Sequence analysis of the internal transcribed spacer 2 (ITS2) region of rDNA for identifying Trichogramma species and evaluating genetic diversity / Análise de sequência do rDNA da região espaçadora do transcrito interno 2 (ITS2) para identificar espécies de Trichogramma e avaliar a diversidade genética

Abstract Species of Trichogramma Westwood, 1833 (Hymenoptera: Trichogrammtidae) are frequently used as biological control agents against Lepidoptera, but practical application of these egg endoparasitoids are complicated because of their complex taxonomy. This study aimed to compare sequences of internal transcribed spacer regions of ribosomal DNA (ITS2-rDNA) of Trichogramma accessions with those deposited in GenBank in order to access the reliability of the ITS2 as a barcode for discriminating species and evaluating the genetic diversity. ITS2-rDNA sequences obtained from seventeen specimens of Trichogramma confirmed previous identifications based on morphological characteristics. Multiple sequence alignment revealed the existence of highly conserved regions in ITS2 sequences while the neighbour-joining dendrogram indicated that the specimens formed three clusters comprising T. manicobai and T. marandobai (group I), T. galloi (group II) and T. pretiosum (group III). The ITS2 marker was shown to be a powerful DNA barcode for discriminating Trichogramma species and could be used to complement the morphological approach.

Resumo Espécies de Trichogramma Westwood, 1833 (Hymenoptera: Trichogrammatidae) são freqüentemente usadas como agentes de controle biológico contra Lepidoptera, esses endoparasitóides de ovos apresentam taxonomia complexa, o que dificulta sua aplicação prática. Este estudo teve como objetivo comparar seqüências de regiões espaçadoras internas transcritas de DNA ribossômico (ITS2-rDNA) de acessos de Trichogramma com aquelas depositadas no GenBank, a fim de avaliar a confiabilidade do ITS2 barcode para discriminar espécies e avaliar a diversidade genética. As seqüências de ITS2-rDNA obtidas de dezessete espécimes de Trichogramma confirmaram identidades anteriores com base em características morfológicas. O alinhamento de múltiplas sequências revelou a existência de regiões altamente conservadas nas sequências ITS2, enquanto o dendrograma indicou que os espécimes formavam três grupos compreendendo T. manicobai e T. marandobai (grupo I), T. galloi (grupo II) e T. pretiosum (grupo III). O marcador ITS2 mostrou ser um poderoso DNA barcode para discriminar espécies de Trichogramma podendo ser usado como complemento da abordagem morfológica.

Sequence analysis of the internal transcribed spacer 2 (ITS2) region of rDNA for identifying Trichogramma species and evaluating genetic diversity.

Species of Trichogramma Westwood, 1833 (Hymenoptera: Trichogrammtidae) are frequently used as biological control agents against Lepidoptera, but practical application of these egg endoparasitoids are complicated because of their complex taxonomy. This study aimed to compare sequences of internal transcribed spacer regions of ribosomal DNA (ITS2-rDNA) of Trichogramma accessions with those deposited in GenBank in order to access the reliability of the ITS2 as a barcode for discriminating species and evaluating the genetic diversity. ITS2-rDNA sequences obtained from seventeen specimens of Trichogramma confirmed previous identifications based on morphological characteristics. Multiple sequence alignment revealed the existence of highly conserved regions in ITS2 sequences while the neighbour-joining dendrogram indicated that the specimens formed three clusters comprising T. manicobai and T. marandobai (group I), T. galloi (group II) and T. pretiosum (group III). The ITS2 marker was shown to be a powerful DNA barcode for discriminating Trichogramma species and could be used to complement the morphological approach.

