Browsing by Author "Sousa, M."
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- Acquired antibiotic resistance among wild animals: the case of Iberian Lynx (Lynx pardinus)Publication . Sousa, M.; Gonçalves, A.; Silva, N.; Serra, R.; Alcaide, E.; Zorrilla, I.; Torres, C.; Caniça, Manuela; IgrejaS, G.; Poeta, P.The selective pressure generated by the clinical misuse of antibiotics has been the major driving force leading to the emergence of antibiotic resistance among bacteria. Antibiotics or even resistant bacteria are released into the environment and contaminate the surrounding areas. Human and animal populations in contact with these sources are able to become reservoirs of these resistant organisms. Then, due to the convergence between habitats, the contact of wild animals with other animals, humans, or human sources is now more common and this leads to an increase in the exchange of resistance determinants between their microbiota. Indeed, it seems that wildlife populations living in closer proximity to humans have higher levels of antibiotic resistance. Now, the Iberian Lynx (Lynx pardinus) is a part of this issue, being suggested as natural reservoir of acquired resistant bacteria. The emerging public health concern regarding microbial resistance to antibiotics is becoming true: the bacteria are evolving and are now affecting unintentional hosts.
- Contribution of casein kinase 2 and spleen tyrosine kinase to CFTR trafficking and protein kinase A-induced activityPublication . Luz, S.; Kongsuphol, P.; Mendes, A.I.; Romeiras, F.; Sousa, M.; Schreiber, R.; Matos, P.; Jordan, P.; Metha, A.; Amaral, M.D.; Kunzelmann, K.; Farinha, C.M.Previously, the pleiotropic "master kinase" casein kinase 2 (CK2) was shown to interact with CFTR, the protein responsible for cystic fibrosis (CF). Moreover, CK2 inhibition abolished CFTR conductance in cell-attached membrane patches, native epithelial ducts, and Xenopus oocytes. CFTR possesses two CK2 phosphorylation sites (S422 and T1471), with unclear impact on its processing and trafficking. Here, we investigated the effects of mutating these CK2 sites on CFTR abundance, maturation, and degradation coupled to effects on ion channel activity and surface expression. We report that CK2 inhibition significantly decreased processing of wild-type (wt) CFTR, with no effect on F508del CFTR. Eliminating phosphorylation at S422 and T1471 revealed antagonistic roles in CFTR trafficking: S422 activation versus T1471 inhibition, as evidenced by a severe trafficking defect for the T1471D mutant. Notably, mutation of Y512, a consensus sequence for the spleen tyrosine kinase (SYK) possibly acting in a CK2 context adjacent to the common CF-causing defect F508del, had a strong effect on both maturation and CFTR currents, allowing the identification of this kinase as a novel regulator of CFTR. These results reinforce the importance of CK2 and the S422 and T1471 residues for regulation of CFTR and uncover a novel regulation of CFTR by SYK, a recognized controller of inflammation.
