Single nucleotide polymorphisms in the interferon gamma gene are associated with distinct types of retinochoroidal scar lesions presumably caused by Toxoplasma gondii infection

The association of single nucleotide polymorphisms (SNPs) in the interferon (IFN)-γ gene ( IFNG ) with different types of retinal scar lesions presumably caused by toxoplasmosis were investigated in a cross-sectional population-based genetic study. Ten SNPs were investigated and after Bonferroni correction, only the associations between SNPs rs2069718 and rs3181035 with retinal/retinochoroidal scar lesions type A (most severe scar lesions) and C (least severe scar lesions), respectively, remained significant. The associations of two different IFNG SNPs with two different types of retinal lesions attributable to toxoplasmosis support the hypothesis that different inflammatory mechanisms underlie the development of these lesions. The in vitro analysis of IFN-γ secretion by peripheral blood mononuclear cells stimulated with Toxoplasma gondii antigens was also investigated. The association between SNP rs2069718 and type A scar lesions revealed that differential IFN-γ levels are correlated with distinct genotypes. However, no correlation was observed with IFN-γ secretion levels and the SNP rs3181035 , which was significantly associated with type C scar lesions. Our findings strongly suggest that immunogenetic studies of individuals with congenital or postnatally acquired infection are needed to better understand the role of IFN-γ and its polymorphisms in the pathogenesis of ocular toxoplasmosis.

Utilização de marcador molecular SCAR na identificação de Fusarium subglutinans, agente causal da malformação da mangueira

O gênero Fusarium é responsável por doenças em diversas plantas economicamente importantes. Entre estas doenças, destaca-se a malformação da mangueira, causada pelo fungo Fusarium subglutinans. O objetivo deste trabalho foi desenvolver oligonucleotídeos iniciadores para reação em cadeia da polimerase (PCR), específicos para o fungo F. subglutinans da mangueira. A amplificação de DNA de oito Fusarium spp. de diferentes hospedeiros, usando o oligonucleotídeo randômico UBC-41 (TTAACCGGGG), produziu um fragmento de aproximadamente 1.300 pb somente para o fungo da mangueira. Tendo em vista que padrões de bandeamento por RAPD não são considerados confiáveis devido à baixa reprodutibilidade dos resultados, o fragmento diferencial foi eluído do gel de agarose, purificado, clonado e seqüenciado. As seqüências nucleotídicas foram utilizadas para identificar e sintetizar quatro pares de oligonucleotídeos específicos, denominados Fs 5, Fs 13, Fs 14 e Fs 15. DNAs de Fusarium spp. de outros hospedeiros (alho, amendoim, cana-de-açúcar, ciclâmen, ervilha, melão e trigo), da planta de mangueira cv. Tommy Atkins sadia e de outros cinco isolados de F. subglutinans de mangueira sintomática, foram submetidos à amplificação com os pares de oligonucleotídeos. Fragmentos amplificados foram visualizados somente para F. subglutinans de mangueira, demonstrando assim a especificidade dos oligonucleotídeos SCAR desenhados.

Development of a scar marker for Pierce's Disease strains of Xylella fastidiosa

The objective of this research was to develop a primer for a polymerase chain reaction specific for Xylella fastidiosa strains that cause Pierce's Disease (PD) in grapes (Vitis vinifera). The DNA amplification of 23 different strains of X. fastidiosa, using a set of primers REP1-R (5'-IIIICGICGIATCCIGGC-3') and REP 2 (5'-ICGICTTATCIGGCCTAC-3') using the following program: 94 ºC/2 min; 35 X (94 ºC/1 min, 45 ºC/1 min and 72 ºC/1 min and 30 s) 72 ºC/5 min, produced a fragment of 630 bp that differentiated the strains that cause disease in grapes from the other strains. However, REP banding patterns could not be considered reliable for detection because the REP1-R and REP 2 primers correspond to repetitive sequences, which are found throughout the bacterial genome. The amplified product of 630 bp was eluted from the agarose gel, purified and sequenced. The nucleotide sequence information was used to identify and synthesize an specific oligonucleotide for X. fastidiosa strains that cause Pierce's Disease denominated Xf-1 (5'-CGGGGGTGTAGGAGGGGTTGT-3') which was used jointly with the REP-2 primer at the following conditions: 94 ºC/2 min; 35 X (94 ºC/1 min, 62 ºC/1 min; 72 ºC/1 min and 30 s) 72 ºC/10 min. The DNAs isolated from strains of X. fastidiosa from other hosts [almond (Prumus amygdalus), citrus (Citrus spp.), coffee (Coffea arabica), elm (Ulmus americana), mulberry (Morus rubra), oak (Quercus rubra), periwinkle wilt (Catharantus roseus), plums (Prunus salicina) and ragweed (Ambrosia artemisiifolia)] and also from other Gram negative and positive bacteria were submitted to amplification with a pair of primers Xf-1/REP 2 to verify its specificity. A fragment, about 350 bp, was amplified only when the DNA from strains of X. fastidiosa isolated from grapes was employed.