Impact of chronic obstructive pulmonary disease on linear and nonlinear dynamics of heart rate variability in patients with heart failure

The objective of this study was to investigate the impact of chronic obstructive pulmonary disease (COPD)-heart failure (HF) coexistence on linear and nonlinear dynamics of heart rate variability (HRV). Forty-one patients (14 with COPD-HF and 27 HF) were enrolled and underwent pulmonary function and echocardiography evaluation to confirm the clinical diagnosis. Heart rate (HR) and R-R intervals (iRR) were collected during active postural maneuver (APM) [supine (10 min) to orthostasis (10 min)], respiratory sinus arrhythmia maneuver (RSA-M) (4 min), and analysis of frequency domain, time domain, and nonlinear HRV. We found expected autonomic response during orthostatic changes with reduction of mean iRR, root mean square of successive differences between heart beats (RMSSD), RR tri index, and high-frequency [HF (nu)] and an increased mean HR, low-frequency [LF (nu)], and LF/HF (nu) compared with supine only in HF patients (P<0.05). Patients with COPD-HF coexistence did not respond to postural change. In addition, in the orthostatic position, higher HF nu and lower LF nu and LF/HF (nu) were observed in COPD-HF compared with HF patients. HF patients showed an opposite response during RSA-M, with increased sympathetic modulation (LF nu) and reduced parasympathetic modulation (HF nu) (P<0.05) compared with COPD-HF patients. COPD-HF directly influenced cardiac autonomic modulation during active postural change and controlled breathing, demonstrating an autonomic imbalance during sympathetic and parasympathetic maneuvers compared with isolated HF.

Impact of chronic obstructive pulmonary disease on linear and nonlinear dynamics of heart rate variability in patients with heart failure.

The objective of this study was to investigate the impact of chronic obstructive pulmonary disease (COPD)-heart failure (HF) coexistence on linear and nonlinear dynamics of heart rate variability (HRV). Forty-one patients (14 with COPD-HF and 27 HF) were enrolled and underwent pulmonary function and echocardiography evaluation to confirm the clinical diagnosis. Heart rate (HR) and R-R intervals (iRR) were collected during active postural maneuver (APM) [supine (10 min) to orthostasis (10 min)], respiratory sinus arrhythmia maneuver (RSA-M) (4 min), and analysis of frequency domain, time domain, and nonlinear HRV. We found expected autonomic response during orthostatic changes with reduction of mean iRR, root mean square of successive differences between heart beats (RMSSD), RR tri index, and high-frequency [HF (nu)] and an increased mean HR, low-frequency [LF (nu)], and LF/HF (nu) compared with supine only in HF patients (P<0.05). Patients with COPD-HF coexistence did not respond to postural change. In addition, in the orthostatic position, higher HF nu and lower LF nu and LF/HF (nu) were observed in COPD-HF compared with HF patients. HF patients showed an opposite response during RSA-M, with increased sympathetic modulation (LF nu) and reduced parasympathetic modulation (HF nu) (P<0.05) compared with COPD-HF patients. COPD-HF directly influenced cardiac autonomic modulation during active postural change and controlled breathing, demonstrating an autonomic imbalance during sympathetic and parasympathetic maneuvers compared with isolated HF.

Genetic growth potential characterization in the Japanese quail: a meta-analysis.

The description of the growth of the Japanese quails is necessary to characterize the genetic potential of these birds raised in different countries. Thus, the aim of this study was to describe the genetic potential of Japanese quails by conducting a meta-analysis considering studies conducted in different countries. Only data about the subspecies Coturnix coturnix japonica were considered; studies regarding Coturnix coturnix coturnix were not examined. The criteria investigated were BW (W), age (t), year of publication and location of the study. Each set of genetic material within a publication was coded as one study. The Gompertz function was used to interpret the growth of laying quails; thus, each study was represented by Gompertz parameters. The W and t data were applied to estimate the values of Gompertz growth parameters, including BW at maturity (Wm), BW at birth (Wi), maturity rate (B) and inflection point (IP). The age at which the maximum growth rate was achieved (t*) was calculated considering the parameters Wm, Wi and B. To estimate these parameters, random regression was used to randomize the parameter Wm. The parameters estimated for each assay were used in exploratory, grouping, and principal component analyses. The values of Wi ranged from 4.1 to 11.6 g. The values of B ranged from 0.0393 to 0.1039/day, and consequently, the values of t* and IP ranged from 14 to 31 days and 9.21 to 31.03 g, respectively. These results show that there is considerable variability in the growth potential of Japanese quails. To better understand this variation, two groups were examined: Brazil and other countries, according to the grouping of Wi, Wm, B and t*; parameter B was the variable that presented the highest specificity, indicating that both groups modified the maturity rate. For the principal component analysis, the year of publication showed a relationship with the growth parameters but only for studies performed in Brazil. For studies carried out in other countries, the changes in growth parameters were not related to the year of publication. In Brazilian studies, there was a decrease in the maturity rate, but the weight at maturity was higher. Therefore, it appears that different strategies of genetic selection were adopted in Brazil compared to other countries.

