Identification of passion fruit (Passiflora edulis) chromosomes using BAC-FISH.

Passiflora edulis, the yellow passion fruit, is the main crop from the Passiflora genus, which comprises 525 species with its diversity center in South America. Genetic maps and a BAC (bacterial artificial chromosome) genomic library are available, but the nine chromosome pairs of similar size and morphology (2n = 18) hamper chromosome identification, leading to different proposed karyotypes. Thus, the aim of this study was to establish chromosome-specific markers for the yellow passion fruit using single-copy and repetitive sequences as probes in fluorescent in situ hybridizations (FISH) to allow chromosome identification and future integration with whole genome data. Thirty-six BAC clones harboring genes and three retrotransposons (Ty1-copy, Ty3-gypsy, and LINE) were selected. Twelve BACs exhibited a dispersed pattern similar to that revealed by retroelements, and one exhibited subtelomeric distribution. Twelve clones showed unique signals in terminal or subterminal regions of the chromosomes, allowing their genes to be anchored to six chromosome pairs that can be identified with single-copy markers. The markers developed herein will provide an important tool for genomic and evolutionary studies in the Passiflora genus.

Chronic cutaneous lupus erythematosus presenting as atypical acneiform and comedonal plaque: case report and literature review.

Introduction Chronic cutaneous lupus erythematosus (CCLE) usually presents as characteristic erythematous patches and infiltrated coin-shaped plaques. However, there are some atypical clinical variants that may mimic other dermatological conditions. Haroon et al. reported in 1972 an unusual presentation of CCLE with hypertrophic follicular scars seen in acne vulgaris. Acneiform presentation is one of the most rarely reported and one of the most confusing, as it resembles a very common inflammatory skin disease. A brief review of the literature using PubMed found only nine other reports. Case report A 32-year-old woman presented with two-year pruritic infiltrated acneiform and comedonal eruption on the right chin treated as acne with isotretinoin without improvement. On examination the patient presented with erythematous-infiltrated plaque, papules, open comedones, pitting scars and hypopigmented atrophic scars on the right chin area and scalp hair loss. An incisional skin biopsy on the chin and scalp lesions was performed and the anatomopathological and immunofluorescence exam showed findings that are consistent with CCLE. Additional tests ruled out systemic involvement. The patient was treated with prednisone and chloroquine diphosphate with great improvement. After four years the lesion is stable, with some scarring. Discussion In a literature review we found nine other cases of acneiform presentation of lupus erythematosus: Three cases were systemic lupus erythematosus (SLE) and seven others were diagnosed as CCLE (including our patient). All three patients who had SLE tested positive for antinuclear antibodies (ANA), and only one patient with CCLE, had a low titer of positive ANA (1:80). Ages varied from 24 to 60 years old, with a median of 32 years old, the same as our patient's age and consistent with the literature. Seven were females and three were males, with a ratio of 2.3:1. Most cases, such as our patient, showed acneiform lesions mainly on the face, a common site of typical CCLE. The present case and literature review illustrates the need to expand the differential diagnosis of atypical acneiform and comedonal lesions. CCLE should be considered especially in a localized lesion, which can be itchy and does not improve with conventional treatment for acne vulgaris.

Activity of Antarctic fungi extracts against phytopathogenic bacteria.