Normal skin and hypertrophic scar fibroblasts differentially regulate collagen and fibronectin expression as well as mitochondrial membrane potential in response to basic fibroblast growth factor

Basic fibroblast growth factor (bFGF) regulates skin wound healing; however, the underlying mechanism remains to be defined. In the present study, we determined the effects of bFGF on the regulation of cell growth as well as collagen and fibronectin expression in fibroblasts from normal human skin and from hypertrophic scars. We then explored the involvement of mitochondria in mediating bFGF-inducedeffects on the fibroblasts. We isolated and cultivated normal and hypertrophic scar fibroblasts from tissue biopsies of patients who underwent plastic surgery for repairing hypertrophic scars. The fibroblasts were then treated with different concentrations of bFGF (ranging from 0.1 to 1000 ng/mL). The growth of hypertrophic scar fibroblasts became slower with selective inhibition of type I collagen production after exposure to bFGF. However, type III collagen expression was affected in both normal and hypertrophic scar fibroblasts. Moreover, fibronectin expression in the normal fibroblasts was up-regulated after bFGF treatment. bFGF (1000 ng/mL) also induced mitochondrial depolarization in hypertrophic scar fibroblasts (P < 0.01). The cellular ATP level decreased in hypertrophic scar fibroblasts (P < 0.05), while it increased in the normal fibroblasts following treatment with bFGF (P < 0.01). These data suggest that bFGF has differential effects and mechanisms on fibroblasts of the normal skin and hypertrophic scars, indicating that bFGF may play a role in the early phase of skin wound healing and post-burn scar formation.

Sequence-characterized amplified region (SCAR) technique used for variety discrimination in vinca (Catharanthus roseus (L.) G.Don)

Sequence-Characterized Amplified Region (SCAR) appears as a useful technique for genetic purity testing and variety discrimination, applicable to species in which some other techniques have failed. In particular, this technique is very attractive with species in which RAPD results were not consistent. The RAPD polymorphic bands were cloned, sequenced and from the sequence information, primers pairs for normal PCR were developed. Since the probability of obtaining successful SCAR primers from RAPD polymorphic bands was about 50%, a larger number of RAPD polymorphic bands are needed to develop sufficient SCAR primers for varietal discrimination in vinca. In addition, the efficiency of the SCAR technique is strongly affected by the quality of DNA extracted from seeds. The SCAR banding patterns obtained from vinca seed were consistent and repeatable making the results reliable for genetic purity testing and variety discrimination. The SCAR technique is simple, fast, relatively inexpensive and allows the use of DNA extracted from dry seeds, which is very important in a seed-quality evaluating program

Tegumentary leishmaniasis in Northern Argentina: distribution of infection and disease, in three municipalities of Salta, 1990-1992

This work describes the epidemiological pattern of tegumentary leishmaniasis in an area north of Salta, Argentina. The prevalence and incidence were estimated by means of a cross-sectional study and two follow-up studies during two consecutive years. The Montenegro Skin Test (MST) was administered to 7336 subjects at baseline. The prevalence and incidence between 1990 and 1992 of infection (MST reactive) was 38‰ persons and 4.5‰ persons/year respectively. The prevalence and incidence of tegumentary leishmaniasis (presence of clinical signs) was 1.8‰ and 0.8‰ persons/year, respectively. A physical examination performed on 264 patients with MST reactive during three years revealed that 130 cases (49.2%) had some evident sign of infection (scar and/or lesion), with a clinical presentation compatible with leishmaniasis. Our study demonstrated that after the epidemic outbreak of 1985 the transmission in the study area returned to endemic levels in 1992, and also demonstrated the presence of the asymptomatic infection in the area.