Optical absorption in complexes of abasic DNA with noble-metal nanoclusters by first principles calculations.

Abasic sites (AP site) in a DNA duplex have been experimentally used to produce fluorescent Ag nanoclusters (NC) with a small number of atoms (n ≤ 6). These AP-DNA:NC complexes act as biological makers that help to locate genes associated with diseases related to single nucleotide polymorphisms (SNP), for example. Abasic sites are the most common SNP genetic variation, and their detection may help predict a host of genetically determined diseases. In this work, we report a theoretical study of the optical absorption spectra of AP-DNA:Ag4 and AP-DNA:Au4 complexes using a fully ab initio methodology. We consider several different base environments for the noble-metal nanocluster occupying the AP site, and compute the absorption spectra of sixteen AP-DNA:Ag4 and sixteen AP-DNA:Au4 complexes. We find that optical absorption in the AP-DNA:Ag4 complexes tends to concentrate in the green-to-violet range of frequencies (2.50 eV ≤ hω ≤ 3.2 eV) and that AP-DNA:Au4 complexes display absorption peaks in the violet-to-ultraviolet interval (hω ≥ 3.0 eV). An analysis of the optical absorption mechanisms in these complexes shows that they can be of local, charge-transfer, or hybrid nature, i.e., AP-DNA:NC complexes display the full variety of optical absorption processes in molecular systems. In particular, we identify both charge-transfer and hybrid processes involving several DNA bases surrounding the NC. Importantly, we find that even sequences where the Ag4 cluster is not in a guanine rich neighborhood display absorption peaks in the visible-light spectrum. Moreover, we obtain that the maximum intensities of the absorption peaks in complexes with pyrimidine vacancies are generally higher than those in complexes with purine vacancies. Regarding the selectivity of single-vacancy AP-DNA to specific noble-metal nanocluster sizes, our calculations show that the four-atom Ag4 (Au4) species fits naturally and binds into the AP-site in a single-vacancy AP-DNA.

Sequence analysis of the internal transcribed spacer 2 (ITS2) region of rDNA for identifying Trichogramma species and evaluating genetic diversity

Abstract Species of Trichogramma Westwood, 1833 (Hymenoptera: Trichogrammtidae) are frequently used as biological control agents against Lepidoptera, but practical application of these egg endoparasitoids are complicated because of their complex taxonomy. This study aimed to compare sequences of internal transcribed spacer regions of ribosomal DNA (ITS2-rDNA) of Trichogramma accessions with those deposited in GenBank in order to access the reliability of the ITS2 as a barcode for discriminating species and evaluating the genetic diversity. ITS2-rDNA sequences obtained from seventeen specimens of Trichogramma confirmed previous identifications based on morphological characteristics. Multiple sequence alignment revealed the existence of highly conserved regions in ITS2 sequences while the neighbour-joining dendrogram indicated that the specimens formed three clusters comprising T. manicobai and T. marandobai (group I), T. galloi (group II) and T. pretiosum (group III). The ITS2 marker was shown to be a powerful DNA barcode for discriminating Trichogramma species and could be used to complement the morphological approach.

