RAPD-based genetic linkage maps of yellow passion fruit (Passiflora edulis Sims. f. flavicarpa Deg.).

A single cross between two clones of passion fruit (Passiflora edulis Sims. f. flavicarpa Deg., 2n = 18) was selected for genetic mapping. The mapping population was composed of 90 F1 plants derived from a cross between 'IAPAR 123' (female parent) and 'IAPAR 06' (male parent). A total of 380 RAPD primers were analyzed according to two-way pseudo-testcross mapping design. The linkage analysis was performed using Mapmaker version 3.0 with LOD 4.0 and a maximum recombination fraction (theta) of 0.30. Map distances were estimated using the Kosambi mapping function. Linkage maps were constructed with 269 loci (2.38 markers/primer), of which 255 segregated 1:1, corresponding to a heterozygous state in one parent and null in the other. The linkage map for 'IAPAR123' consisted of 135 markers. A total of nine linkage groups were assembled covering 727.7 cM, with an average distance of 11.20 cM between framework loci. The sizes of the linkage groups ranged from 56 to 144.6 cM. The linkage map for 'IAPAR 06' consisted of 96 markers, covering 783.5 cM. The average distance between framework loci was 12.2 cM. The length of the nine linkage groups ranged from 20.6 to 144.2 cM. On average, both maps provided 61% genome coverage. Twenty-four loci (8.9%) remained unlinked. Among their many applications, these maps are a starting point for the identification of quantitative trait loci for resistance to the main bacterial disease affecting passion fruit orchards in Brazil, caused by Xanthomonas campestris pv. passiflorae, because parental genotypes exhibit diverse responses to bacterial inoculation.     

Yarrowia lipolytica lipase production enhanced by increased air pressure

Aims: To study the cellular growth and morphology of Yarrowia lipolytica W29 and its lipase and protease production under increased air pressures. Methods and Results: Batch cultures of the yeast were conducted in a pressurized bioreactor at 4 and 8 bar of air pressure and the cellular behaviour was compared with cultures at atmospheric pressure. No inhibition of cellular growth was observed by the increase of pressure. Moreover, the improvement of the oxygen transfer rate (OTR) from the gas to the culture medium by pressurization enhanced the extracellular lipase activity from 96·6 U l^?1 at 1 bar to 533·5 U l^?1 at 8 bar. The extracellular protease activity was reduced by the air pressure increase, thereby eliciting further lipase productivity. Cell morphology was slightly affected by pressure, particularly at 8 bar, where cells kept the predominant oval form but decreased in size. Conclusions: OTR improvement by total air pressure rise up to 8 bar in a bioreactor can be applied to the enhancement of lipase production by Y. lipolytica. Significance and Impact of the Study: Hyperbaric bioreactors can be successfully applied for yeast cells cultivation, particularly in high‐density cultures used for enzymes production, preventing oxygen limitation and consequently increasing overall productivity.     

Insecticidal action of Bauhinia monandra leaf lectin (BmoLL) against Anagasta kuehniella (Lepidoptera: Pyralidae), Zabrotes subfasciatus and Callosobruchus maculatus (Coleoptera: Bruchidae)

Bruchid beetle larvae cause major losses in grain legume crops throughout the world. Some bruchid species, such as the cowpea weevil (Callosobruchus maculatus) and the Mexican bean weevil (Zabrotes subfasciatus), are pests that damage stored seeds. The Mediterranean flour moth (Anagasta kuehniella) is of major economic importance as a flour and grain feeder; it is often a severe pest in flour mills. Plant lectins have been implicated as antibiosis factors against insects. Bauhinia monandra leaf lectin (BmoLL) was tested for anti-insect activity against C. maculatus, Z. subfasciatus and A. kuehniella larvae. BmoLL produced ca. 50% mortality to Z. subfaciatus and C. maculatus when incorporated into an artificial diet at a level of 0.5% and 0.3% (w/w), respectively. BmooLL up to 1% did not significantly decrease the survival of A. kuehniella larvae, but produced a decrease of 40% in weight. Affinity chromatography showed that BmoLL bound to midgut proteins of the insect C. maculatus. 33 kDa subunit BmoLL was not digested by midgut preparations of these bruchids. BmoLL-fed C. maculatus larvae increased the digestion of potato starch by 25% compared with the control. The transformation of the genes coding for this lectin could be useful in the development of insect resistance in important agricultural crops.     

