Drug susceptibility of Brazilian strains of Mycobacterium bovis using traditional and molecular techniques

Transmission of Mycobacterium bovis from cattle to humans has been reported and can cause tuberculosis (Tb) and a problem in certain risk populations. Therefore, knowledge of resistance of M. bovis towards antibiotics used for therapy of human Tb could help avoiding cure delay and treatment cost increase when dealing with drug resistant organisms. We therefore evaluated the susceptibility of M. bovis isolates towards streptomycin, isoniazide, rifampicin, ethambutol, and ethionamide, the first line antibiotics for human Tb. Therefore, 185 clinical samples from cattle with clinical signs of tuberculosis were processed and submitted to culturing and bacterial isolates to identification and drug susceptibility testing using the proportion method. Among 89 mycobacterial strains, 65 were identified as M. bovis and none were resistant to any of the antibiotics used. Confirmation of present results by future studies, enrolling a large number of isolates and designed to properly represent Brazilian regions, may favor the idea of using isoniazide preventive therapy as part of a Tb control strategy in special situations. Also, nucleic acids from bacterial isolates were submitted to rifoligotyping, a recently described reverse hybridization assay for detection of mutations causing resistance towards rifampicin. Concordance between the conventional and the molecular test was 100%, demonstrating the use of such methodology for rapid evaluation of drug susceptibility in M. bovis.     

Southernmost finding of Lymnaea viatrix orbigny, 1835 (Pulmonata: Lymnaeidae), intermediate host of Fasciola hepatica (Linnaeus, 1758) (Trematoda: Digenea), in urban and rural areas of Patagonia, Argentina

We report the first finding of Lymnaea viatrix south of parallel 41 masculine S, in rural and urban areas from Argentina. Ninety snails were collected during year 2000, from a concrete pond at a Public Square in El Bolsón Village, Río Negro province, and 811 snails in November 1999, and during 2000 from waterbodies within a farm at Cholila locality, Chubut province. Fasciola hepatica infection was detected in 0.9% snails from the rural area. We discuss the potential risk of L. viatrix to public health in urban areas and its epidemiological importance in rural areas of the Andean Patagonian region.     

Modelling of ex vivo expansion/maintenance of hematopoietic stem cells

In this study, we described the modelling of the expansion/maintenance of human hematopoietic stem/progenitor cells from adult human bone marrow. CD 34(+)-enriched cell populations from bone marrow were cultured in the presence and absence of human stroma in serum-free media containing bFGF, SCF, LIF and Flt-3 ligand for several days. The cells in the culture were analysed for expansion and phenotype by flow cytometry. Although significant expansion of bone marrow cultures occurred in the presence and absence of human stroma, the results of expansion were effectively better in the presence of a stromal layer. In both situations the phenotypic analysis demonstrated a great expansion of CD 34(+)38(-) cells. The differentiative potential of bone marrow CD 34(+) cells co-cultured with human stroma was primarily shifted towards the myeloid lineage with the presence of CD 15 and CD 33.     

Assisted-selection of naturally caffeine-free coffee cultivars—characterization of SNPs from a methyltransferase gene

Breeding of caffeine-free coffee cultivars require tools for an early selection of progenies bearing this later trait. Genes from caffeine synthesis and degradation represent major targets for the development of molecular markers for assisted selection. In this study, we characterized SNPs identified on the caffeine synthase gene from AC1 mutant, a naturally caffeine-free arabica coffee plant. Molecular analysis of normal and mutant sequences indicates the occurrence of SNPs in protein domains, potentially associated with caffeine synthesis in coffee. Progenies F₂, F₁BC₁ and BC from crosses of AC mutants and elite cultivars were evaluated regarding caffeine content in grains and genomic segregation profile of selected SNPs. Genotyping analysis allowed the discrimination between homozygous and heterozygous plants. Quantification of caffeine content indicated a significant variability among progenies and a low frequency of caffeine-free plants. Statistical analyses of genotyping and phenotyping results showed significant association between presence of selected SNPs and reduced caffeine content. Moreover, this association occurs through all evaluated genetic backgrounds and generations, indicating an inheritance stability of both trait and markers. The molecular markers described here represent a successful case of assisted-selection in coffee, indicating their potential use for breeding of caffeine-free cultivars.     

Genetic and dermatoglyphic distances among Basque Valleys.

Palmar dermatoglyphics of a sample including 552 males and 701 females from 8 Basque valleys were analyzed. We studied the frequency of palmar pattern types and compared them using correspondence the frequency of palmar pattern types and compared them using correspondence analysis. The results of this comparative study show that there is diversity among valleys and also that this diversity depends on the trait and on sex. Genetic drift could explain this variability found in the Basque population.     

Diffusion-Weighted MRI in the Assessment of Early Treatment Response in Patients with Squamous-Cell Carcinoma of the Head and Neck: Comparison with Morphological and PET/CT Findings

Objective

To evaluate changes in apparent diffusion coefficients (ADC) as measured by magnetic resonance imaging (MRI) before and after the treatment of primary tumors and cervical metastases in patients with squamous-cell carcinoma (SCC) of the head and neck, and to compare these values to the results of widely used morphological criteria and [18F]-FDG PET/CT findings.