Resumo Espécies de Trichogramma Westwood, 1833 (Hymenoptera: Trichogrammatidae) são freqüentemente usadas como agentes de controle biológico contra Lepidoptera, esses endoparasitóides de ovos apresentam taxonomia complexa, o que dificulta sua aplicação prática. Este estudo teve como objetivo comparar seqüências de regiões espaçadoras internas transcritas de DNA ribossômico (ITS2-rDNA) de acessos de Trichogramma com aquelas depositadas no GenBank, a fim de avaliar a confiabilidade do ITS2 barcode para discriminar espécies e avaliar a diversidade genética. As seqüências de ITS2-rDNA obtidas de dezessete espécimes de Trichogramma confirmaram identidades anteriores com base em características morfológicas. O alinhamento de múltiplas sequências revelou a existência de regiões altamente conservadas nas sequências ITS2, enquanto o dendrograma indicou que os espécimes formavam três grupos compreendendo T. manicobai e T. marandobai (grupo I), T. galloi (grupo II) e T. pretiosum (grupo III). O marcador ITS2 mostrou ser um poderoso DNA barcode para discriminar espécies de Trichogramma podendo ser usado como complemento da abordagem morfológica.

Differential physiological response of the grapevine varieties Touriga Nacional and Trincadeira to combined heat, drought and light stresses.

Worldwide, extensive agricultural losses are attributed to drought, often in combination with heat in Mediterranean climate regions, where grapevine traditionally grows. The available scenarios for climate change suggest increases in aridity in these regions. Under natural conditions plants are affected by a combination of stresses, triggering synergistic or antagonistic physiological, metabolic or transcriptomic responses unique to the combination. However the study of such stresses in a controlled environment can elucidate important mechanisms by allowing the separation of the effects of individual stresses. To gather those effects, cuttings of two grapevine varieties, Touriga Nacional (TN) and Trincadeira (TR), were grown under controlled conditions and subjected to three abiotic stresses (drought - WS, heat - HS and high light - LS) individually and in combination two-by-two (WSHS, WSLS, HSLS) or all three (WSHSLS). Photosynthesis, water status, contents of H2 O2 , abscisic acid and metabolites of the ascorbate-glutathione cycle were measured in the leaves. Common and distinct response features were identified in the different stress combinations. Photosynthesis was not hindered in TN by LS, while even individual stresses severely affect photosynthesis in TR. Abscisic acid may be implicated in grapevine osmotic responses since it is correlated with tolerance parameters, especially in combined stresses involving drought. Overall, the responses to drought-including treatments were clearly distinct to those without drought. From the specific behaviours of the varieties, it can be concluded that TN shows a higher capacity for heat dissipation and for withstanding high light intensities, indicating better adjustment to warm conditions, provided that water supply is plentiful.

Detection of parasite antigens in Leishmania infantum-infected spleen tissue by monoclonal antibody-, piezoelectric-based immunosensors.

Diseases such as leishmaniases are important causes of morbidity and mortality in Brazil, and their diagnoses need to be improved. The use of monoclonal antibodies has ensured high specificity to immunodiagnosis. The development of an immunosensor, coupling a monoclonal antibody to a bioelectronic device capable of quickly detecting Leishmania sp. antigens both qualitatively and quantitatively, is a promising alternative for the diagnosis of leishmaniasis due to its high specificity, low cost, and portability, compared with conventional methods. The present work was aimed at developing an immunosensor-based assay for detecting Leishmania infantum antigens in tissues of infected hosts. Four hybridomas producing monoclonal antibodies against L. infantum had their specificity confirmed by enzyme-linked immunosorbent assay. These antibodies were immobilized on a gold surface, covered with a thin film of 2-aminoethanethiol (cysteamine) and glutaraldehyde, blocked with glycine, and placed into contact with extracts of L. infantum -infected and noninfected control hamster spleens. The assay was able to detect 1.8 × 10(4) amastigotes/g of infected tissue. These results demonstrated that this assay may be useful for quantifying L. infantum amastigotes in organs of experimental animals for studies on pathogenesis and immunity and that it is a promising tool for the development of a diagnostic method, based on antigen detection, of human and dog visceral leishmaniasis.

