Characterization of leptospiral proteins that afford partial protection in hamsters against lethal challenge with Leptospira interrogans

Leptospirosis is a worldwide zoonosis caused by pathogenic Leptospira. The whole-genome sequence of Leptospira interrogans serovar Copenhageni together with bioinformatic tools allow us to search for novel antigen candidates suitable for improved vaccines against leptospirosis. This study focused on three genes encoding conserved hypothetical proteins predicted to be exported to the outer membrane. The genes were amplified by PCR from six predominant pathogenic serovars in Brazil. The genes were cloned and expressed in Escherichia coli strain BL21-SI using the expression vector pDEST17. The recombinant proteins tagged with N-terminal 6xHis were purified by metal-charged chromatography. The proteins were recognized by antibodies present in sera from hamsters that were experimentally infected. Immunization of hamsters followed by challenge with a lethal dose of a virulent strain of Leptospira showed that the recombinant protein rLIC12730 afforded statistically significant protection to animals (44 %), followed by rLIC10494 (40 %) and rLIC12922 (30 %). Immunization with these proteins produced an increase in antibody titres during subsequent boosters, suggesting the involvement of a T-helper 2 response. Although more studies are needed, these data suggest that rLIC12730 and rLIC10494 are promising candidates for a multivalent vaccine for the prevention of leptospirosis.

Humoral responses and immune protection in mice immunized with irradiated T. gondii tachyzoites and challenged with three genetically distinct strains of T. gondii

Toxoplasma gondii is an obligate intracellular parasite that infects a variety of mammals and birds. T. gondii also causes human toxoplasmosis; although toxoplasmosis is generally a benign disease, ocular, congenital or reactivated disease is associated with high numbers of disabled people. Infection occurs orally through the ingestion of meat containing cysts or by the intake of food or water contaminated with oocysts. Although the immune system responds to acute infection and mediates the clearance of tachyzoites, parasite cysts persist for the lifetime of the host in tissues such as the eye, muscle, and CNS. However, T. gondii RH strain tachyzoites irradiated with 255 Gy do not cause residual infection and induce the same immunity as a natural infection. To assess the humoral response in BALB/c and C57BL/6J mice immunized with irradiated tachyzoites either by oral gavage (p.o.) or intraperitoneal (i.p.) injection, we analyzed total and high-affinity IgG and IgA antibodies in the serum. High levels of antigen-specific IgG were detected in the serum of parenterally immunized mice, with lower levels in mice immunized via the oral route. However, most serum antibodies exhibited low affinity for antigen in both mice strain. We also found antigen specific IgA antibodies in the stools of the mice, especially in orally immunized BALB/c mice. Examination of bone marrow and spleen cells demonstrated that both groups of immunized mice clearly produced specific lgG, at levels comparable to chronic infection, suggesting the generation of IgG specific memory. Next, we challenged i.p. or p.o. immunized mice with cysts from ME49. VEG or P strains of T. gondii. Oral immunization resulted in partial protection as compared to challenged naive mice: these findings were more evident in highly pathogenic ME49 strain challenge. Additionally, we found that while mucosal IgA was important for protection against infection, antigen-specific IgG antibodies were involved with protection against disease and disease pathogenesis. Most antigen responsive cells in culture produced specific high-affinity IgG after immunization, diverse of the findings in serum IgG or from cells after infection, which produced low proportion of high-avidity IgG. (C) 2011 Elsevier B.V. All rights reserved.

Changes in Perfusion-Weighted Magnetic Resonance Imaging after Carotid Angioplasty with Stent

