Browsing by Author "Vale, Filipa F."
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- Antimicrobial activity of prophage endolysins against critical Enterobacteriaceae antibiotic-resistant bacteriaPublication . Gonçalves, Tiago; Marques, Andreia T.; Manageiro, Vera; Tanoeiro, Luis; Vital, Joana S.; Duarte, Aida; Vítor, Jorge M.B.; Caniça, Manuela; Gaspar, Maria Manuela; Vale, Filipa F.Enterobacteriaceae species are part of the 2017 World Health Organization antibiotic-resistant priority pathogens list for development of novel medicines. Multidrug-resistant Klebsiella pneumoniae is an increasing threat to public health and has become a relevant human pathogen involved in life-threatening infections. Phage therapy involves the use of phages or their lytic endolysins as bioagents for the treatment of bacterial infectious diseases. Gram-negative bacteria have an outer membrane, making difficult the access of endolysins to the peptidoglycan. Here, three endolysins from prophages infecting three distinct Enterobacterales species, Kp2948-Lys from K. pneumoniae, Ps3418-Lys from Providencia stuartii, and Kaer26608-Lys from Klebsiella aerogenes, were purified and exhibited antibacterial activity against their specific bacterium species verified by zymogram assays. These three endolysins were successfully associated to liposomes composed of dimyristoyl phosphatidyl choline (DMPC), dioleoyl phosphatidyl ethanolamine (DOPE) and cholesteryl hemisuccinate (CHEMS) at a molar ratio (4:4:2), with an encapsulation efficiency ranging from 24 to 27%. Endolysins encapsulated in liposomes resulted in higher antibacterial activity compared to the respective endolysin in the free form, suggesting that the liposome-mediated delivery system enhances fusion with outer membrane and delivery of endolysins to the target peptidoglycan. Obtained results suggest that Kp2948-Lys appears to be specific for K. pneumoniae, while Ps3418-Lys and Kaer26608-Lys appear to have a broader antibacterial spectrum. Endolysins incorporated in liposomes constitute a promising weapon, applicable in the several dimensions (human, animals and environment) of the One Health approach, against multidrug-resistant Enterobacteriaceae.
- Cryptic Prophages Contribution for Campylobacter jejuni and Campylobacter coli IntrogressionPublication . Tanoeiro, Luís; Oleastro, Mónica; Nunes, Alexandra; Marques, Andreia T.; Duarte, Sílvia Vaz; Gomes, João Paulo; Matos, António Pedro Alves; Vítor, Jorge M.B.; Vale, Filipa F.Campylobacter coli and C. jejuni, the causing agents of campylobacteriosis, are described to be undergoing introgression events, i.e., the transference of genetic material between different species, with some isolates sharing almost a quarter of its genome. The participation of phages in introgression events and consequent impact on host ecology and evolution remain elusive. Three distinct prophages, named C. jejuni integrated elements 1, 2, and 4 (CJIE1, CJIE2, and CJIE4), are described in C. jejuni. Here, we identified two unreported prophages, Campylobacter coli integrated elements 1 and 2 (CCIE1 and CCIE2 prophages), which are C. coli homologues of CJIE1 and CJIE2, respectively. No induction was achieved for both prophages. Conversely, induction assays on CJIE1 and CJIE2 point towards the inducibility of these prophages. CCIE2-, CJIE1-, and CJIE4-like prophages were identified in a Campylobacter spp. population of 840 genomes, and phylogenetic analysis revealed clustering in three major groups: CJIE1-CCIE1, CJIE2-CCIE2, and CJIE4, clearly segregating prophages from C. jejuni and C. coli, but not from human- and nonhuman-derived isolates, corroborating the flowing between animals and humans in the agricultural context. Punctual bacteriophage host-jumps were observed in the context of C. jejuni and C. coli, and although random chance cannot be fully discarded, these observations seem to implicate prophages in evolutionary introgression events that are modulating the hybridization of C. jejuni and C. coli species.
- Genome Sequencing of 10 Helicobacter pylori Pediatric Strains from Patients with Nonulcer Dyspepsia and Peptic Ulcer DiseasePublication . Nunes, Alexandra; Rocha, Raquel; Vale, Filipa F.; Vieira, Luís; Sampaio, Daniel A.; Dias, Ricardo; Gomes, João Paulo; Oleastro, MónicaWe present draft genome sequences of 10 Helicobacter pylori clinical strains isolated from children. This will be important for future studies of comparative genomics in order to better understand the virulence determinants underlying peptic ulcer disease.