The influence of fish culture in floating net cages on microbial indicators of water quality.

This work was carried out to analyse the microbiological parameters of the water quality of a reservoir used for the irrigation and culture of tilapia (Oreochromis niloticus) in floating net cages. The physico-chemical parameters, counts of mesophilic total aerobic bacteria, total and thermotolerant coliforms and fecal streptococci, and the presence of Escherichia coli in samples of water collected in three sites of the reservoir (pre-culture site, culture site, post-culture site) were analysed. The levels of ammonia (0.047-0.059 mg/L), nitrite (0.001-0.021 mg/L) and total phosphorus (0.050-0.355 mg/L) in the water did not show significant differences (p > 0.05) between sampling sites. The levels of total bacteria in the water varied between 1.3 x 10(4) and 67.3 x 10(4) CFU/100 mL. The MPN values of thermotolerant coliforms (< 930 MPN/100 mL) were within values recommended for water used for fish culture and/or irrigation. The presence of E. coli and fecal streptococci were verified in 48% and 56% of analysed samples, respectively. The site with floating net cages showed more samples contaminated with E. coli and fecal streptococci than other sampling points.

The influence of fish culture in floating net cages on microbial indicators of water quality / Influencia da piscicultura em tanques-rede sobre os microrganismos indicadores da qualidade de agua

This work was carried out to analyse the microbiological parameters of the water quality of a reservoir used for the irrigation and culture of tilapia (Oreochromis niloticus) in floating net cages. The physico-chemical parameters, counts of mesophilic total aerobic bacteria, total and thermotolerant coliforms and fecal streptococci, and the presence of Escherichia coli in samples of water collected in three sites of the reservoir (pre-culture site, culture site, post-culture site) were analysed. The levels of ammonia (0.047-0.059 mg/L), nitrite (0.001-0.021 mg/L) and total phosphorus (0.050-0.355 mg/L) in the water did not show significant differences (p > 0.05) between sampling sites. The levels of total bacteria in the water varied between 1.3 x 104 and 67.3 x 104 CFU/100 mL. The MPN values of thermotolerant coliforms (< 930 MPN/100 mL) were within values recommended for water used for fish culture and/or irrigation. The presence of E. coli and fecal streptococci were verified in 48% and 56% of analysed samples, respectively. The site with floating net cages showed more samples contaminated with E. coli and fecal streptococci than other sampling points.

Este trabalho foi realizado para analisar os parâmetros microbiológicos da qualidade da água de um reservatório destinado a irrigação e à cultivo de tilapia do Nilo (Oreochromis niloticus) em tanques redes. Foi realizada análise de parâmetros físico-químicos, quantificação de bactérias totais aeróbias mesófilas, coliformes totais e termotolerantes, estreptococos fecais e a detecção de E. coli em amostras de água coletada em três pontos distintos do açude (pré-cultivo, cultivo, pós-cultivo). Os níveis de amônia (0,047-0,059 mg/L), nitrito (0,001-0,021 mg/L) e fósforo total (0,050-0,355 mg/L) na água não apresentaram diferenças significativas (p > 0.05) entre os pontos de coleta. As contagens de bactérias aeróbias mesófilas variaram entre 1,3 x 104 e 67,3 x 104 UFC/100 mL nas amostras de água. O NMP de coliformes termotolerantes (< 930 NMP/100 mL) se apresentou dentro do limite permitido para águas doces destinadas a piscicultura e/ou irrigação. A presença de E. coli foi constatada em 48% e de estreptococos fecais em 56% das amostras de água analisadas. O local de cultivo de tilápia em tanques-rede apresentou maior número de amostras contaminadas com E. coli e estreptococos fecais que outros pontos de amostragem.

A proteomic approach to identify proteins from Trichuris trichiura extract with immunomodulatory effects.

