Biomedical subjects

Jorge Machado

Publications and source records attributed to Jorge Machado.

9 recordsLinked to original sources

PubMed2006

Fourier transform infrared (FT-IR) spectroscopy in bacteriology: towards a reference method for bacteria discrimination.

Rapid and reliable discrimination among clinically relevant pathogenic organisms is a crucial task in microbiology. Microorganism resistance to antimicrobial agents increases prevalence of infections. The possibility of Fourier transform infrared (FT-IR) spectroscopy to assess the overall molecular composition of microbial cells in a non-destructive manner is reflected in the specific spectral fingerprints highly typical for different microorganisms. With the objective of using FT-IR spectroscopy for discrimination between diverse microbial species and strains on a routine basis, a wide range of chemometrics techniques need to be applied. Still a major issue in using FT-IR for successful bacteria characterization is the method for spectra pre-processing. We analyzed different spectra pre-processing methods and their impact on the reduction of spectral variability and on the increase of robustness of chemometrics models. Different types of the Enterococcus faecium bacterial strain were classified according to chromosomal DNA restriction patterns produced by pulsed-field gel electrophoresis (PFGE). Samples were collected from human patients. Collected FT-IR spectra were used to verify if the same classification was obtained. In order to further optimize bacteria classification we investigated whether a selected combination of the most discriminative spectral regions could improve results. Two different variable selection methods (genetic algorithms (GAs) and bootstrapping) were investigated and their relative merit for bacteria classification is reported by comparing with results obtained using the entire spectra. Discriminant partial least-squares (Di-PLS) models based on corrected spectra showed improved predictive ability up to 40% when compared to equivalent models using the entire spectral range. The uncertainty in estimating scores was reduced by about 50% when compared to models with all wavelengths. Spectral ranges with relevant chemical information for Enterococcus faecium bacteria discrimination were outlined.

Bacterial Typing Techniques↗

PubMed2006

Characterization of antimicrobial resistance and class 1 and 2 integrons in Salmonella enterica isolates from different sources in Portugal.

OBJECTIVES: The antimicrobial resistance profiles of 1183 Salmonella isolates collected during 2002-2003 from several sources (human, food products and environment) were evaluated. The occurrence, distribution and cassette content of class 1 and 2 integrons among the sulphonamide-resistant population, as well as the role of particular clones to the spread of these genetic elements, were investigated. METHODS: The isolates were examined for susceptibility to antimicrobial agents. The characterization of class 1 and 2 integrons was investigated using PCR, PCR-RFLP (restriction fragment length polymorphism) and sequencing in the sulphonamide-resistant isolates. Conjugation assays and clonality analysis by PFGE were performed. RESULTS: The most common resistance phenotypes were to nalidixic acid, tetracycline, streptomycin, sulfamethoxazole and ampicillin (ranging from 31% to 17%). Resistance to sulphonamides (n=200) was associated with resistance to other antimicrobial agents, with 75% of the isolates carrying one or two class 1 integrons while only 3% simultaneously carried class 1 and 2 integrons. Integrons were observed among at least 11 serotypes (mainly Typhimurium) and in a reduced number of PFGE clones (20). Eight class 1 integron types were found, with the aadA genes (aadA1, aadA2 and aadA5) alone or downstream of a trimethoprim (dfrA1, dfrA12 and dfrA17) or a beta-lactamase resistance gene (blaoxa-30) and the blaPSE-1 gene alone. Most of the class 1 integron types were shared by several clones from the same or different serotypes obtained either from humans or food products of animal origin, especially pork products. However, some Typhimurium-specific integrons were found: aadA2 plus blaPSE-1 and blaoxa-30-aadA1. CONCLUSIONS: Apart from the hypothetical contribution of the conjugative transfer of integrons, the incidence of Salmonella carrying these genetic units seems to rely on the ability of certain clones to spread or persist in particular animal niches. Our data suggest that food-producing animals might be simultaneously considered as a reservoir of clones and integrons carrying antibiotic resistance genes, thus making the food chain, especially pork products, a possible source of multidrug-resistant isolates in humans.

Anti-Bacterial Agents↗

PubMed2005

Dissemination of sulfonamide resistance genes (sul1, sul2, and sul3) in Portuguese Salmonella enterica strains and relation with integrons.

In 200 sulfonamide-resistant Portuguese Salmonella isolates, 152 sul1, 74 sul2, and 14 sul3 genes were detected. Class 1 integrons were always associated with sul genes, including sul3 alone in some isolates. The sul3 gene has been identified in isolates from different sources and serotypes, which also carried a class 1 integron with aadA and dfrA gene cassettes.