Epidemiological, clinical, and operational aspects of leprosy patients assisted at a referral service in the state of Maranhão, Brazil

INTRODUCTION: Leprosy is an infectious disease caused by Mycobacterium leprae. The aim of this study was to describe the epidemiological, clinical, and operational aspects of leprosy carriers. METHODS: A cross-sectional study leprosy patients assisted in São Luis, MA, was performed. RESULTS: Of the 85 cases analyzed, 51.7% were male participants, and 60% were brown. Concerning the age, 54.8% of women were between 35 and 49 years, and 57.6% of men were between 20 and 34 years. Lepromatous leprosy was found in 42.3% of cases, and the multibacillary form was found in 72.9%. The skin smear was positive in 42.3%. The occurrence of reaction was found in 43.5% of cases, and 83.5% had no Bacillus Calmette-Guérin scar. Leprosy in the family was reported by 44.7% of the patients. Most of the individuals (96.4%) lived in houses made of brick with more than three rooms (72.6%) and two persons per room (65.1%). Concerning the level of education, 41.4% of women and 34.1% of men had more than one to three years of education. The most evaluated age group in the beginning of the treatment was that of 35 to 49 years with a Grade 0 incapability (64.5%), and that in the end was the age group of 20 to 34 (29.9%) with Grade 0, 30.7% Grade 1, and 11.5% Grade 2. CONCLUSIONS: The frequency of multibacillary forms found in this study and the cases in family members point out delayed diagnoses. Thus, early diagnosis and appropriate treatment are important in decreasing the outcome of disabilities.

Schistosoma mansoni granuloma in late evolutive phase, in a case of tumoral form in man

INTRODUCTION: Authors describe human schistosomal granuloma in late chronic phase, from the morphological and evolutionary viewpoints. METHODS: The study was based on a histological analysis of two fragments obtained from a surgical biopsy of peritoneum and large intestine of a 42-year-old patient, with a pseudotumoral form mimicking a peritoneal carcinomatosis associated to the schistosomiasis hepatointestinal form. RESULTS: Two hundred and three granulomas were identified in the pseudotumor and 27 in the intestinal biopsy, with similar morphological features, most in the late chronic phase, in fibrotic healing. A new structural classification was suggested for granulomas: zone 1 (internal), 2 (intermediate) and 3 (external). CONCLUSIONS: Regarding granuloma as a whole, we may conclude that fibrosis is likely to be controlled by different and independent mechanisms in the three zones of the granuloma. Lamellar fibrosis in zone 3 seems to be controlled by matrix mesenchymal cells (fibroblasts and myoepithelial cells) and by inflammatory exudate cells (lymphocytes, plasmocytes, neutrophils, eosinophils). Annular fibrosis in zone 2, comprising a dense fibrous connective tissue, with few cells in the advanced phase, would be controlled by epithelioid cells involving zone 1 in recent granulomas. In zone 1, replacing periovular necrosis, an initialy loose and tracery connective neoformation, housing stellate cells or with fusiform nuclei, a dense paucicellular nodular connctive tissue emerges, probably induced by fibroblasts. In several granulomas, one of the zones is missing and granuloma is represented by two of them: Z3 and Z2, Z3 and Z1 or Z2 and Z1 and, ultimately, by a scar.

Active and latent tuberculosis in prisoners in the Central-West Region of Brazil

Introduction Jailed populations exhibit high rates of tuberculosis (TB) infection and active disease. Methods A cross-sectional study was performed to estimate the prevalence of latent and active TB and to identify factors associated with latent infection in inmates. Results The prevalence of latent TB was 49%, and the prevalence of active TB was 0.4%. The presence of a Bacille Calmette-Guérin (BCG) scar (prevalence ratio (PR)=1.65; 95% confidence interval (CI): 1.09-2.50; p=0.0162) and the World Health Organization (WHO) score for active TB in prisons (PR=1.07; 95% CI: 1.01-1.14; p=0.0181) were correlated with infection. Conclusions The identification of associated factors and the prevalence of latent and active TB allows the development of plans to control this disease in jails.