- Genomic structure and insertion sites of Helicobacter pylori prophages from various geographical originsPublication . Vale, Filipa F.; Nunes, Alexandra; Oleastro, Mónica; Gomes, João P.; Sampaio, Daniel A.; Rocha, Raquel; Vítor, Jorge M. B.; Engstrand, Lars; Pascoe, Ben; Berthenet, Elvire; Sheppard, Samuel K.; Hitchings, Matthew D.; Mégraud, Francis; Vadivelu, Jamuna; Lehours, PhilippeHelicobacter pylori genetic diversity is known to be influenced by mobile genomic elements. Here we focused on prophages, the least characterized mobile elements of H. pylori. We present the full genomic sequences, insertion sites and phylogenetic analysis of 28 prophages found in H. pylori isolates from patients of distinct disease types, ranging from gastritis to gastric cancer, and geographic origins, covering most continents. The genome sizes of these prophages range from 22.6-33.0 Kbp, consisting of 27-39 open reading frames. A 36.6% GC was found in prophages in contrast to 39% in H. pylori genome. Remarkably a conserved integration site was found in over 50% of the cases. Nearly 40% of the prophages harbored insertion sequences (IS) previously described in H. pylori. Tandem repeats were frequently found in the intergenic region between the prophage at the 3' end and the bacterial gene. Furthermore, prophage genomes present a robust phylogeographic pattern, revealing four distinct clusters: one African, one Asian and two European prophage populations. Evidence of recombination was detected within the genome of some prophages, resulting in genome mosaics composed by different populations, which may yield additional H. pylori phenotypes.
- Population genetic structure of Helicobacter pylori strains from Portuguese-speaking countriesPublication . Oleastro, Mónica; Rocha, Raquel; Vale, Filipa F.The human gastric colonizer Helicobacter pylori is useful to track human migrations given the agreement between the bacterium phylogeographic distribution and human migrations. As Portugal was an African and Brazilian colonizer for over 400 years, we hypothesized that Portuguese isolates were likely genetically closer with those from countries colonized by Portuguese in the past. We aimed to characterize the population structure of several Portuguese-speaking countries, including Portugal, Brazil, Angola, and Cape Verde.
- Repeated out-of-Africa expansions of Helicobacter pylori driven by replacement of deleterious mutationsPublication . Thorpe, Harry A.; Tourrette, Elise; Yahara, Koji; Vale, Filipa F.; Liu, Siqi; Oleastro, Mónica; Alarcon, Teresa; Perets, Tsachi-Tsadok; Latifi-Navid, Saeid; Yamaoka, Yoshio; Martinez-Gonzalez, Beatriz; Karayiannis, Ioannis; Karamitros, Timokratis; Sgouras, Dionyssios N.; Elamin, Wael; Pascoe, Ben; Sheppard, Samuel K.; Ronkainen, Jukka; Aro, Pertti; Engstrand, Lars; Agreus, Lars; Suerbaum, Sebastian; Thorell, Kaisa; Falush, DanielHelicobacter pylori lives in the human stomach and has a population structure resembling that of its host. However, H. pylori fromEurope and the Middle East trace substantially more ancestry from modern African populations than the humans that carry them. Here, we use a collection of Afro-Eurasian H. pylori genomes to show that this African ancestry is due to at least three distinct admixture events. H. pylori from East Asia, which have undergone little admixture, have accumulated many more non-synonymous mutations than African strains. European and Middle Eastern bacteria have elevated African ancestry at the sites of these mutations, implying selection to remove them during admixture. Simulations show that population fitness can be restored after bottlenecks bymigration and subsequent admixture of small numbers of bacteria from non-bottlenecked populations. We conclude that recent spread of African DNA has been driven by deleterious mutations accumulated during the original out-of-Africa bottleneck.
- Trends in Helicobacter pylori resistance to clarithromycin: from phenotypic to genomic approachesPublication . Marques, Andreia T.; Vítor, Jorge M.B.; Santos, Andrea; Oleastro, Mónica; Vale, Filipa F.For a long time Helicobacter pylori infections have been treated using the macrolide antibiotic, clarithromycin. Clarithromycin resistance is increasing worldwide and is the most common cause of H. pylori treatment failure. Here we review the mechanisms of antibiotic resistance to clarithromycin, detailing the individual and combinations of point mutations found in the 23S rRNA gene associated with resistance. Additionally, we consider the methods used to detect clarithromycin resistance, emphasizing the use of high-throughput next-generation sequencing methods, which were applied to 17 newly sequenced pairs of H. pylori strains isolated from the antrum and corpus of a recent colonized paediatric population. This set of isolates was composed of six pairs of resistant strains whose phenotype was associated with two point mutations found in the 23S rRNA gene: A2142C and A2143G. Other point mutations were found simultaneously in the same gene, but, according to our results, it is unlikely that they contribute to resistance. Further, among susceptible isolates, genomic variations compatible with mutations previously associated with clarithromycin resistance were detected. Exposure to clarithromycin may select low-frequency variants, resulting in a progressive increase in the resistance rate due to selection pressure.