Carotid artery stenosis due to arteriosclerosis increases the risk of cerebral ischemia via embolic phenomena or reduced blood flow. The changes in cerebral perfusion that may occur after treatment are not clearly understood. This study evaluated the changes in cerebral microcirculation following carotid angioplasty with stenting (CAS) under cerebral protection with filters using ultrafast gradient echo (GRE) perfusion weighted imaging (PWI) with magnetic resonance imaging (MRI). Prospectively, 21 cervical carotid stenosis patients, mean age 69.95 years, underwent MRI 12 h before and 72 h after CAS. PWI parameters were collected for statistical analysis: cerebral blood volume (CB V), mean transit time (MTT) and time to peak (TTP). Statistical analysis was applied to absolute parameters and to values normalized against those from the contralateral parenchyma. The main finding of this study was improved hemodynamics for the normalized data after CAS, shown by reduced MTT (p<0.001) and TTP (p=0.019) in the territory fed by the middle cerebral artery ipsilateral to the CAS. Absolute data showed increased blood volume in the cerebral hemispheres after CAS, which was more accentuated on the stent side (p=0.016) than the contralateral side (p=0.029). Early improvements in cerebral perfusion, mainly seen in the normalized data, were clearly demonstrated in the timing parameters - TTP & MTT - after CAS.

Reliability assessment of Cobb angle measurements using manual and digital methods

BACKGROUND CONTEXT: The vertebral spine angle in the frontal plane is an important parameter in the assessment of scoliosis and may be obtained from panoramic X-ray images. Technological advances have allowed for an increased use of digital X-ray images in clinical practice. PURPOSE: In this context, the objective of this study is to assess the reliability of computer-assisted Cobb angle measurements taken from digital X-ray images. STUDY DESIGN/SETTING: Clinical investigation quantifying scoliotic deformity with Cobb method to evaluate the intra- and interobserver variability using manual and digital techniques. PATIENT SAMPLE: Forty-nine patients diagnosed with idiopathic scoliosis were chosen based on convenience, without predilection for gender, age, type, location, or magnitude of the curvature. OUTCOME MEASURES: Images were examined to evaluate Cobb angle variability, end plate selection, as well as intra- and interobserver errors. METHODS: Specific software was developed to digitally reproduce the Cobb method and calculate semiautomatically the degree of scoliotic deformity. During the study, three observers estimated the Cobb angle using both the digital and the traditional manual methods. RESULTS: The results showed that Cobb angle measurements may be reproduced in the computer as reliably as with the traditional manual method, in similar conditions to those found in clinical practice. CONCLUSIONS: The computer-assisted method (digital method) is clinically advantageous and appropriate to assess the scoliotic curvature in the frontal plane using Cobb method. (C) 2010 Elsevier Inc. All rights reserved.

Validity and reliability of a computer method to estimate vertebral axial rotation from digital radiographs

Axial vertebral rotation, an important parameter in the assessment of scoliosis may be identified on X-ray images. In line with the advances in the field of digital radiography, hospitals have been increasingly using this technique. The objective of the present study was to evaluate the reliability of computer-processed rotation measurements obtained from digital radiographs. A software program was therefore developed, which is able to digitally reproduce the methods of Perdriolle and Raimondi and to calculate semi-automatically the rotation degree of vertebra on digital radiographs. Three independent observers estimated vertebral rotation employing both the digital and the traditional manual methods. Compared to the traditional method, the digital assessment showed a 43% smaller error and a stronger correlation. In conclusion, the digital method seems to be reliable and enhance the accuracy and precision of vertebral rotation measurements.

Mycobacterium tuberculosis Culture Filtrate Proteins plus CpG Oligodeoxynucleotides Confer Protection to Mycobacterium bovis BCG-Primed Mice by Inhibiting Interleukin-4 Secretion

Culture filtrate proteins (CFP) are potential targets for tuberculosis vaccine development. We previously showed that despite the high level of gamma interferon (IFN-gamma) production elicited by homologous immunization with CFP plus CpG oligodeoxynucleotides (CFP/CpG), we did not observe protection when these mice were challenged with Mycobacterium tuberculosis. In order to use the IFN-gamma-inducing ability of CFP antigens, in this study we evaluated a prime-boost heterologous immunization based on CFP/CpG to boost Mycobacterium bovis BCG vaccination in order to find an immunization schedule that could induce protection. Heterologous BCG-CFP/CpG immunization provided significant protection against experimental tuberculosis, and this protection was sustained during the late phase of infection and was even better than that conferred by a single BCG immunization. The protection was associated with high levels of antigen-specific IFN-gamma and interleukin-17 (IL-17) and low IL-4 production. The deleterious role of IL-4 was confirmed when IL-4 knockout mice vaccinated with CFP/CpG showed consistent protection similar to that elicited by BCG-CFP/CpG heterologous immunization. These findings show that a single dose of CFP/CpG can represent a new strategy to boost the protection conferred by BCG vaccination. Moreover, different immunological parameters, such as IFN-gamma and IL-17 and tightly regulated IL-4 secretion, seem to contribute to the efficacy of this tuberculosis vaccine.