Infections with Trichuris trichiura and other trichurid nematodes have been reported to display protective effects against atopy, allergic and autoimmune diseases. The aims of the present study were to investigate the immunomodulatory properties of T. trichiura adult worm extract (TtE) and its fractions (TtEFs) on the production of cytokines by peripheral blood mononuclear cells and to identify their proteinaceous components. Fourteen TtEFs were obtained by ion exchange chromatography and tested for effects on cytokine production by peripheral blood mononuclear cells. The molecular constituents of the six most active fractions were evaluated using nano-LC/mass spectrometry. The homology between T. trichiura and the related nematode Trichinella spiralis was used to identify 12 proteins in TtEFs. Among those identified, fructose biphosphate aldolase, a homologue of macrophage migration inhibitory factor and heat-shock protein 70 may contribute to the immunomodulatory effects of TtEFs. The identification of such proteins could lead to the development of novel drugs for the therapy of allergic and other inflammatory diseases.

Reversal of the anticoagulant and anti-hemostatic effect of low molecular weight heparin by direct prothrombin activation

Lopap, found in the bristles of Lonomia obliqua caterpillar, is the first exogenous prothrombin activator that shows serine protease-like activity, independent of prothrombinase components and unique lipocalin reported to interfere with hemostasis mechanisms. To assess the action of an exogenous prothrombin activator reversing the anticoagulant and antihemostatic effect induced by low molecular weight heparin (LMWH), male New Zealand rabbits (N = 20, weighing 3.8-4.0 kg) allocated to 4 groups were anticoagulated with 1800 IU/kg LMWH (iv) over 2 min, followed by iv administration of saline (SG) or recombinant Lopap (rLopap) at 1 µg/kg (LG1) or 10 µg/kg (LG10), 10 min after the injection of LMWH, in a blind manner. Control animals (CG) were treated only with saline. The action of rLopap was assessed in terms of activated partial thromboplastin time (aPTT), prothrombin fragment F1+2, fibrinogen, and ear puncture bleeding time (BT) at 5, 10, 15, 17, 20, 30, 40, 60, and 90 min after initiation of LMWH infusion. LG10 animals showed a decrease of aPTT in more than 50% and BT near to normal baseline. The level of prothrombin fragment F1+2 measured by ELISA had a 6-fold increase with rLopap treatment (10 µg/kg) and was inversely proportional to BT in LMWH-treated animals. Thus, Lopap, obtained in recombinant form using E. coli expression system, was useful in antagonizing the effect of LMWH through direct prothrombin activation, which can be a possible strategy for the reversal of bleeding and anticoagulant events.

Reversal of the anticoagulant and anti-hemostatic effect of low molecular weight heparin by direct prothrombin activation.

Lopap, found in the bristles of Lonomia obliqua caterpillar, is the first exogenous prothrombin activator that shows serine protease-like activity, independent of prothrombinase components and unique lipocalin reported to interfere with hemostasis mechanisms. To assess the action of an exogenous prothrombin activator reversing the anticoagulant and antihemostatic effect induced by low molecular weight heparin (LMWH), male New Zealand rabbits (N = 20, weighing 3.8-4.0 kg) allocated to 4 groups were anticoagulated with 1800 IU/kg LMWH (iv) over 2 min, followed by iv administration of saline (SG) or recombinant Lopap (rLopap) at 1 µg/kg (LG1) or 10 µg/kg (LG10), 10 min after the injection of LMWH, in a blind manner. Control animals (CG) were treated only with saline. The action of rLopap was assessed in terms of activated partial thromboplastin time (aPTT), prothrombin fragment F1+2, fibrinogen, and ear puncture bleeding time (BT) at 5, 10, 15, 17, 20, 30, 40, 60, and 90 min after initiation of LMWH infusion. LG10 animals showed a decrease of aPTT in more than 50% and BT near to normal baseline. The level of prothrombin fragment F1+2 measured by ELISA had a 6-fold increase with rLopap treatment (10 µg/kg) and was inversely proportional to BT in LMWH-treated animals. Thus, Lopap, obtained in recombinant form using E. coli expression system, was useful in antagonizing the effect of LMWH through direct prothrombin activation, which can be a possible strategy for the reversal of bleeding and anticoagulant events.