Anti-Bacterial Agents↗

PubMed2004

Dissemination amongst humans and food products of animal origin of a Salmonella typhimurium clone expressing an integron-borne OXA-30 beta-lactamase.

OBJECTIVES: Characterization of the molecular basis for beta-lactam resistance and evaluation of the clonal relatedness among nine isolates of multidrug-resistant Salmonella typhimurium recovered from seven clinical human samples and two pork end products. METHODS: The isolates were examined for susceptibility to antimicrobial agents. The relationships between resistance genes, class 1 integrons, plasmids and isolates were screened by molecular methods such as polymerase chain reaction and restriction fragment length analysis. RESULTS: A bla(OXA-30) gene, located in a class 1 integron, was detected in all isolates. This integron was present on a conjugative plasmid in all but one isolate. By pulsed-field gel electrophoresis, it was determined that all strains share the same chromosomal type. CONCLUSIONS: This study demonstrates the spread of an OXA-30-producing S. typhimurium in Portugal, suggesting dissemination of a resistant clone through the food chain.

Animals↗

PubMed2004

Paracellular pathway in the shell epithelium of Anodonta cygnea.

Ultrastructural study of cell-cell connections in the outer mantle epithelium (OME) on high-pressure-frozen specimens revealed zonula adherens, septate junctions and gap junctions in Anodonta cygnea. In order to evaluate the permeability of the paracellular pathway, the OME was incubated under gradients of lanthanum and calcium. After lanthanum incubation (4 mM) from the basal side, the septate junctions were penetrated completely by this tracer. When applied from the apical side, lanthanum deposits were located similarly over the entire length of the septate junctions up to the first dilatations of the intercellular space. Calcium deposits were also present in paracellular areas only when OME had been incubated simultaneously with calcium (6 mM) and lanthanum (4 mM) gradients. Lanthanum and calcium deposits were detected with ESI (Electron Spectroscopic Imaging) and identified with EELS (Electron Energy Loss Spectroscopy). On the other hand, electrophysiological observations showed a 48% reduction of conductance when the OME was bathed on both sides with solutions containing lanthanum (4 mM) and calcium (6 mM), compared to bathing with lanthanum-free solution (control). The conductance reduction was 52% when calcium was removed from the control solution. Supported by morphological and physiological evidence, it appears that, under in vivo conditions, calcium ions may diffuse paracellularly from the haemolymph towards the extrapallial fluid and vice-versa across the septate junctions in the OME of A. cygnea. Permeability of the septate junctions depended proportionally on the calcium concentration in fluids.

Adherens Junctions↗

PubMed2003

Effects of controlled pH on organic and inorganic composition in haemolymph, epidermal tissue and cuticle of mud crab Scylla serrata.

Analysis of organic and inorganic compounds in plasma, epidermal tissue and cuticle were accomplished in the intermolt (C3 stage) of crab Scylla serrata incubated in different pH media. Significant changes with similar trends for protein, carbohydrates, glycosaminoglycans (GAG), sulphur, calcium, magnesium, potassium, phosphorus and copper in the plasma suggested higher dissolution in an acidic medium while the deposition increased in alkaline medium. Similar decreases in protein, carbohydrate and GAG in the epidermal compartment were observed from pH 4 to pH 12. However, significantly higher contents of sodium, chloride, potassium, phosphorus, magnesium, sulphur and copper were measured at pH 7.5 with a symmetrical decrease profile in both acidic and alkaline media, resulting from synergistic effects in the osmotic regulation. Clear changes in calcium concentrations were observed with a sharp increase from lower contents at pH 7.5 to higher at pH 12. In the cuticle, the acidic condition induced a significant dissolution of HCl-protein, GAG, calcium and magnesium contents. On the other hand, the alkaline condition induced a significant decrease in carbohydrate, calcium, chloride, sulphur and potassium. A reduction trend is seen for NaOH and H(2)O-protein contents in the cuticle. These observations suggest that GAG and HCl-protein might constitute the most soluble fraction with high affinity for calcium binding and easily removed in acidic conditions. Additionally, it is possible to speculate that the carbohydrates associated with the NaOH and H(2)O-proteins may form an interface between the soluble matrix fraction and the chitin framework. Sulphur groups seem to present a strong linkage role in this interface fraction, maybe only broken by a specific enzyme in extreme alkaline conditions with subsequent release of significant calcium from the shell.

Animals↗

PubMed2002

Cuticle Ultrastructure Changes in the Crab Scylla serrata Over the Molt Cycle.

