The enzymatic properties of Arabidopsis thaliana DNA polymerase λ suggest a role in base excision repair.

Base excision repair (BER) generates gapped DNA intermediates containing a 5'-terminal 2-deoxyribose-5-phosphate (5'-dRP) group. In mammalian cells, gap filling and dRP removal are catalyzed by Pol ß, which belongs to the X family of DNA polymerases. In higher plants, the only member of the X family of DNA polymerases is Pol λ. Although it is generally believed that plant Pol λ participates in BER, there is limited experimental evidence for this hypothesis. Here we have characterized the biochemical properties of Arabidopsis thaliana Pol λ (AtPol λ) in a BER context, using a variety of DNA repair intermediates. We have found that AtPol λ performs gap filling inserting the correct nucleotide, and that the rate of nucleotide incorporation is higher in substrates containing a C in the template strand. Gap filling catalyzed by AtPol λ is most efficient with a phosphate at the 5'-end of the gap and is not inhibited by the presence of a 5'-dRP mimic. We also show that AtPol λ possesses an intrinsic dRP lyase activity that is reduced by mutations at two lysine residues in its 8-kDa domain, one of which is present in Pol λ exclusively and not in any Pol ß homolog. Importantly, we also found that the dRP lyase activity of AtPol λ allows efficient completion of uracil repair in a reconstituted short-patch BER reaction. These results suggest that AtPol λ plays an important role in plant BER.

Ready-to-eat street food: a potential source for dissemination of multidrug-resistant Escherichia coli epidemic clones in Quito, Ecuador.

Ready-to-eat food contamination with ESBL-producing Escherichia coli is a growing health concern. Some of these strains also are epidemic clones and can cause community-associated infections that are difficult to treat. In this study, the occurrence of ESBL-producing E. coli contaminated ready-to-eat street food in Quito, Ecuador was evaluated. In total, 150 samples were collected randomly in the most crowded sites of the city. In all, 34 samples (34/150; 22·6%) were positive for total thermotolerant (44·5°C) coliforms resistant to cefotaxime. MALDI-TOF analysis identified that the E. coli was found in 20 food samples (20/34; 59%). ESBL gene blaCTX-M-55 was identified in nine isolates, blaCTX-M-15 in six isolates, blaCTX-M-14 in two isolates, and one isolate each harboured blaCTX-M-24 , blaCTX-M-65 , blaCTX-M-55 and blaCTX-M-8 . Phylogenetic groups like A and B1 were the most common, followed by groups D and B2. MLST analysis identified 12 different sequence types (STs), the most common was ST162. Recognized epidemic clonal groups ST410, ST131 and ST744 were encountered. Ready-to-eat street food is a potential way of spreading ESBL-producing E. coli epidemic clones in Quito, Ecuador. SIGNIFICANCE AND IMPACT OF THE STUDY: This study identified ESBL-producing Escherichia coli epidemic clones: ST131, ST410 and ST744 in ready-to-eat street food samples. Street food is a possible way to spread harm multidrug-resistant (MDR) E. coli strains in the community. Studies to identify the contamination sources of this kind of food are needed to tackle MDR E. coli dissemination.

Genetic diversity and drug resistance of Mycobacterium tuberculosis in Ecuador.

BACKGROUND: The genetic diversity of Mycobacterium tuberculosis in Quito, Ecuador is not well known. OBJECTIVE: To investigate mutations related to drug resistance and bacterial genotypes in M. tuberculosis strains in Ecuador. DESIGN: This was a retrospective study of M. tuberculosis isolates from 104 patients. Isolates were phenotypically resistant to rifampicin (RMP) and/or isoniazid (INH). The genotype was determined using 24-locus mycobacterial interspersed repetitive units-variable-number tandem repeats (MIRU-VNTR). RESULTS: Isolates showed mutations in the rpoB and katG genes, and the inhA promoter. In rpoB, we found 13 genetic alterations at codons 511, 513, 514, 515, 516, 526 and 531. Forty-six (44.2%) RMP-resistant isolates belonged to codon 531. In katG, there were nine genetic alterations at codons 296, 312, 314, 315, 322, 324 and 351. Fifty-three (51%) INH-resistant isolates belonged to codon 315. Five mutations not previously described were identified in katG: Thr324Ser, Thr314Ala, Ala312Pro, Trp351Stop and deleted G at 296 codon. The Latin American Mediterranean (LAM) (33.7%) and Ghana (30.8%) lineages presented most of the main mutations observed. CONCLUSION: This is the first report from Ecuador; it describes five new mutations in katG and indicates that LAM is the most prevalent lineage.

Colistin-resistant Escherichia coli clinical isolate harbouring the mcr-1 gene in Ecuador.

Colistin resistance mediated by the mcr-1 gene has been reported worldwide, but to date not from the Andean region, South America. We report the first clinical isolate of Escherichia coli harbouring the mcr-1 gene in Ecuador. The strain was isolated from peritoneal fluid from a 14-year-old male with acute appendicitis, and subjected to molecular analysis. The minimum inhibitory concentration of colistin for the strain was 8 mg/ml and it was susceptible to carbapenems but resistant to tigecycline. The strain harboured mcr-1 and bla CTX-M-55 genes and was of sequence type 609. The recognition of an apparently commensal strain of E. coli harbouring mcr-1 serves as an alert to the presence in the region of this recently described resistance mechanism to one of the last line of drugs available for the treatment of multi-resistant Gram-negative infections.

Characterization of the first report of Mycobacterium timonense infecting an HIV patient in an Ecuadorian hospital.

Mycobacterium timonense is a non-tuberculous mycobacteria (NTM) described in southern France in 2009, and to our knowledge, not reported again as a human pathogen in indexed literature. The aim of this work was to characterize the first clinical isolate of M. timonense in Ecuador. Time of growth, biochemical tests, thin layer growth test, PCR-RFLP analysis of the hsp65 gene and MALDI-TOF spectra analysis were not able to identify the species. The species identification was achieved through sequencing of rrs, hsp65 and rpoB genes. The results highlight the necessity to set up a sequencing method to identify emerging NTM in Ecuadorian clinical facilities.