Effects of circular myotomy on the healing of esophageal suture anastomosis: an experimental study

For esophageal reconstruction in newborns with esophageal atresia, esophageal reunion with an end-to-end anastomosis is the ideal procedure, although it may result in leaks and strictures due to tension on the suture line, mainly in cases with a wide gap between the ends. Circular myotomy (Livaditis' procedure) is the best method to elongate the proximal esophageal pouch and reduce anastomotic tension. This experimental investigation in dogs was undertaken to attempt to verify that circular myotomy decreases the anastomotic leak rate in newborns with wide gap esophageal atresia, and to analyze whether the technique promotes morphologic changes in the anastomotic scar. A pilot study demonstrated that it is necessary to resect more than 8 cm (40% of the total esophageal length) in order to obtain high leak rates. In the experimental project, such resection was performed in dogs divided into two groups (control group, anastomosis only, and experimental group, anastomosis plus circular myotomy in the proximal esophageal segment). The animals were killed in the 14th postoperative day, submitted to autopsy, and were evaluated as to the presence of leaks and strictures, as well as to the features (macroscopic and microscopic aspects) of the anastomosis. Leak rates were the same in both groups. Morphometric analysis revealed that in animals in the experimental group, the anastomotic scar was thinner than the control animals, and the isolated muscular manchette distal to the site of myotomy was replaced by fibrous tissue. Correspondingly, a decreased number of newly formed small vessels were noted in the experimental animals, compared to control animals. We concluded that circular myotomy does not decrease the incidence of anastomotic leaks, and it also promotes deleterious changes in anastomotic healing.

Prognostic Value of Pulmonary Vascular Resistance by Magnetic Resonance in Systolic Heart Failure

Abstract Background: Pulmonary hypertension is associated with poor prognosis in heart failure. However, non-invasive diagnosis is still challenging in clinical practice. Objective: We sought to assess the prognostic utility of non-invasive estimation of pulmonary vascular resistances (PVR) by cardiovascular magnetic resonance to predict adverse cardiovascular outcomes in heart failure with reduced ejection fraction (HFrEF). Methods: Prospective registry of patients with left ventricular ejection fraction (LVEF) < 40% and recently admitted for decompensated heart failure during three years. PVRwere calculated based on right ventricular ejection fraction and average velocity of the pulmonary artery estimated during cardiac magnetic resonance. Readmission for heart failure and all-cause mortality were considered as adverse events at follow-up. Results: 105 patients (average LVEF 26.0 ±7.7%, ischemic etiology 43%) were included. Patients with adverse events at long-term follow-up had higher values of PVR (6.93 ± 1.9 vs. 4.6 ± 1.7estimated Wood Units (eWu), p < 0.001). In multivariate Cox regression analysis, PVR ≥ 5 eWu(cutoff value according to ROC curve) was independently associated with increased risk of adverse events at 9 months follow-up (HR2.98; 95% CI 1.12-7.88; p < 0.03). Conclusions: In patients with HFrEF, the presence of PVR ≥ 5.0 Wu is associated with significantly worse clinical outcome at follow-up. Non-invasive estimation of PVR by cardiac magnetic resonance might be useful for risk stratification in HFrEF, irrespective of etiology, presence of late gadolinium enhancement or LVEF.

Immunopathology of american cutaneous leishmaniasis. Modulation of MHC class II gene products by Keratinocytes before and after glucantime therapy

Epidermal changes from 32 cutaneous and 3 mucosal American leishmaniasis (ACL) active lesions were studied for HLA-DR, -DP expression, Lanerhans cells and lymphocyte infiltration. In addition to a DR and DQ positivity at the surface of the cells of the inflammatory infiltrate, a strong reaction for DR antigens was detected on keratinocytes. Hyperplasia of Langerhans cells was present in al cutaneous lesions and epidermis was infiltrated by T lymphocytes. When healed lesions of 14 of these subjects were re-biopsied 1 to 12 months after the end of pentavalent antimonial therapy, MHC class antigens could no longer be seen on keratinocytes. Our data represrn evidence for hhe reversibility of the abnormal HLA-DR expression by keratinocytes in ACL after Glucantime therapy or spontaneous scar formation, demonstrating that this expresion is restricted to the period of active lesions. The present findings can be regarded as an indirect evidence that keratinocytes may be involved in the immunopathology of ACL.