Serine-like proteolytic enzymes correlated with differential pathogenicity in patients with acute Acanthamoeba keratitis.

Acute ocular infection due to free-living amoebae of the genus Acanthamoeba is characterized by severe pain, loss of corneal transparency and, eventually, blindness. Proteolytic enzymes secreted by trophozoites of virulent Acanthamoeba strains have an essential role in the mechanisms of pathogenesis, including adhesion, invasion and destruction of the corneal stroma. In this study, we analysed the relationship between the extracellular proteases secreted by clinical isolates of Acanthamoeba and the clinical manifestations and severity of disease that they caused. Clinical isolates were obtained from patients who showed typical symptoms of Acanthamoeba keratitis. Trophozoites were cultivated axenically, and extracellular proteins were collected from cell culture supernatants. Secreted enzymes were partially characterized by gelatin and collagen zymography. Acanthamoeba trophozoites secreted proteases with different molecular masses, proteolysis rates and substrate specificities, mostly serine-like proteases. Different enzymatic patterns of collagenases were observed, varying between single and multiple collagenolytic activities. Low molecular weight serine proteases were secreted by trophozoites associated with worse clinical manifestations. Consequently, proteolytic enzymes of some Acanthamoeba trophozoites could be related to the degree of their virulence and clinical manifestations of disease in the human cornea.

Serine-like proteolytic enzymes correlated with differential pathogenicity in patients with acute Acanthamoeba keratitis

Acute ocular infection due to free-living amoebae of the genus Acanthamoeba is characterized by severe pain, loss of corneal transparency and, eventually, blindness. Proteolytic enzymes secreted by trophozoites of virulent Acanthamoeba strains have an essential role in the mechanisms of pathogenesis, including adhesion, invasion and destruction of the corneal stroma. In this study, we analysed the relationship between the extracellular proteases secreted by clinical isolates of Acanthamoeba and the clinical manifestations and severity of disease that they caused. Clinical isolates were obtained from patients who showed typical symptoms of Acanthamoeba keratitis. Trophozoites were cultivated axenically, and extracellular proteins were collected from cell culture supernatants. Secreted enzymes were partially characterized by gelatin and collagen zymography. Acanthamoeba trophozoites secreted proteases with different molecular masses, proteolysis rates and substrate specificities, mostly serine-like proteases. Different enzymatic patterns of collagenases were observed, varying between single and multiple collagenolytic activities. Low molecular weight serine proteases were secreted by trophozoites associated with worse clinical manifestations. Consequently, proteolytic enzymes of some Acanthamoeba trophozoites could be related to the degree of their virulence and clinical manifestations of disease in the human cornea.

Craniocervical posture analysis in patients with temporomandibular disorder / Análise da postura cranio-cervical em pacientes com disfunção temporomandibular

OBJECTIVE: To compare head positioning and cervical spine alignment between individuals with and without temporomandibular disorders (TMDs), by means of positional evaluation using photographs, radiographs and visual observation, and to investigate whether the type of TMD influences head posture and cervical spine positioning. METHODS: Ninety randomly chosen women were diagnosed using the research diagnostic criteria for TMDs (RDC/TMD) by a trained examiner and were divided into three groups: Group 1, with a diagnosis of myofascial dysfunction (group I of RDC axis I); Group 2, with mixed TMD (groups I, II and III of RDC axis I); and Control, without TMD. Following this, the participants were photographed in frontal and lateral views by a single examiner. To produce these photos, the following anatomical points were marked out on the skin: occipital protuberance, C4, C7, acromioclavicular joint and sternoclavicular joint. From these points, different angles were analyzed by means of the ALCimagem-2000 application. These same photos were then evaluated qualitatively (visual evaluation). Next, lateral teleradiography and radiography of the cervical spine was requested. The examiner was blind when analyzing the images. To compare the results, the chi-squared test and analysis of variance were used, with significance levels of 5 percent. RESULTS: Regardless of the method used, the results revealed that head and cervical spine posture did not differ between the groups with and without TMD, independent of the diagnostic group. CONCLUSION: The posture of individuals with myogenic or arthrogenous TMD does not differ from the posture of individuals without TMD. The presence of TMD does not influence the head and cervical spine posture.