This study aims to obtain secondary metabolites extracts from filamentous fungi isolated from soil and marine sediments from Antarctic ecosystems and to assess its potential antibacterial activity on Xanthomonas euvesicatoria and Xanthomonas axonopodis pv. passiflorae (phytopathogenic bacteria causing diseases in pepper and tomato and passionfruit, respectively). Among the 66 crude intracellular and extracellular extracts obtained from fungi recovered from soil and 79 obtained from marine sediment samples, 25 showed the ability to prevent the growth of X. euvesicatoria in vitro and 28 showed the ability to prevent the growth of X. axonopodis pv. passiflorae in vitro. Intracellular and extracellular extracts from soil fungi inhibited around 97% of X. euvesicatoria and 98% of X. axonopodis pv. passiflorae at 2·1 mg ml-1 . The average inhibition rates against X. euvesicatoria and X. axonopodis pv. passiflorae for intracellular and extracellular extracts from marine sediments fungi were around 96 and 97%, respectively, at 3·0 mg ml-1 . Extracts containing secondary metabolites with antimicrobial activity against X. euvesicatoria and X. axonopodis pv. passiflorae were obtained, containing possible substitutes for the products currently used to control these phytopathogens. SIGNIFICANCE AND IMPACT OF THE STUDY: Micro-organisms from extreme ecosystems, such as the Antarctic ecosystem, need to survive in harsh conditions with low temperatures, low nutrients and high UV radiation. Micro-organisms adapt to these conditions evolving diverse biochemical and physiological adaptations essential for survival. All this makes these micro-organisms a rich source of novel natural products based on unique chemical scaffolds. Discovering novel bioactive compounds is essential because of the rise in antibiotic-resistant micro-organisms and the emergence of new infections. Fungi from Antarctic environments have been proven to produce bioactive secondary metabolites against various micro-organisms, but few studies have shown activity against Xanthomonas phytopathogens.

Prevalence of Leptospira antibodies in wild boars (Sus scrofa) from Northern Portugal: risk factor analysis.

Leptospirosis is a zoonosis of worldwide distribution, caused by infection with pathogenic spirochaetes of the genus Leptospira. The wild boar (Sus scrofa), an important hunting species in Europe, seems to play a significant role in the epidemiological cycle of leptospirosis. A total of 101 serum samples from wild boar hunted in Northern Portugal were analysed for leptospiral antibodies detection by microscopic agglutination test. Sera were collected during hunting seasons (2011-2013) and tested with 17 different pathogenic serovars of Leptospira. Antibodies against nine serovars were detected in 66 (65·4%) of these sera. Serovars Tarassovi and Altodouro exhibited the highest seroreactivity rates (23·8% and 16·8%, respectively), followed by Autumnalis (7·9%) and Bratislava (6·9%). Age and district of origin were found to be risk factors for the presence of leptospiral antibodies in contrast to gender. From a One Health perspective, this study revealed that wild boar should be considered as a potential source of leptospirosis dissemination for humans and animal species (domestic and wild) in shared environments, particularly in the Trás-os-Montes region.

Genetic Variability of Beauveria bassiana and a DNA Marker for Environmental Monitoring of a Highly Virulent Isolate Against Cosmopolites sordidus.

The banana weevil Cosmopolites sordidus (Germar) is one of a number of pests that attack banana crops. The use of the entomopathogenic fungus Beauveria bassiana as a biological control agent for this pest may contribute towards reducing the application of chemical insecticides on banana crops. In this study, the genetic variability of a collection of Brazilian isolates of B. bassiana was evaluated. Samples were obtained from various geographic regions of Brazil, and from different hosts of the Curculionidae family. Based on the DNA fingerprints generated by RAPD and AFLP, we found that 92 and 88 % of the loci were polymorphic, respectively. The B. bassiana isolates were attributed to two genotypic clusters based on the RAPD data, and to three genotypic clusters, when analyzed with AFLP. The nucleotide sequences of nuclear ribosomal DNA intergenic spacers confirmed that all isolates are in fact B. bassiana. Analysis of molecular variance showed that variability among the isolates was not correlated with geographic origin or hosts. A RAPD-specific marker for isolate CG 1024, which is highly virulent to C. sordidus, was cloned and sequenced. Based on the sequences obtained, specific PCR primers BbasCG1024F (5'-TGC GGC TGA GGA GGA CT-3') and BbasCG1024R (5'-TGC GGC TGA GTG TAG AAC-3') were designed for detecting and monitoring this isolate in the field.

Genetic diversity and a PCR-based method for Xanthomonas axonopodis detection in passion fruit.