Morphological and chemical studies on the cuticle during the molt cycle of the crab Scylla serrata were performed in order to understand the layer formation. Cuticle ultrastructure was studied by scanning electron microscopy (SEM). Energy-dispersive, X-ray diffraction, and X-ray fluorescence analysis were used for identification of the elements and phases in the inner surface of the cuticle. In the first stage (A) of cuticle formation, a thin pellicle organized as an irregular fragmented structure is built. It is composed mainly of alpha-chitin/protein beta-keratin-like complexes where heterogeneous mineral nucleation occur. It is impregnated by ferric concretions, responsible for the brown colour of the carapace. At the beginning of the mineralization process, a spheroidal inorganic phase appears consisting of dicalcium phosphate dihydrate (DCPD) Ca/P=1.00, octacalcium phosphate (OCP) Ca/P=1.33 associated with hydromagnesite and bromapatite traces. During further cuticle development in the remaining A stage and in the beginning of the B stage, calcite and magnesian calcite are formed from the precursor calcium phosphate phase. The next development in the C stages is characterized by intense calcareous thickening consisting mainly of calcite and of magnesian calcite, which become the major mineral fraction of the cuticle. Organic-inorganic complex precipitations exhibit different aspects as spongiform, filamentary helicoidal, and concentric radial arrangements during C1, C2, and C3, respectively. During different stages of the cuticle formation in Scylla serrata, these mineral deposits may partially result from the balance among different organic contents, mainly between alpha-chitin and protein beta-keratin-like compounds. On the other hand, the calcium crystallization on apatite and calcite polymorphic structures may be influenced by variations of physico-chemical factors in the cuticle compartment. J. Exp. Zool. 293:414-426, 2002.

Animals↗

PubMed2002

Organic and inorganic compound variations in haemolymph, epidermal tissue and cuticle over the molt cycle in Scylla serrata (Decapoda).

Analysis of organic and inorganic compounds in haemolymph, epidermal tissue and cuticle, allowed the correlation of their content variation during the molt cycle in Scylla serrata (Decapoda). The extrusion of carbohydrate, chitin, H(2)O- and NaOH-soluble proteins of the epidermal tissue from D2 stage on suggested an early formation of the organic fraction in the new cuticle. The additional secretion of HCl-soluble protein, glycosaminoglycans, calcium, phosphorus and sulfur in D2-D3 stages suggests mineral nucleation shortly before or at ecdysis. This is consistent with the dominant content of proteins and chitin associated with peaks of carbohydrates and GAG-sulfur groups detected in the cuticle immediately after ecdysis. Furthermore, the maximal phosphorus content at this moment suggests calcium, magnesium and manganese phosphate deposits. After an intense accumulation of calcium, chitin and sharp decrease of protein from A to B stages, a constant rate of their deposition in intermolt and dissolution in premolt occurred. The variations concerning proteins, carbohydrates, glycosaminoglycans, calcium, magnesium and sulfur in the haemolymph, before and after ecdysis, suggest a transitory phenomenon for calcium binding and releasing. Other major elements such as, sodium, potassium and chloride may function as osmotic regulators in the haemolymph and in epidermal compartments. The copper profile presented an indicator role for variations of osmolality over the molt cycle.

Animals↗

PubMed2002

Anatomic study of the filum terminale and its correlations with the tethered cord syndrome.

OBJECTIVE: To evaluate the intradural anatomic features of the filum terminale (FT) in fresh human cadavers, analyzing morphological parameters relevant for the diagnosis of the tethered cord syndrome. METHODS: Forty-one fresh cadavers were dissected, and the following parameters were evaluated: cadaver height, weight, and age, FT length, FT diameters at the initial point and midpoint, and topographic relationships of the initial and fusion points of the FT to the adjacent vertebrae. RESULTS: The mean FT length was 156.44 mm (range, 112.8-211.1 mm), the mean initial diameter was 1.38 mm (range, 0.4-2.5 mm), and the mean midpoint diameter was 0.76 mm (range, 0.1-1.55 mm). Four specimens (9.76%) exhibited FT thicknesses of more than 2 mm at their initial points. The FT most frequently started at the middle L1 level (19.51%) and fused with the dura mater at the upper S2 level (31.71%). Two fila (4.88%) started below the L2 level. Statistically significant correlations were observed (Pearson correlation, P < 0.05) between specimen weight and height (P = 0.019), initial point and midpoint diameters of the FT (P < 0.001), initial vertebral level and specimen height (P = 0.012), and initial and fusion vertebral levels (P = 0.004). CONCLUSION: Variation ranges were large for almost all anatomic parameters of the FT. Six cadavers (14.63%) in our study fulfilled one of the anatomic diagnostic criteria for tethered cord syndrome. Neurosurgeons must remember the normal variations in the anatomic features of this region when establishing the diagnosis of tethered cord syndrome or performing surgical procedures involving the caudal part of the dural sac.

Adult↗