OBJETIVO: Comparar o posicionamento da cabeça e o alinhamento da coluna cervical em indivíduos com e sem DTM, por meio da avaliação postural por fotografias, radiografias e por observação visual e verificar se o tipo de DTM influencia na postura da cabeça e no posicionamento da coluna cervical. MÉTODOS: Noventa mulheres escolhidas aleatoriamente foram diagnosticadas por meio dos Critérios para Diagnóstico em Pesquisa para Disfunções Temporomandibulares (RDC/TMD) por um examinador treinado e divididas em três grupos: grupo 1, diagnóstico de disfunção miofascial (grupo I do eixo I do RDC); grupo 2, com DTM mista (grupo I, II e III do eixo I do RDC) e controle, sem DTM . Em seguida, foram fotografadas em vista anterior e em perfil por um único examinador. Para a realização dessas fotografias, foram demarcados sobre a pele os seguintes pontos anatômicos: protuberância occiptal, C4, C7, articulação acromioclavicular e esternoclavicular. A partir desses pontos, foram analisados diferentes ângulos por meio do aplicativo ALCimagem-2000. Essas mesmas fotos foram posteriormente avaliadas qualitativamente (avaliação visual). Em seguida, foi solicitada uma radiografia e uma telerradiografia em perfil. O examinador foi cego ao analisar as imagens. Para comparação dos resultados, foi utilizado o teste qui-quadrado e a análise de variância com nível de significância de 5 por cento. RESULTADOS: Independentemente do método utilizado, os resultados revelaram que a postura da cabeça e da coluna cervical não diferem entre o grupo com DTM e sem DTM, independentemente do grupo diagnosticado. CONCLUSÃO: A postura do indivíduo com DTM miogênica ou artrogênica não é diferente do indivíduo sem DTM. A presença da DTM não influencia na postura da cabeça e da coluna cervical.

Angiotensin II-mediated vasodilation is reduced in adult spontaneously hypertensive rats despite enhanced expression of AT2 receptors.

1. The vasodilator action of angiotensin (Ang) II has not yet been demonstrated in spontaneously hypertensive rats (SHR), nor have any possible changes in this response during the development of hypertension. 2. In the present study, the vasodilator effect of AngII was evaluated in the rat isolated, preconstricted mesenteric arterial bed (MAB) from 6- (young) and 24-week-old (adult) SHR and compared with effects on MAB from age-matched normotensive rats (control). 3. Angiotensin II (10-300 nmol) induced vasodilation in noradrenaline (NA)-preconstricted MAB that was greater in vessels from young compared with adult rats in both the control and SHR groups. Angiotensin II-induced vasodilation was reduced by the angiotensin AT(2) receptor antagonist PD 123319 (10 micromol/L), the angiotensin-(1-7) receptor antagonist A779 (1 micromol/L) and the bradykinin B(2) receptor antagonist HOE-140 (0.01 micromol/L), but not by the AT(1) receptor antagonist losartan (30 micromol/L). Expression of AT(2) receptors was weak in vessels from adult control rats compared with that in young control rats, whereas in young SHR AT(2) receptor expression was increased compared with that in young control rats. This increased expression of AT(2) receptors was maintained in adult SHR and there was no significant difference in AT(2) receptor expression between young and old SHR. 4. The findings of the present suggest that AngII induces an AT(2) receptor-mediated vasodilator effect in the MAB via activation of angiotensin-(1-7) and bradykinin receptors, an action that is reduced in adult control rats and adult SHR. In adult control rats, the attenuated response of AngII is probably due to endothelial dysfunction and reduced expression of AT(2) receptors, whereas in adult SHR it is associated with endothelial dysfunction alone. Increased expression of AT(2) receptors in SHR may represent a counteracting response for modulating blood pressure.