Thymidylate synthase polymorphisms are associated to therapeutic outcome of advanced non-small cell lung cancer patients treated with platinum-based chemotherapy

Thymidylate synthase (TYMS) has three polymorphisms that may modulate thymidylate synthase (TS) expression levels: (1) 28 base pairs (bp) variable number tandem repeat (VNTR) (rs34743033); (2) single nucleotide polymorphism (SNP) C>G at the twelfth nucleotide of the second repeat of 3R allele (rs2853542); and (3) 6 bp sequence deletion (1494del6, rs34489327). This study was conducted to evaluate the influence of TYMS polymorphisms on the survival of Portuguese patients with advanced non-small cell lung cancer (NSCLC) undergoing platinum-based chemotherapy. Our results showed no statistically significant differences between VNTR genotypes; although, considering the SNP C>G, homozygotes 3RG presented a better prognostic at 36 months (p = 0.004) and overall survival (p = 0.003) when compared to 2R3RG patients. Patients with “median/high expression genotypes” demonstrated a better survival at 12 months (p = 0.041) when compared to “low expression genotypes”. Furthermore, 6 bp? carriers (p = 0.006) showed a better survival at 12 months when compared to 6 bp+ homozygotes patients. When analyzing TYMS haplotypes, better survival at 12 months was observed for patients carrying haplotypes with the 6 bp? allele (2R6 bp?; p = 0.026 and 3RG6 bp?; p = 0.045). This is the first report that evaluates the three major TYMS polymorphisms in the therapeutic outcome of NSCLC in Portugal. According to our results, the TYMS polymorphisms may be useful tools to predict which advanced NSCLC patients could benefit more from platinum-based chemotherapy regimens.     

Experimental paracoccidioidomycosis of hamster inoculated in the cheek pouch

We compared the granuloma morphology and immune response of hamsters inoculated withParacoccidioides brasiliensis (Pb) into the cheek pouch, which lacks lymphatic drainage, and into the footpad, which is rich in lymphatics. Our objective was to better understand the modulation ofPb granuloma in an immunocompetent animal inoculated in an immunologically privileged site. The humoral immune response (ELISA) and cell mediated immunity (footpad test) became positive on days 7 and 14, respectively in animals inoculated into footpad and on days 35 and 60 in animals inoculated into the pouch. Typical epithelioid granulomas were observed at both sites on day 14. The number of fungi gradually decreased from the beginning of the experiment in footpad lesions, but only after day 35 in pouch granulomas, when cell mediated immunity was detectable. The results indicate that typical epithelioid paracoccidioidomycotic granulomas may develop in the absence of a detectable immune response; however, they are incapable of controlling fungal reproduction. Lack of lymphatic drainage delays the appearance of a detectable immune response, but with time fungi escape from the pouch, elicit an immune response and reach other organs. Our results further indicate the importance of the lymphatics in the pathogenesis of paracoccidioidomycosis.Abbreviations HCP hamster cheek pouch - Pb Paracoccidioides brasiliensis - Pbmycosis Paracoccidioidomycosis     

Correlations between cresty neck scores and post-mortem nape fat measurements in horses,obtained after photographic image analysis

Background

Obesity and emaciation in horses have major detrimental effects on health and morbidity, reproductive failure, work performance or carcass quality. Scoring is a current management tool used to assess and monitor equine body condition due to its simplicity and low cost. However, accurate assessment of obesity remains a challenge, even though a number of approaches have been tested, particularly for research purposes on adiposity. Their merit is usually validated by comparison with standard scoring methods. The overall aim of this study was to establish the correlation between post-mortem nape fat measurements obtained after photographic image analysis and cresty neck score (CNS) in horses. Data were collected from seventeen horses with a hot carcass weight of 165 ± 51 kg. Pre-slaughter CNS measurements were obtained using a six-point scale (from 0 to 5). Image capture was performed post-mortem, in the slaughter line; for each carcass, images of the dorsal and medial views were collected and afterwards transferred to a computer for analysis. After outlining the cresty neck fat, its area, major axis and thickness were determined. Correlation coefficients between nape fat measurements, CNS and carcass fatness were determined.

Results

The horses in the study show similar variation for CNS and hot carcass weight [Coefficient of variation (CV) = 32 and 31 %, respectively], but a high variation for carcass fattening (CV = 41 %). The nape fat area measurement was the parameter exhibiting the greatest variation (CV = 50 %). Correlations established between CNS and the variables tested revealed the existence of moderate to strong correlations among CNS, nape fat measurements, and carcass fatness. The highest correlation coefficients were found between CNS and nape fat thickness (r = 0.882; P < 0.01). The linear regression between CNS and nape fat thickness accounted for 77 % of the recorded variation for nape fat thickness.

Conclusions

The present study showed that there is a strong correlation between horse CNS and post-mortem nape fat measurements or carcass fatness.

     

Enantioselective determination of metoprolol acidic metabolite in plasma and urine using liquid chromatography chiral columns: applications to pharmacokinetics

Enantioselective separations on chiral stationary phases with or without derivatization were developed and compared for the HPLC analysis of (+)-(R)- and (-)-(S)-metoprolol acidic metabolite in human plasma and urine. The enantiomers were analysed in plasma and urine without derivatization on a Chiralcel OD-R column, and in urine after derivatization using methanol in acidic medium on a Chiralcel OD-H column. The quantitation limits were 17 ng of each enantiomer/ml plasma and 0.5 microgram of each enantiomer/ml urine using both methods. The confident limits show that the methods are compatible with pharmacokinetic investigations of the enantioselective metabolism of metoprolol. The methods were employed in a metabolism study of racemic metoprolol administered to a patient phenotyped as an extensive metabolizer of debrisoquine. The enantiomeric ratio (+)-(R)/(-)-(S)-acid metabolite was 1.1 for plasma and 1.2 for urine. Clearances were 0.41 and 0.25 l/h/kg, respectively, for the (+)-(R)- and (-)-(S)-enantiomers. The correlation coefficients between the urine concentrations of the acid metabolite enantiomers obtained by the two methods were >0.99. The two methods demonstrated interchangeable application to pharmacokinetics.     

