Percorrer por autor "Perdigao, Joao"
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- Dynamics and Development of Extensively Drug-resistant Tuberculosis, PortugalPublication . Perdigao, Joao; Silva, D.; Pereira, V.; Macedo, Rita; Silva, Carla; Machado, Diana; Couto, Isabel; Viveiros, Miguel; Jordão, Luísa; Portugal, IsabelThe development of multidrug-resistant (MDR) and extensive drug-resistant (XDR) tuberculosis(TB) combined with subsequent transmission constitutes a serious threat to the effective control of tuberculosis in several countries. Lisbon Health Region, despite great progresses in TB management still presents a high number of MDR/XDR-TB cases. The development of this type of resistance is the result of adaptative selection of Mycobacterium tuberculosis strains that acquire and accumulate specific mutations at specific genes. The presently known mechanisms of drug resistance include the modification or overexpression of drug targets, inactivation of drug- activator enzymes and overexpression of drug-modifying enzymes. Although the molecular basis of resistance of MDR/XDR-TB strains circulating in Lisbon has already been addressed in different studies, the dynamics or mode of resistance acquisition that have lead to the different circulating strains is still partially unclear. In the present study we have genotyped and screened a set of 44 MDR/XDR-TB isolates for mutations in tlyA, gyrA, rrs and eis genes. We have determined the most prevalent mutations found in each gene to be Ins755GT in tlyA, A1401G in rrs, G-10A in eis and S91P in gyrA. Two genetic clusters previously known to be associated with XDR-TB were detected, Lisboa3 and Q1, containing 27 and 17 isolates, respectively. Lisboa3 strains isolated in the 90’s with the same mutational profile of recent XDR-TB Lisboa3 strains were found, emphasizing the ancient XDR-TB problem in the region. Also investigated was the resistance level conferred by eis G-10A mutations, revealing that eis G-10A mutations may result in an undetectable AMK resistance. We concluded by analyzing the mutational distribution found by genetic cluster that in Q1 cluster two mutations, gyrA D94A and rrs A1401G, were enough to ensure development of XDR-TB from a multidrug resistant strain. Moreover, in Lisboa3 cluster it was possible to determine that the development of kanamycin low-level resistance mediated by eis promoter mutations was at the origin of independent emergence of several XDRTB strains that can be discriminated within Lisboa3 genetic cluster by tlyA mutations.
- Exploring a link between mycobacteria structure and virulencePublication . Silva, Carla; Perdigao, Joao; Alverca, Elsa; Matos, António Pedro Alves; Carvalho, Patrícia A; Portugal, Isabel; Jordão, LuísaTuberculosis (TB) is a major health problem. The emergence of multidrug resistant (MDR)Mycobacterium tuberculosis (Mtb) isolates confounds treatment strategies. In Portugal, cases of MDR-TB are reported annually with increased incidence noted in Lisbon. The majority of these MDR-TB cases are due to closely-related mycobacteria known collectively as Lisboa family and Q1 cluster. The genetic determinants linked to drug resistance have been exhaustively studied resulting in the identification of family and cluster specific mutations. Nevertheless, little is known about other factors involved in drug resistance development. Here we focused on the study of morphological and structural features of Mtb isolates, collected during 2008-2009 in Lisbon, in order to complement the genetic analysis. For this propose scanning and transmission electron microscopy techniques were used. Particular attention was given to Lisboa family and Q1 cluster isolates since together they account for the majority of reported MDR-TB cases. This analysis allowed the identification of structural differences, such as cell envelope thickness, between Mtb clinical isolates, which are correlated with antibiotic resistance. The infection of human monocyte derived macrophages allowed us to document the relative selective advantage of Lisboa family isolates over other circulating Mtb isolates.
- Exploring the contribution of mycobacteria characteristics in the interaction with human macrophagesPublication . Silva, Carla; Perdigao, Joao; Alverca, Elsa; de Matos, António Pedro Alves; Carvalho, Patrícia A; Portugal, Isabel; Jordão, LuísaTuberculosis (TB) is a major health problem. The emergence of multidrug resistant (MDR) Mycobacterium tuberculosis (Mtb) isolates confounds treatment strategies. In Portugal, cases of MDR-TB are reported annually with increased incidence noted in Lisbon. The majority of these MDR-TB cases are due to closely-related mycobacteria known collectively as Lisboa family and Q1 cluster. The genetic determinants linked to drug resistance have been exhaustively studied resulting in the identification of family and cluster specific mutations. Nevertheless little is known about other factors involved in drug resistance development. Here, for the first time, we complemented the genetic analysis with the study of morphological and structural features of Lisboa family and Q1 cluster isolates by using scanning and transmission electron microscopy. This analysis allowed the identification of structural differences, such as cell envelope thickness, between Mtb clinical isolates, which are correlated with antibiotic resistance. The infection of human monocyte derived macrophages allowed us to document the relative selective advantage of Lisboa family isolates over other circulating Mtb isolates.
- Molecular analysis on resistant TB isolates in PortugalPublication . Silva, Carla; Perdigao, Joao; Jordão, Luísa; Portugal, IsabelPortugal has one of the highest tuberculosis (TB) incidence (20/100 000 inhabitants) in Europe. The emergence of multidrug resistant (MDR) TB and extensive drug-resistant (XDR) TB infection is the biggest threat to TB control. Most strains of MDR-TB circulating in the Lisbon area belong to a particular family of genetically related strains, the Lisboa family, detected in the 90’s. The prevalence of this family of strains has increased over the years, and represented more than 85% of the MDR-TB strains in the year of 2008. XDR-TB has been recently derived from MDR-TB strains and account for about 50% of these, the majority belonging to Lisboa family. Lisboa family represents a serious problem regarding TB control in Portugal and its prevalance in recent years suggests that these strains may have selective advantages over others. In order to establish a link between the most prevalent mutations in drug-resistance associated genes and spread of Lisboa strains in the Portuguese setting, 54 resistant-TB clinical isolates in Portugal were study. The isolates were characterized by 24 loci MIRU-VNTR and analyzed for inhA, katG, rpoB, rpsL, rrs, embB and pncA genes, for resistance to first-line drugs. The MDR isolates (n=35) were further analyzed for mutations in gyrA, tlyA, eis and rrs for resistance to fluoroquinolones and second-line injectables. MIRU-VNTR analysis showed that Lisboa family strains and Q1 cluster were the most prevalent, with 26 (48%) and 6 (11%) isolates respectively, including the majority of the MDR-TB and XDR- TB isolates. No mutations in first-line drug resistance associated genes specifically related with MDR were found. However, mutation analysis to second-line drug resistance in 17 MDR-TB isolates shown that specific mutations arepresent in particular families. Therefore, XDR-TB from Lisboa family exhibits gyrA D94G/S91P, tlyA Ins755GT and eis G-10A mutations, and XDR-TB from Q1 presents gyrA D94A and rrs A1401G mutations. The remaining isolates are still under study, and further analyses are ongoing. We conclude that XDR-TB isolates from Lisboa family and Q1 cluster have shown a marked difference between them, regarding second-line mutations. Such analysis may be useful to define mutation profiles that distinguish Lisboa family from Q1 isolates.