Xanthomonas axonopodis pv. passiflorae causes bacterial spot in passion fruit. It attacks the purple and yellow passion fruit as well as the sweet passion fruit. The diversity of 87 isolates of pv. passiflorae collected from across 22 fruit orchards in Brazil was evaluated using molecular profiles and statistical procedures, including an unweighted pair-group method with arithmetical averages-based dendrogram, analysis of molecular variance (AMOVA), and an assigning test that provides information on genetic structure at the population level. Isolates from another eight pathovars were included in the molecular analyses and all were shown to have a distinct repetitive sequence-based polymerase chain reaction profile. Amplified fragment length polymorphism technique revealed considerable diversity among isolates of pv. passiflorae, and AMOVA showed that most of the variance (49.4%) was due to differences between localities. Cluster analysis revealed that most genotypic clusters were homogeneous and that variance was associated primarily with geographic origin. The disease adversely affects fruit production and may kill infected plants. A method for rapid diagnosis of the pathogen, even before the disease symptoms become evident, has value for producers. Here, a set of primers (Xapas) was designed by exploiting a single-nucleotide polymorphism between the sequences of the intergenic 16S-23S rRNA spacer region of the pathovars. Xapas was shown to effectively detect all pv. passiflorae isolates and is recommended for disease diagnosis in passion fruit orchards.

Synergistic effect of serine protease inhibitors and a bothropic antivenom in reducing local hemorrhage and coagulopathy caused by Bothrops jararaca venom.

Snakebite accidents are a public health problem that affects the whole world, causing thousands of deaths and amputations each year. In Brazil, snakebite envenomations are caused mostly by snakes from the Bothrops genus. The local symptoms are characterized by pain, swelling, ecchymosis, and hemorrhages. Systemic disturbances can lead to necrosis and amputations. The present treatment consists of intravenous administration of bothropic antivenom, which is capable of reversing most of the systemic symptoms, while presenting limitations to treat the local effects, such as hemorrhage and to neutralize the snake venom serine protease (SVSP). In this context, we aimed to evaluate the activity of selective serine protease inhibitors (pepC and pepB) in combination with the bothropic antivenom in vivo. Further, we assessed their possible synergistic effect in the treatment of coagulopathy and hemorrhage induced by Bothrops jararaca venom. For this, we evaluated the in vivo activity in mouse models of local hemorrhage and a series of in vitro hemostasis assays. Our results showed that pepC and pepB, when combinated with the antivenom, increase its protective activity in vivo and decrease the hemostatic disturbances in vitro with high selectivity, possibly by inhibiting botropic proteases. These data suggest that the addition of serine protease inhibitor to the antivenom can improve its overall potential.

Resistance to Passion fruit woodiness virus in Transgenic Passionflower Expressing the Virus Coat Protein Gene.

We report the use of the coat protein (CP) gene from Passion fruit woodiness virus (PWV) to produce resistant transgenic plants of yellow passion fruit. A full-length CP gene from a severe PWV isolate from the state of São Paulo, Brazil (PWV-SP) was cloned into pCAMBIA 2300 binary vector, which was further introduced into Agrobacterium tumefaciens strain EHA 105. Leaf disks were used as explants for transformation assays, e.g., 2,700 and 2,730 disks excised from plants from the Brazilian cultivars IAC-275 and IAC-277, respectively. In vitro selection was performed in kanamycin. After transferring to the elongation medium, 119 and 109 plantlets of IAC-275 and IAC-277, respectively, were recovered. Integration of the PWV CP gene was confirmed in seven of eight plants evaluated by Southern blot analysis, showing different numbers of insertional events for the CP gene. Three transgenic plants (T3, T4, and T7) expressed the expected transcript, but the 32 kDa PWV CP was detected by Western blot in only two plants (T3 and T4). The results of three successive mechanical inoculations against the transgenic plants using three PWV isolates showed that the primary transformant T2 of IAC-277 was immune to all isolates.

Identification by genomic typing of non-DR3 HLA class II genes associated with myasthenia gravis.