Molecular and cellular basis of hepatic fibrogenesis in experimental schistosomiasis mansoni infection

Morbidity in schistosomiasis mansoni occurs primaryly as a result of the complications of hepatic fibrosis. Yet, the pathogenesis of schistosomal hepatic fibrosis is poorly understood. The fact that hepatic egg granuloma is the hallmark of this infection suggests a potential role for granulomatous inflamation in hepatic fibrogenesis. Our studies in a murine schistosomiasis model indicate that hepatic granuloma cells secrete a variety of fibrogenic cytokines that may initiate the scarring process. Among these cytokines, we identified a novel protein that we designated fibroplast stimulating factor-1 (FsF-1). FsF-1 is a lymphokine that can stimulate fibroplast growth and matrix synthesis. A notable feature of hepatic fibrosis in this model is that production of FsF-1 and other granuloma-derived fibrogenic cytokines is down-regulated in chronic infection, an event that may be under immunological control. The spontaneous reduction of FsF-1 secretion presumably accounts for reduced scar formation late in infection of mice. In the context of relevant clinical studies, our findings engender the hypothesis that Symmer's fibrosis may develop in a small suppopulation of individuals as a result of immunogenetically-determined dysregulation of fibrogenic cytokine production.

Tuberculin Skin Test and CD4+/CD8+ T Cell Counts in Adults Infected with the Human Immunodeficiency Virus in Medellín, Colombia

To evaluate the effect of BCG vaccination and T lymphocyte subpopulations on the reactivity to the tuberculin skin test, 113 asymtomatic HIV+ individuals were tuberculin tested by intradermal injection of 5TU of purified protein derivative and the levels of circulating lymphocyte (CD3, CD4 and CD8) subpopulations determined by indirect immunofluorescence. Ninety-two percent of the subjects included in the study were males. The mean age of the group was 32.1±7.4 years. Sixty-two percent presented a BCG scar. However, only 22% exhibited positive tuberculin reactions (³5mm) irrespective of the presence of the BCG scar. Tuberculin positive individuals exhibited higher CD4+ cell counts (p=0.004) and CD4+/CD8+ ratios (p=0.006) than tuberculin negative (<5mm) HIV+ individuals. The number of individuals with positive tuberculin reactions was significantly higher in subjects with more than 500 CD4+ lymphocytes/ml (p=0.02) or CD4+/CD8+ ratios ³1.12 (p=0.002). These results suggest that a prior BCG vaccination does not influence the reactivity to the tuberculin skin test in HIV+ asymptomatic individuals and that the number of CD4+ lymphocytes and the CD4+/CD8+ ratio positively correlate with the tuberculin reactivity

Frequency of specific anti-Toxoplasma gondii IgM, IgA and IgE in Colombian patients with acute and chronic ocular toxoplasmosis

We studied the frequency of specific anti-Toxoplasma IgM, IgA and IgE antibodies in serum of 28 immunocompetent Colombian patients, selected by ophthalmologists and with lesions that were compatible with ocular toxoplasmosis. Patients were classified in three groups: (i) group 1 consisted of ten patients with a first episode; (ii) group 2, with seven patients with a recurrence and (iii) group 3, consisted of eleven patients with chronic chorioretinal lesion without uveitis. We found that 10/28 (35%) of Colombian patients with ocular toxoplasmosis possessed at least one serological marker for Toxoplasma infection different from IgG. In group 1 (first episode), we found simultaneous presence of specific IgM plus IgA plus IgE in 1/10 (10%). In group 2 (recurrences) in 1/7 (14%) we found IgM and IgA test positives and in 1/7 (14%) we found IgM and IgE tests positives. In group 3 (toxoplasmic chorioretinal scar) the IgA serological test was positive in 2/11 (18%). These results show that serum IgM or IgA or IgE can be present during recurrences.