l-3,4-Dihydroxyphenylalanine (l-DOPA) secreted by oyster (Crassostrea gigas) mantle cells: functional aspects

l-DOPA is present in many molluscan periostraca and is thought to play an important role in the shell protein sclerotization mechanism. We analyzed l-DOPA content in organic membranes produced by the oyster (Crassostrea gigas) mantle as a reaction against shell damage, such as induced by Polydora sp. infestation, an artificial perforation of the shell, and in normal oysters mantle. When oysters are secreting the protective organic membrane against a shell perforation, the area of mantle close to the repairing region had a greater l-DOPA content than the remaining mantle. Mantle border as a whole had the same l-DOPA content as the central mantle. Nevertheless there is a variation pattern in mantle border l-DOPA content, the area near the umbo having a higher content and decreasing towards the frontal region. l-DOPA content in younger oysters was greater than that in older oysters. The results of perforation experiment suggest a good correlation between organic membrane synthesis and l-DOPA mantle border content. Concerning normal oysters, a higher l-DOPA content in mantle border than in central mantle would be expected but we couldn’t detect any differences. From all the results there is some evidence that mantle l-DOPA is related to other functions in addition to participating in the sclerotization mechanism.     

Evolutionary origin of human and primate malarias: evidence from the circumsporozoite protein gene

We have analyzed the conserved regions of the gene coding for the circumsporozoite protein (CSP) in 12 species of Plasmodium, the malaria parasite. The closest evolutionary relative of P. falciparum, the agent of malignant human malaria, is P. reichenowi, a chimpanzee parasite. This is consistent with the hypothesis that P. falciparum is an ancient human parasite, associated with humans since the divergence of the hominids from their closest hominoid relatives. Three other human Plasmodium species are each genetically indistinguishable from species parasitic to nonhuman primates; that is, for the DNA sequences included in our analysis, the differences between species are not greater than the differences between strains of the human species. The human P. malariae is indistinguishable from P. brasilianum, and P. vivax is indistinguishable from P. simium; P. brasilianum and P. simium are parasitic to New World monkeys. The human P. vivax-like is indistinguishable from P. simiovale, a parasite of Old World macaques. We conjecture that P. malariae, P. vivax, and P. vivax-like are evolutionarily recent human parasites, the first two at least acquired only within the last several thousand years, and perhaps within the last few hundred years, after the expansion of human populations in South America following the European colonizations. We estimate the rate of evolution of the conserved regions of the CSP gene as 2.46 x 10(-9) per site per year. The divergence between the P. falciparum and P. reichenowi lineages is accordingly dated 8.9 Myr ago. The divergence between the three lineages leading to the human parasites is very ancient, about 100 Myr old between P. malariae and P. vivax (and P. vivax-like) and about 165 Myr old between P. falciparum and the other two.      

Cost-Effectiveness Analysis of Universal Vaccination of Adults Aged 60 Years with 23-Valent Pneumococcal Polysaccharide Vaccine versus Current Practice in Brazil

Objective

To evaluate the cost-effectiveness of introducing universal vaccination of adults aged 60 years with the 23-valent pneumococcal polysaccharide vaccine (PPV23) into the National Immunization Program (NIP) in Brazil.

Methods

Economic evaluation using a Markov model to compare two strategies: (1) universal vaccination of adults aged 60 years with one dose of PPV23 and 2) current practice (vaccination of institutionalized elderly and elderly with underlying diseases). The perspective was from the health system and society. Temporal horizon was 10 years. Discount rate of 5% was applied to costs and benefits. Clinical syndromes of interest were invasive pneumococcal disease (IPD) including meningitis, sepsis and others and pneumonia. Vaccine efficacy against IPD was obtained from a meta-analysis of randomized control trials and randomized studies, whereas vaccine effectiveness against pneumonia was obtained from cohort studies. Resource utilization and costs were obtained from the Brazilian Health Information Systems. The primary outcome was cost per life year saved (LYS). Univariate and multivariate sensitivity analysis were performed.

Results

The universal vaccination strategy avoided 7,810 hospitalizations and 514 deaths, saving 3,787 years of life and costing a total of USD$31,507,012 and USD$44,548,180, respectively, from the health system and societal perspective. The universal immunization would result in ICERs of USD$1,297 per LYS, from the perspective of the health system, and USD$904 per LYS, from the societal perspective.

Conclusion

The results suggest that universal vaccination of adults aged 60 years with the 23-valent pneumococcal polysaccharide vaccine (PPV23) is a very cost-effective intervention for preventing hospitalization and deaths for IPD and pneumonia is this age group in Brazil.