HLA association with myasthenia gravis (MG) has been studied in a series of 114 patients using class I and class II genotyping after PCR amplification. Positive association was found with DR3, particularly in women (RR = 2.6) and in early MG onset (RR = 3.4). DRB1, DRB3, DQB1, DQA1 and B (B8 and B18) genotyping revealed that the association was predominantly with the B8 DRB1*03 DRB3*0101 DQB1*0201 DQA1*0501 ancestral haplotype. This haplotype frequency was also increased in patients with thymic hyperplasia (RR = 3.5) and was greatly reduced in patients with thymoma (RR = 0.35). Sixteen out of 48 patients carrying this 8.1 ancestral haplotype showed absence of B8 (n = 4) or of DR3 (n = 12). HLA class II genotyping further revealed the existence of two other significant associations. MG was positively associated with the DQB1*0604 allele (RR = 3.4), particularly in patients with thymoma (RR = 5.7). Furthermore, the disease was negatively associated with DR1 in females (RR = 0.32). These data suggest that MG is placed under the control of at least three distinct genes: (1) a class II predisposing gene in the 8.1 ancestral haplotype; (2) a thymoma-associated class II allele on the DQB1*0604 haplotype; and (3) a protective allele DR1.

Polymorphism of antigen processing (TAP, LMP) and HLA class II genes in celiac disease.

Susceptibility to CD is strongly associated with particular HLA class II molecules. However, additive genetic factors are likely to be required for the development of the disease. The polymorphic TAP and LMP genes, located within the HLA class II region, are involved in the antigen presentation pathway and thus represent candidate susceptibility genes. HLA class II DRB1, DRB3, DQA1, DQB1, and DPB1 as well as TAP1, TAP2, and LMP2 polymorphism was studied in 80 Caucasian CD patients and 213 normal controls by DNA oligotyping. The DQB1*0201 allele was found in 96.3% of CD patients and provided the highest risk (RR = 50), whereas only 89% of CD patients carried the DQ alpha 501/beta 201 heterodimer (RR = 30). The participation of the DRB3 and DPB1 locus was ruled out as it was attributed to a linkage disequilibrium on the DR3 haplotype. TAP1 and LMP2 allelic distribution was not significantly different among CD patients and controls. The TAP2-C allele was completely absent from the CD population, while it was found in 22.5% of controls. Although linkage disequilibrium between TAP2 and class II loci clearly exists in some haplotypes, TAP could act as additional susceptibility genes.

Higher incidence of malignant neoplasms after heart transplantation for treatment of chronic Chagas' heart disease.

BACKGROUND: Heart transplantation is a new therapeutic procedure to treat heart failure resulting from Chagas' disease. Experimental studies have demonstrated neoplastic effects of benznidazole, which is used for treatment of Trypanosoma cruzi infection. We compared the incidence and characteristics of neoplasia after heart transplantation for treatment of chronic Chagas' disease with those of other diseases. METHODS: Sixteen patients with Chagas' disease and 75 patients with other diseases underwent heart transplantation. Benznidazole was administered to 14 patients with Chagas's disease either for prophylaxis (4 patients) or for treatment of Chagas' disease reactivation (10 patients). RESULTS: The survival rate of patients in the nonchagasic group was 90% at 1 year and 82.4% at 2 years, and the survival rate in the chagasic group was 63% at 1 year and 57% at 2 years. Six of 16 patients (37.5%) with Chagas' disease had malignant tumors after a mean follow-up time of 25.3+/-2.1 months in contrast to 2 of 75 patients (2.7%) in the nonchagasic group after 34.6+/-3.6 months of follow-up. In the chagasic group, lymphoproliferative disorder was diagnosed in three patients, Kaposi's sarcoma in two, and squamous cell carcinoma in one patient. Reactivation of T. cruzi infection was diagnosed in all patients who had lymphoproliferative disorder. One patient without Chagas' disease had lymphoproliferative disorder in the lung, and another had malignant schwannoma affecting the skin. CONCLUSIONS: We found a higher incidence of malignant neoplasia after heart transplantation for treatment of chronic Chagas' disease. It is likely that the neoplasia is the result of chronic infection with an immunomodulator protozoan, immunosuppression, reactivation of the T. cruzi infection, or the toxicity of therapeutic intervention with benznidazole.