Material and Method

This was a longitudinal, prospective, single-center nonrandomized trial involving patients with head and neck SCC treated with chemotherapy alone or in combination with radiotherapy. Imaging examinations ([18F]-FDG PET/CT and diffusion-weighted MRI) were performed on the same day, up to one day prior to the beginning of the first treatment cycle, and on the 14th day of the first chemotherapy cycle. Treatment response was evaluated based on the Response Evaluation Criteria in Solid Tumors (RECIST) and World Health Organization (WHO) morphological criteria, as well as PET Response Criteria in Solid Tumors (PERCIST) metabolic criteria.

Results

Seventy-five lesions were examined in 23 patients. Pre- and post-treatment comparisons of data pertaining to all target lesions revealed reductions in tumor size and SUV, as well as increases in ADC values, all of which were statistically significant. The increase in ADC following treatment was significantly higher in patients classified as complete responders by both morphological criteria than that observed in any of the other patient groups of response. Patients with a complete metabolic response also showed greater increases in ADC values as compared to the remaining groups.

Conclusion

The assessment of tumor response based on diffusion-weighted MRI showed an increase in the ADC of cervical lesions following treatment, which was corroborated by morphological and metabolic findings. Associations between changes in ADC values and treatment response categories using morphologic criteria and [18F]-FDG PET/CT were only identified in complete responders.     

A rheumatoid factor paradox: inhibition of rituximab effector function

Introduction

Rituximab (RTX) therapy of rheumatoid arthritis (RA) exhibits enhanced effectiveness in seropositive patients. Using patient sera, we tested if this improved efficacy was associated with enhanced RTX mediated complement-dependent cytotoxicity (RTX-CDC).

Methods

We developed an in vitro assay for RTX-CDC using patient sera and the Daudi human B cell line. Using propidium iodide uptake and flow cytometry, we compared RTX-CDC with rheumatoid factor (RF)+ sera relative to normal volunteer, non-RA and RF- sera. Additional studies examined mixing studies of RF+ and RF- sera, as well as the effect of monoclonal IgA or IgM RF. Finally, the effect of RF on RTX mediated trogocytosis of normal B cells was evaluated.

Results

Using human sera, addition of RTX resulted in rapid and profound (> 50%) Daudi cell death that was complement dependent. Surprisingly, RF+ patient sera exhibited reduced RTX-CDC relative to RF- sera, with an inverse relationship of RTX-CDC and RF titer. Mixing studies indicated the presence of an inhibitor of RTX-CDC in RF+ sera. The addition of monoclonal IgM or IgA RF to RF- sera markedly inhibited RTX-CDC. This effect was specific for RF binding to the Fc portion of RTX as it was not apparent with the F(ab)'' domains of RTX engineered onto IgG3 heavy chain. RF also modestly inhibited RTX mediated trogocytosis.

Conclusions

Contrary to expectations, RF+ sera exhibits reduced RTX-CDC due to the presence of RF. The enhanced efficacy of RTX in seropositive RA patients cannot be attributed to improved B cell depletion through CDC. This result indicates that high RF levels may potentially modulate the efficacy of any therapeutic monoclonal antibody dependent on Fc effector function.     

Therapeutic cold: An effective kind to modulate the oxidative damage resulting of a skeletal muscle contusion

Muscular contusions affect the function of the skeletal muscle system. This study investigated the oxidative damage as well as the main morphological changes related to a skeletal muscle contusion in the gastrocnemius muscle of rats and also the capacity of therapeutic cold to modulate these parameters. The therapeutic cold modulated the increase of oxidative stress markers and also modulated the reduction in the antioxidants levels in the injured muscle. In enzyme assays, therapeutic cold was also effective in normalizing the muscle Na(+)/K(+) and Ca(2+) ATPases, lactate dehydrogenase and myeloperoxidase activities. Similarly, the lesioned non-treated animals presented evident impairments in the mitochondrial functions and in the muscle morphology which were diminished by the cold treatment. The therapeutic cold was able to modulate the oxidative damage possibly by its capacity to limit the inflammatory response intensity, to attenuate the impairment of the mitochondrial function and also to preserve the skeletal muscle morphology.     

European Ancestry Predominates in Neuromyelitis Optica and Multiple Sclerosis Patients from Brazil

Background

Neuromyelitis optica (NMO) is considered relatively more common in non-Whites, whereas multiple sclerosis (MS) presents a high prevalence rate, particularly in Whites from Western countries populations. However, no study has used ancestry informative markers (AIMs) to estimate the genetic ancestry contribution to NMO patients.

Methods

Twelve AIMs were selected based on the large allele frequency differences among European, African, and Amerindian populations, in order to investigate the genetic contribution of each ancestral group in 236 patients with MS and NMO, diagnosed using the McDonald and Wingerchuck criteria, respectively. All 128 MS patients were recruited at the Faculty of Medicine of Ribeirão Preto (MS-RP), Southeastern Brazil, as well as 108 healthy bone marrow donors considered as healthy controls. A total of 108 NMO patients were recruited from five Neurology centers from different Brazilian regions, including Ribeirão Preto (NMO-RP).

Principal Findings

European ancestry contribution was higher in MS-RP than in NMO-RP (78.5% vs. 68.7%) patients. In contrast, African ancestry estimates were higher in NMO-RP than in MS-RP (20.5% vs. 12.5%) patients. Moreover, principal component analyses showed that groups of NMO patients from different Brazilian regions were clustered close to the European ancestral population.

Conclusions

Our findings demonstrate that European genetic contribution predominates in NMO and MS patients from Brazil.