CpG-ODN combined with Neospora caninum lysate, but not with excreted-secreted antigen, enhances protection against infection in mice

CpG oligodeoxynucleotides (ODN) have shown to be potent immunoadjuvants for several pathogens, but there is limited information concerning their use in immunization protocols against neosporosis. This study aimed to evaluate the potential of CpG-ODN combined with Neosporar lysate antigen (NLA) or excreted-secreted antigen (NcESA) to induce protective immune response against Neospora caninum infection in mice. C57BL/6 mice were vaccinated subcutaneously three times at 2-week intervals with NLA, NLA+CpG, NcESA, NcESA+CpG, CpG (adjuvant control) or PBS (infection control). Serological assays showed an increased specific IgG2a response in animals immunized with either antigen plus adjuvant and elevated levels of the IgG1 isotype in those vaccinated with antigens alone. Splenocyte proliferative responses upon antigen stimulation were higher in groups immunized with NLA OF NcESA combined with CpG, showing increased IL-12 levels. Also, mice vaccinated with NcESA or NcESA+CpG demonstrated higher IFN-gamma levels and IFN-gamma/IL-10 ratio. After lethal challenge, mice immunized with NLA+CpG or NLA had lower Morbidity score and body weight changes in comparison to other groups, and animals did not succumb during acute infection. In contrast, NcESA+CpG or NcESA groups exhibited the highest morbidity scores, body weight impairment and mortality rates, associated with greatest brain parasite burden and inflammation. In conclusion, CpG-ODN was able to induce a Th1-type humoral immune response with predominant IgG2a levels for either NLA or NcESA, but resulting in an effective Th1-driven cellular immune response and total Protection only when combined with NLA. Vaccination with NcESA alone or combined with CpG resulted in a strong cellular immune response associated with high levels of IFN-gamma and inflammation, rendering mice more susceptible to parasite challenge. (C) 2009 Elsevier Ltd. All rights reserved.

Bovine Peritoneum Protection Role on Intestinal Adhesions to a Vascular Graft: A Morphological Analysis

The present study aimed to experimentally evaluate the protection role of glycerin preserved bovine peritoneum (BP) against intestinal adhesions to a vascular graft. Experiments were performed on 24 adult rabbits, randomly dived into two groups. All animals were submitted to a vascular graft over the infra-renal aorta and vena cava. Group I (12 animals) was submitted to a BP patch on the retroperitoneal opening, between the vascular prosthetic graft and the intestinal loops. Group II (12 animals) had the retroperitoneal opening sutured. After 7, 14, 28 and 60 days, 3 animals of each group were randomly killed and the retro peritoneum, with or without the BP patch, was removed for histological analysis. The histological analysis showed that the BP stimulated a moderate to intense inflammatory reaction at the beginning of the experiments and on the 60-day evaluation, the inflammatory reaction was mild, limited to the BP border with its histological structure preserved. In conclusion, the BP is a safe and cheap interposition material to be used between vascular grafts and intestinal loops, presenting a protection role against adhesions between them.

HLA-DRB1*allele-associated genetic susceptibility and protection against multiple sclerosis in Brazilian patients