Blood pressure and the risk of complex arrhythmia in renal insufficiency, hemodialysis, and renal transplant patients.

Complex arrhythmia is frequent in hemodialysis patients but it is not clear if this is a consequence of dialysis or uremia or is secondary to the hemodynamic and cardiovascular alterations often associated with chronic renal failure. The incidence of complex ventricular arrhythmia (frequent multiform premature beats, couplets, and runs) in 31 subjects who had their uremic status recently corrected by renal transplant (Group 1) and in 23 predialysis (Group 2) and 73 hemodialysis (Group 3) chronic renal failure patients were studied with 24-h Holter monitoring. Patients were not receiving antiarrhythmic drugs or digitalis and significant coronary artery disease was excluded by clinical and noninvasive methods. Complex arrhythmia was two times more frequent in dialysis patients but the difference did not reach statistical significance (Group 1: 16%; Group 2: 17%; Group 3: 34%; chi2 4.9, P = .086). The stepwise model of logistic regression analysis identified systolic blood pressure (odds ratio 1.015, 95% confidence interval [CI] 1.001-1.027, P = .03) and left ventricular systolic dysfunction (odds ratio 7.04, 95% CI 1.3-36.7, P = .02) as the only factors that independently influenced the probability of complex arrhythmia. Age, gender, race, diabetes, smoking status, body mass index, diastolic blood pressure, serum creatinine, hematocrit, left ventricular mass index, and use of diuretics, beta-blockers, angiotensin converting enzyme (ACE) inhibitors, sympatolytics, and calcium channel blockers did not influence the occurrence of complex arrhythmia. The data indicate that blood pressure and myocardial dysfunction are more important determinants of complex arrhythmia than dialysis or uremia in chronic renal disease patients.

Sex differences in the play fighting activity of golden hamster infants.

Play fighting of homosexual and heterosexual dyads in golden hamsters was studied. The behaviors of male-male (MM), female-female (FF), and female-male (FM) dyads were recorded from the 25th to the 31st day of age. Play fighting activity of heterosexual dyads was depressed since MF and FM dyads exhibited a significantly high physical contact time and engaged in less play fighting, with a significantly lower time spent on such activity. Alternation ratio in the pinning posture was very high and subjects did not differ in the mean number of active (top) and passive pinning (under) in each dyad, showing an inexistence of sex dominance. In most cases, differences in the performances of MM and FF dyads were not significant. We have concluded that play fighting in golden hamsters may be predominantly modulated by environmental stimuli (sex of the partner) rather than by endogenous stimuli (androgen level).

Effects of blindness on play fighting in golden hamster infants.

The effects of bilateral optic enucleation on play fighting in golden hamster infants were studied. The behaviors of blind-blind (BB), blind-intact (BI) and intact-intact (II) dyads were recorded from the 30th to the 36th day of age. It has been noted that the differences in physical contact, play fighting and grooming time, as well as in the percentage of time spent in play fighting, the amount of locomotion and other activities of BB, BI and II were small and statistically insignificant. The mean number of active and passive pinning in all groups was equivalent and the alternation of active and passive pinning, or vice versa, was around 50%, showing that posture during play fighting is not an indicator of dominance. It has been concluded that vision is not important for either the initiation or the modulation of play fighting in golden hamsters.

Play fighting in juvenile golden hamsters (Mesocricetus auratus): Effects of litter size and analysis of social interaction among males.