- First report on MRSA CC398 recovered from wild boars in the north of PortugalPublication . Sousa, M.; Silva, N.; Manageiro, V.; Ramos, S.; Coelho, A.; Gonçalves, D.; Caniça, M.; Torres, C.; Igrejas, G.; Poeta, P.More than 47% of the Portuguese hospital S. aureus isolates are methicillin-resistant (MRSA): one of the highest rates in Europe [1]. Anyhow, MRSA are becoming increasingly prevalent in community-acquired infections and, in recent years, new genetic lineages of MRSA were associated to livestock animals (LA-MRSA) [2, 3]. Nevertheless, less information do exists about the prevalence of MRSA in wild animals but, since 2013, these animals are pointed as natural hosts of MRSA strains [4]. The aim of the present study was to evaluate the antimicrobial resistance in Staphylococcus aureus recovered from wild boars, to analyze their genetic lineages, and to investigate the susceptibility to oxacillin. Samples from mouth and nose of 45 wild boars (Sus scrofa) were collected during hunt activity from November 2012 to January 2013 in the North of Portugal. S. aureus isolates were recovered from 30 of these samples (33%); one isolate/sample was further studied. The susceptibility of the isolates was tested by disk-diffusion test against 14 antimicrobial agents and minimal inhibitory concentration was used to test oxacillin according to EUCAST 2014 guidelines. The genetic lineages of S. aureus were characterized by agr-typing, spa-typing and MLST. From the 30 isolates, 18 S. aureus were susceptible to all antibiotics tested and 7 presented resistance to one or more of the following antibiotics: penicillin (n=3), oxacillin (n=4), cefoxitin (n=1), clindamycin (n=2), gentamicin (n=1), fusidic acid (n=1), ciprofloxacin (n=2), tetracycline (n=1) and linezolid (n=1). One MRSA CC398 (spa-type t899) isolate was detected (oxacillin MIC=32mg/L and mecApositive), which presented resistance to penicillin, tetracycline, and ciprofloxacin and contained the genes of immune evasion cluster (IEC) system (type B). The 29 methicillin-susceptible isolates were typed as ST1 (t1533), ST133 (t3583), ST1643 (t10712), ST2328 (t3750) and the new STs (3220, 3222, 3223, 3224) associated to new spa-types t14311 and t14312. The agr types I, II, III and IV were identified. It is a matter of concern when MRSA and some specific lineages of S. aureus are taken as commensal habitants of the skin and nose of wild animals and are characterized with resistance to various antimicrobial agents in clinical use.
- Human spermatogenic failure purges deleterious mutation load from the autosomes and both sex chromosomes, including the gene DMRT1Publication . Lopes, Alexandra; Aston, Kenneth I.; Thompson, Emma E; Carvalho, Filipa; Gonçalves, João; Huang, N.; Matthiesen, Rune; Noordam, Michiel J.; Quintela, Ines; Ramu, Avinash; Seabra, Catarina; Wilfert, Amy B.; Dai, Juncheng; Downie, Jonathan; Fernandes, Susana; Guo, Xuejiang; Shah, Jiahao; Amorim, Antonio; Barros, Alberto; Carracedo, A.; Hu, Z.; Hurles, M.E.; Moskovtsev, S.; Ober, C.; Paduch, D.A.; Schiffman, J.D.; Schlegel, P.N.; Sousa, M.; Carrell, D.T.; Conrad, D.F.Gonadal failure, along with early pregnancy loss and perinatal death, may be an important filter that limits the propagation of harmful mutations in the human population. We hypothesized that men with spermatogenic impairment, a disease with unknown genetic architecture and a common cause of male infertility, are enriched for rare deleterious mutations compared to men with normal spermatogenesis. After assaying genomewide SNPs and CNVs in 323 Caucasian men with idiopathic spermatogenic impairment and more than 1,100 controls, we estimate that each rare autosomal deletion detected in our study multiplicatively changes a man’s risk of disease by 10% (OR 1.10 [1.04–1.16], p,261023), rare X-linked CNVs by 29%, (OR 1.29 [1.11–1.50], p,161023), and rare Y-linked duplications by 88% (OR 1.88 [1.13–3.13], p,0.03). By contrasting the properties of our case-specific CNVs with those of CNV callsets from cases of autism, schizophrenia, bipolar disorder, and intellectual disability, we propose that the CNV burden in spermatogenic impairment is distinct from the burden of large, dominant mutations described for neurodevelopmental disorders. We identified two patients with deletions of DMRT1, a gene on chromosome 9p24.3 orthologous to the putative sex determination locus of the avian ZW chromosome system. In an independent sample of Han Chinese men, we identified 3 more DMRT1 deletions in 979 cases of idiopathic azoospermia and none in 1,734 controls, and found none in an additional 4,519 controls from public databases. The combined results indicate that DMRT1 loss-of-function mutations are a risk factor and potential genetic cause of human spermatogenic failure (frequency of 0.38% in 1306 cases and 0% in 7,754 controls, p = 6.261025). Our study identifies other recurrent CNVs as potential causes of idiopathic azoospermia and generates hypotheses for directing future studies on the genetic basis of male infertility and IVF outcomes.
- New biomarkers to fight urogenital schistosomiasis: a major neglected tropical disease [editorial]Publication . Botelho, M.C.; Sousa, M.