Multiple sclerosis (MS) is a chronic autoimmune demyelinating disease of the central nervous system that causes neurological disorders in young adults. Previous studies in various populations highlighted an association between the HLA-DRB1*1.5 allele and MS. This study investigated the association between HLA-DRB1*15 and other HLA-DRB1 alleles and MS in a Brazilian Caucasian population sample from Londrina, Southern Brazil. HLA-DRB1 alleles were analyzed by polymerase chain reaction with specific sequence oligonucleotide primers in 119 MS patients and in 305 healthy blood donors as a control. Among the MS patients, 89 (75.0%) presented with relapsing remitting MS, 24 (20.0%) with secondary progressive MS and 6 (5.0%) with primary progressive MS. The frequency of the HLA-DRB1*15 allele observed in the MS Brazilian patients was similar to findings reported in previous studies carried out in populations worldwide. However, the results showed a higher frequency of the HLA-DRB1*15 allele in the MS patients compared to the controls, with a relative frequency of 0.1050 (10.50%) and 0.0443 (4.4%), respectively (OR=2.53; 95% CI 1.43-4.46; p=0.0009). A protector allele was also detected. The frequency of the HLA-DRB1*11 allele was reduced in the MS patients compared to the controls, with a relative frequency of 0.1345 (13.4%) and 0.1869 (18.7%), respectively (OR=0.67; 95% CI 0.44-1.03; p=0.0692). The results demonstrated that the HLA-DRB1*15 allele in heterozygosity is positively associated with MS (p=0.0079), and may be considered a genetic marker of susceptibility to the disease. A negative association between the HLA-DRB1*11 allele in homozygosity and MS was also verified (p=0.0418); this allele may be considered a genetic marker of resistance to MS in the Brazilian population.

TNF microsatellite alleles may confer protection against the development of lipodystrophy syndrome in Brazilian HIV patients

P>The aim of this study was to evaluate the frequency of TNFa-e microsatellites and the promoter region (TNF-308 and TNF-238) in HIV/AIDS-infected patients presenting or not lipodystrophy syndrome (LS). The design is the genetic case-control association study. Microsatellite and the TNF promoter region polymorphisms were amplified by PCR and submitted to polyacrylamide gel electrophoresis. The genotypes and allele frequencies for 67 HIV-positive patients with lipodystrophy were compared with 50 HIV-positive patients with no evidence of lipodystrophy and with 131 healthy HIV-negative individuals. The presence of the TNFa5 allele could provide HIV/AIDS patients with protection against developing LS. The presence of TNF-308G allele, as well as of its homozygote TNF-308GG, were associated with susceptibility to developing LS. In addition, the presence of the haplotype TNFe3-d3-238G-308A-c1-a5-b7 suggests protection against developing that syndrome. This study highlights that polymorphic sites spanning the region nearby the TNF locus are associated with LS development in HIV/AIDS patients.

Comparison of pelvic floor muscle strength between women undergoing vaginal delivery, cesarean section, and nulliparae using a perineometer and digital palpation

Objectives. To compare pelvic floor muscle (PFM) strength between women undergoing vaginal delivery, cesarean section, and nulliparae, investigating the factors associated with PFM strength, and observing the correlation between vaginal digital palpation and use of a perineometer. Methods. A cross-sectional study was conducted, including 31 women following vaginal delivery, 30 women following cesarean section, and 30 nulliparous women. PFM strength was measured by vaginal digital palpation and use of a perineometer. Multiple linear regression analysis with adjustment for covariables was used to compare the mean PFM strength and identify its associated factors. Results. The mean PFM strength of women undergoing vaginal delivery and cesarean section was 25.6 +/- 14.5 cmH(2)O and 39.6 +/- 22.0 cmH(2)O (p < 0.01, adjusted for covariables), respectively. A correlation was observed between measurements of PFM strength obtained by vaginal digital palpation and use of a perineometer (tau = 0.82; p < 0.01). The non-white race/ethnicity was negatively associated with PFM strength (coefficient: -10.2424; p = 0.02). Conclusions. A lower PFM strength was observed in women with a history of vaginal delivery compared to those undergoing cesarean section. Non-white race/ethnicity negatively affected PFM strength. Our data suggest that vaginal digital palpation may be used in clinical practice because of its expressive correlation with use of a perineometer.

