in order to improve the PZQ-NP release profile.

Thermodynamic-based retention time predictions of endogenous steroids in comprehensive two-dimensional gas chromatography.

This work evaluates the application of a thermodynamic model to comprehensive two-dimensional gas chromatography (GC × GC) coupled with time-of-flight mass spectrometry for anabolic agent investigation. Doping control deals with hundreds of drugs that are prohibited in sports. Drug discovery in biological matrices is a challenging task that requires powerful tools when one is faced with the rapidly changing designer drug landscape. In this work, a thermodynamic model developed for the prediction of both primary and secondary retention times in GC × GC has been applied to trimethylsilylated hydroxyl (O-TMS)- and methoxime-trimethylsilylated carbonyl (MO-TMS)-derivatized endogenous steroids. This model was previously demonstrated on a pneumatically modulated GC × GC system, and is applied for the first time to a thermally modulated GC × GC system. Preliminary one-dimensional experiments allowed the calculation of thermodynamic parameters (ΔH, ΔS, and ΔC p ) which were successfully applied for the prediction of the analytes' interactions with the stationary phases of both the first-dimension column and the second-dimension column. The model was able to predict both first-dimension and second-dimension retention times with high accuracy compared with the GC × GC experimental measurements. Maximum differences of -8.22 s in the first dimension and 0.4 s in the second dimension were encountered for the O-TMS derivatives of 11ß-hydroxyandrosterone and 11-ketoetiocholanolone, respectively. For the MO-TMS derivatives, the largest discrepancies were from testosterone (9.65 ) for the first-dimension retention times and 11-keto-etiocholanolone (0.4 s) for the second-dimension retention times.

Nanobiosensors exploiting specific interactions between an enzyme and herbicides in atomic force spectroscopy.

The development of sensitive methodologies for detecting agrochemicals has become important in recent years due to the increasingly indiscriminate use of these substances. In this context, nanosensors based on atomic force microscopy (AFM) tips are useful because they provide higher sensitivity with operation at the nanometer scale. In this paper we exploit specific interactions between AFM tips functionalized with the enzyme acetolactate synthase (ALS) to detect the ALS-inhibitor herbicides metsulfuron-methyl and imazaquin. Using atomic force spectroscopy (AFS) we could measure the adhesion force between tip and substrate, which was considerably higher when the ALS-functionalized tip (nanobiosensor) was employed. The increase was approximately 250% and 160% for metsulfuron-methyl and imazaquin, respectively, in comparison to unfunctionalized probes. We estimated the specific enzyme-herbicide force by assuming that the measured force comprises an adhesion force according to the Johnson-Kendall-Roberts (JKR) model, the capillary force and the specific force. We show that the specific, biorecognition force plays a crucial role in the higher sensitivity of the nanobiosensor, thus opening the way for the design of similarly engineered tips for detecting herbicides and other analytes.

Nanobiosensors based on chemically modified AFM probes: a useful tool for metsulfuron-methyl detection.

The use of agrochemicals has increased considerably in recent years, and consequently, there has been increased exposure of ecosystems and human populations to these highly toxic compounds. The study and development of methodologies to detect these substances with greater sensitivity has become extremely relevant. This article describes, for the first time, the use of atomic force spectroscopy (AFS) in the detection of enzyme-inhibiting herbicides. A nanobiosensor based on an atomic force microscopy (AFM) tip functionalised with the acetolactate synthase (ALS) enzyme was developed and characterised. The herbicide metsulfuron-methyl, an ALS inhibitor, was successfully detected through the acquisition of force curves using this biosensor. The adhesion force values were considerably higher when the biosensor was used. An increase of ~250% was achieved relative to the adhesion force using an unfunctionalised AFM tip. This considerable increase was the result of a specific interaction between the enzyme and the herbicide, which was primarily responsible for the efficiency of the nanobiosensor. These results indicate that this methodology is promising for the detection of herbicides, pesticides, and other environmental contaminants.

Comparative analysis of expressed genes from cacao meristems infected by Moniliophthora perniciosa.

BACKGROUND AND AIMS: Witches' broom disease is caused by the hemibiotrophic basidiomycete Moniliophthora perniciosa, and is one of the most important diseases of cacao in the western hemisphere. Because very little is known about the global process of such disease development, expressed sequence tags (ESTs) were used to identify genes expressed during the Theobroma cacao-Moniliophthora perniciosa interaction. METHODS: Two cDNA libraries corresponding to the resistant (RT) and susceptible (SP) cacao-M. perniciosa interactions were constructed from total RNA, using the DB SMART Creator cDNA library kit (Clontech). Clones were randomly selected, sequenced from the 5' end and analysed using bioinformatics tools including in silico analysis of the differential gene expression. KEY RESULTS: A total of 6884 ESTs were generated from the RT and SP cDNA libraries. These ESTs were composed of 2585 singlets and 341 contigs for a total of 2926 non-redundant sequences. The redundancy of the libraries was low and their specificity high when compared with the few other cacao libraries already published. Sequence analysis allowed the assignment of a putative functional category for 54 % of sequences, whereas approx. 22 % of sequences corresponded to unknown function and approx. 24 % of sequences did not show any significant similarity with other proteins present in the database. Despite the similar overall distribution of the sequences in functional categories between the two libraries, qualitative differences were observed. Genes involved during the defence response to pathogen infection or in programmed cell death were identified, such as pathogenesis related-proteins, trypsin inhibitor or oxalate oxidase, and some of them showed an in silico differential expression between the resistant and the susceptible interactions. CONCLUSIONS: As far as is known this is the first EST resource from the cacao-M. perniciosa interaction and it is believed that it will provide a significant contribution to the understanding of the molecular mechanisms of the resistance and susceptibility of cacao to M. perniciosa, to develop strategies to control witches' broom, and as a source of polymorphism for molecular marker development and marker-assisted selection.

Equine protease inhibitor system as a marker for the diagnosis of chronic obstructive pulmonary disease (COPD)

The protease inhibitor system (PI) was investigated to ascertain if it can be used as a marker of chronic obstructive pulmonary disease (COPD) in thoroughbred horses. Serum samples were taken from healthy thoroughbreds (n = 13) and those diagnosed as having COPD (n = 24) or inflammatory airway disease (IAD, n = 38) as well as from 3,600 undiagnosed thoroughbred horses. PI allelic and genotypic frequencies were estimated using protein electrophoresis on starch and polyacrylamide gels. The four groups of horses showed high genotypic similarity and none of the observed alleles or genotypes of the equine PI system were found to be associated with COPD.