- Rare double sex and mab-3-related transcription factor 1 regulatory variants in severe spermatogenic failurePublication . Lima, A.C.; Carvalho, F.; Gonçalves, J.; Fernandes, S.; Marques, P.I.; Sousa, M.; Barros, A.; Seixas, A.; Amorim, A.; Conrad, D.F.; Lopes, M.The double sex and mab-3-related transcription factor 1 (DMRT1) gene has long been linked to sex-determining pathways across vertebrates and is known to play an essential role in gonadal development and maintenance of spermatogenesis in mice. In humans, the genomic region harboring the DMRT gene cluster has been implicated in disorders of sex development and recently DMRT1 deletions were shown to be associated with non-obstructive azoospermia (NOA). In this work, we have employed different methods to screen a cohort of Portuguese NOA patients for DMRT1 exonic insertions and deletions [by multiplex ligation probe assay (MLPA); n = 68] and point mutations (by Sanger sequencing; n = 155). We have found three novel patient-specific non-coding variants in heterozygosity that were absent from 357 geographically matched controls. One of these is a complex variant with a putative regulatory role (c.-223_-219CGAAA>T), located in the promoter region within a conserved sequence involved in Dmrt1 repression. Moreover, while DMRT1 domains are highly conserved across vertebrates and show reduced levels of diversity in human populations, two rare synonymous substitutions (rs376518776 and rs34946058) and two rare non-coding variants that potentially affect DMRT1 expression and splicing (rs144122237 and rs200423545) were overrepresented in patients when compared with 376 Portuguese controls (301 fertile and 75 normozoospermic). Overall our previous and present results suggest a role of changes in DMRT1 dosage in NOA potentially also through a process of gene misregulation, even though DMRT1 deleterious variants seem to be rare.
- Staphylococci among Wild European Rabbits from the Azores: A Potential Zoonotic Issue?Publication . Sousa, M.; Silva, V.; Silva, A.; Silva, N.; Ribeiro, J.; Tejedor-Junco, M.T.; Capita, R.; Chenouf, N.S.; Alonso-Calleja, C.; Rodrigues, T.M.; Leitão, M.; Gonçalves, D.; Caniça, M.; Torres, C.; Igrejas, G.; Poeta, P.The prevalence and diversity of Staphylococcus species from wild European rabbits (Oryctolagus cuniculus) in the Azores were investigated, and the antibiotic resistance phenotype and genotype of the isolates were determined. Nasal samples from 77 wild European rabbits from São Jorge and São Miguel islands in Azores were examined. Antibiotic susceptibility of the isolates was determined using the Kirby-Bauer disk diffusion method, and the presence of antimicrobial resistance genes and virulence factors was determined by PCR. The genetic lineages of S. aureus isolates were characterized by spa typing and multilocus sequence typing. A total of 49 staphylococci were obtained from 35 of the 77 wild rabbits. Both coagulase-positive (8.2%) and coagulase-negative (91.8%) staphylococci were detected: 4 S. aureus, 17 S. fleurettii, 13 S. sciuri, 7 S. xylosus, 4 S. epidermidis, and 1 each of S. simulans, S. saprophyticus, S. succinus, and S. equorum. The four S. aureus isolates showed methicillin susceptibility and were characterized as spa type t272/CC121, Panton-Valentine leukocidin negative, and hlB positive. Most of the coagulase-negative staphylococci showed resistance to fusidic acid and beta-lactams, and multidrug resistance was identified especially among S. epidermidis isolates. The mecA gene was detected in 20 isolates of the species S. fleurettii and S. epidermidis, associated with the blaZ gene in one S. epidermidis isolate. Five antimicrobial resistance genes were detected in one S. epidermidis isolate (mecA,dfrA,dfrG,aac6'-aph2'', and ant4). Our results highlight that wild rabbits are reservoirs or "temporary hosts" of Staphylococcus species with zoonotic potential, some of them carrying relevant antimicrobial resistances.