Gentamicin attenuates gentamicin-induced ototoxicity - Self-protection

Aminoglycoside antibiotics cause considerable toxicity to the inner ear. A progressive hearing loss at high frequencies resulted from the loss of hair cells in the base of the cochlea and a constant preoccupation with finding a treatment that protects against their toxic effects. A self-protection phenomenon to high ototoxic doses of gentamicin is proposed in this paper. Thirty-eight adult guinea pigs with normal hearing were tested using Preyer`s reflex and the distortion product otoacoustic emission (DPOAE) test, and their cochleae were analyzed by scanning electron microscopy. To the four groups investigated, group I (control) and group II (low dose, 10 mg/kg/day for 30 days) showed a normal DPOEA and normal outer hair cells; group III (high dose, 160 mg/kg/day for 10 days) showed the absence of DPOEA and damage to the outer hair cells; and group IV (low dose, 10 mg/kg/day for 30 days followed by a high dose of 160 mg/kg/day for 10 days) showed a normal DPOEA and normal outer hair cells. These results demonstrate that there was a considerable self-protection phenomenon by gentamicin.

Protection levels of vaccinated pigeons (Columba livia) against a highly pathogenic newcastle disease virus strain

The purposes of this study were to model a vaccination regimen for Newcastle disease virus (NDV) in pigeons, and to evaluate the susceptibility and behavior of vaccinated birds against a highly pathogenic NDV Brazilian strain. Antibody response was assessed by means of hemagglutination inhibition test (HI), and viral genome excretion by means of RT-PCR. Vaccinal strains (La Sota and Ulster) induced high antibody titers without any adverse effects, both in inoculated and in sentinel birds. A viral strain pathogenic for chickens did not produce clinical signs of the disease in experimentally infected pigeons. Only 4 out of 10 vaccinated pigeons shed NDV genome, and just for two days. Results confirmed the high infectivity of the vaccinal strains used, as all nonvaccinated pigeons showed antibody titers as high as those of vaccinated birds.

Interface between domestic and wild carnivores in an environmental protection area in the brazilian Amazon: indicators and aplications for conservation

The presence of domestic animals in protected areas has been a growing world concern, including in the Brazilian Amazon. Introduced domestic carnivores can put the conservation of carnivores and other wild mammals at risk in protected areas. These risks can be associated with direct factors, such as the dogs` hunting habit, and indirect factors, such as the domestic carnivores` potential for transmitting infectious agents to wild populations. The objective of this study was to analyze the potential implications of humans and domestic animals staying in fragmented and altered areas such as the Wildlife Protection Areas (WPA) in the Tucuruf Lake Environmental Protection Area (Para, Brazil), created for the full protection of its fauna and flora. This evaluation was made through interviews conducted with riparian inhabitants living in the WPA and surrounding area, involving issues related to the presence of dogs (number of animals per domicile, birth rate, mortality rate, vaccination status, hunting habit) and wild carnivores (occurrence and location) in the study area. The results indicated risks mainly due to the presence of on average three dogs per domicile, as well as the high canine birth rate, their hunting habits, low vaccination rates and evidence of direct and indirect contact with the human, canine and wild carnivore populations. These factors represent risks to the health and conservation of Tucurui EPA wild carnivores since they provide favorable conditions for the transmission of pathogens from domestic fauna to wild fauna, as well as the risks brought about by hunting.

Accuracy of working length determination using 3 electronic apex locators and direct digital radiography

Objectives. The objectives of this study were to assess the accuracy of working length determination using 3 electronic apex locators and direct digital radiography and to compare the results with those obtained using the visual method (control measurement). Study design. Twenty extracted human maxillary premolars were selected: 17 two-rooted and 3 single-rooted (total of 37 canals). Working length was measured using electronic apex locators Elements Diagnostic, Root ZX, and Just II. Subsequently, teeth were positioned in the alveolar bone of a dry skull and submitted to direct digital radiography. A variation of +/- 1 mm was considered as acceptable. Results were analyzed using the Wilcoxon and the chi(2) tests. Results. Results presented an accuracy of 94.6% for Elements Diagnostic, 91.9% for Root ZX, 73.0% for Just II, and 64.9% for direct digital radiography when considering the margin of +/- 1 mm in relation to the control measurement. Comparisons with the actual control measurements resulted in accuracy results of 13.51%, 13.51%, 10.10%, and 2.70%, respectively. Conclusions. Root ZX and Elements Diagnostic are more accurate in determining working length when compared with Just II and Schick direct digital radiography. (Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2011;111:e44-e49)