Previous studies have shown that different social experiences, such as the interaction involving partners of different ages or among individuals of different sexes, significantly alter the play rate. In the present study, two experiments are described, the hamster being used as subject, to investigate the effects of litter size and to analyse the play fighting behavior among males. It was found that there was no significant difference in the time spent in play fighting by an animal when it was placed in groups of 2, 4 or 6 individuals (Exp. 1). Moreover, the majority of animals did not show preferences during play fighting when they were placed in groups formed by four males (Exp. 2). Through these data it has been noted that the hamster distributes the total time spent in play fighting among the available partners. This fact can be explained by the hamster's social organization, because it is a solitary animal, which does not develop a clear and durable interaction with its playmates. Also, through other results, a certain stability was found in most of the behavioral categories registered such that they were not significantly affected by the manipulated variables.

Cryopreservation of Arachis species by vitrification of in vitro-grown shoot apices and genetic stability of recovered plants.

A storage protocol at cryogenic temperature was established for shoot apices from in vitro plants of the cultivated groundnut (Arachis hypogaea) and wild Arachis species (A. retusa and A. burchellii) using a basic vitrification protocol with direct immersion in liquid nitrogen (LN). The effect of pre-treatments of donor-plants with ABA as well as of different supplements in the post-thaw culture medium was studied. After rapid warming at 40 C, the explants were cultured on MS medium devoid of growth regulators (MS0) or MS supplemented with 4.4(M benzylaminopurine (BAP) and 0.5(M naphthalene acetic acid (NAA) plus 5(M silver nitrate (AgNO3), 0.25% polyvinylpyrrolidone (PVP) or 0.2% activated charcoal. Non-frozen explants from the three species formed one shoot through meristematic amplification when cultured on MS0 medium. These explants also developed callus on MS supplemented with growth regulators (4.4(M BAP and 0.5(M NAA) alone or plus PVP or AgNO3. Callus formation was suppressed in the presence of activated charcoal. Post-thaw regeneration ocurred only through indirect organogenesis on media containing AgNO3 or PVP. Preculturing on medium supplemented with abscisic acid (ABA) improved regrowth rate in these media. Recovery failed to occur in the presence of activated charcoal. The genetic stability of shoots of A. burchellii originated from shoot apices was analyzed through Random Amplified Polymorphic DNA (RAPD) markers.

Clinical aspects of pulpal-periodontal lesions.

It is well known that the main purpose of dentistry is to keep teeth in the oral cavity so that they may fulfill their various roles. This case report demonstrates a method to treat a situation involving a pulpal-periodontal lesion, by using a simple and efficient combined therapy. The clinician can thereby determine the cause of the lesion (pulpal and/or periodontal) and subsequently establish the correct treatment, thus improving the prognosis of the case.

[Thrombosis in normal coronary arteries causing acute myocardial infarction in an alcoholic patient]. / Trombose em artérias coronárias normais causando infarto agudo do miocárdio em paciente alcoólatra.

A case of a young patient, heavy drinker, who suffered an acute myocardial infarction (AMI), and whose cinecoronariography, early post unsuccessful intravenous thrombolysis, showed the presence of thrombi in multiple arteries is reported. A second cinecoronariography performed on the 7th day post AMI revealed normal coronary arteries. There were no plasmatic lipid or coagulation abnormalities, nor other risk factors for coronary artery disease. We speculate about the role of the heavy alcohol ingestion in this particular case.

[Aneurysms of the left and non coronary sinus of Valsalva associated to mitral and aortic rheumatic valves and obstructive coronary disease]. / Aneurismas dos seios de Valsalva esquerdo e não-coronário associados à valvopatia reumática mitro-aórtica e à coronariopatia obstrutiva.

A 30 years old, male, patient with rare multiple aneurysms of sinus of Valsalva associated to mitral and aortic regurgitation and total occlusion of circumflex artery had the clinical diagnosis made accidentally. The multiple aneurysms were detected by transesophageal echodopplercardiography and hemodynamic study. The diagnosis were confirmed at surgery and the patient underwent correction of the aneurysms through the use of pericardial patches and mitral and aortic valve replacement.