- The role of estrogens and estrogen receptor signaling pathways in cancer and infertility: the case of schistosomesPublication . Botelho, M.C.; Alves, H.; Barros, A.M.; Rinaldi, G.; Brindley, P.J.; Sousa, M.Schistosoma haematobium, a parasitic flatworm that infects more than 100 million people, mostly in the developing world, is the causative agent of urogenital schistosomiasis, and is associated with a high incidence of squamous cell carcinoma (SCC) of the bladder. Schistosomiasis haematobia also appears to negatively influence fertility, and is particularly associated with female infertility. Given that estrogens and estrogen receptors are key players in human reproduction, we speculate that schistosome estrogen-like molecules may contribute to infertility through hormonal imbalances. Here, we review recent findings on the role of estrogens and estrogen receptors on both carcinogenesis and infertility associated with urogenital schistosomiasis and discuss the basic hormonal mechanisms that might be common in cancer and infertility.
- Untreated sewage contamination of beach sand from a leaking underground sewage systemPublication . Brandão, J.; Albergaria, I.; Albuquerque, J.; José, S.; Grossinho, J.; Ferreira, F.C.; Raposo, A.; Rodrigues, R.; Silva, C.; Jordao, L.; Sousa, M.; Rebelo, M.H.; Veríssimo, C.; Sabino, R.; Amaro, T.; Cardoso, F.; Patrão-Costa, M.; Solo-Gabriele, H.Thirty people (mostly children) experienced an episode of skin rash days after a sand sifting beach operation at Porto Pim Beach in Faial, Azores during June 2019. An environmental and epidemiologic investigation was conducted to identify the cause of the outbreak of skin rash. The epidemiologic investigation found that some of the patients experiencing symptoms had never entered the beach water. During the pollution period and throughout the epidemiologic investigation, faecal indicator bacteria levels (94 CFU/100 ml for intestinal enterococci and 61 CFU/100 ml for Escherichia coli) in water remained under the limits used for the ninety-five percentile calculation of an Excellent coastal and transitional bathing water defined in the Portuguese Legislation (100 CFU/100 ml for intestinal enterococci and 250 CFU/100 ml for Escherichia coli). Thus sand contact was considered as a likely primary exposure route. Sand microbiological analysis for faecal indicator organisms and electron microscopy strongly suggested faecal contamination. Chemical analysis of the sand also revealed a concomitant substance compatible with sodium-hypochlorite as analysed using gas chromatography and subsequently confirmed by free chlorine analysis. Inspection of the toilet facilities and sewage disposal system revealed a leaking sewage distribution box. Collectively, results suggest that the cause of the outbreak was the leaking underground sewage distribution box that serviced the beach toilet facilities (40 m from beach), where sodium-hypochlorite was used for cleaning and disinfection. This sewage then contaminated the surficial sands to which beach goers were exposed. Chlorine being an irritant substance, was believed to have been the cause of the symptoms given the sudden presentation and dissipation of skin rashes. No gastro-intestinal illness was reported during this episode and during the following 30 days. Like water, beach sand should also be monitored for safety, especially for areas serviced by aged infrastructure.
- Urinary Estrogen Metabolites and Self-Reported Infertility in Women Infected with Schistosoma haematobiumPublication . Santos, J.; Gouveia, M.J.; Vale, N.; Delgado, M. de L.; Gonçalves, A.; da Silva, J.M.; Oliveira, C.; Xavier, P.; Gomes, P.; Santos, L.L.; Lopes, C.; Barros, A.; Rinaldi, G.; Brindley, P.J.; da Costa, J.M.; Sousa, M.; Botelho, M.C.Schistosomiasis is a neglected tropical disease, endemic in 76 countries, that afflicts more than 240 million people. The impact of schistosomiasis on infertility may be underestimated according to recent literature. Extracts of Schistosoma haematobium include estrogen-like metabolites termed catechol-estrogens that down regulate estrogen receptors alpha and beta in estrogen responsive cells. In addition, schistosome derived catechol-estrogens induce genotoxicity that result in estrogen-DNA adducts. These catechol estrogens and the catechol-estrogen-DNA adducts can be isolated from sera of people infected with S. haematobium. The aim of this study was to study infertility in females infected with S. haematobium and its association with the presence of schistosome-derived catechol-estrogens.