Daniela Marlene da Silva Machado
@ucp.pt
Centre of Biotechnology and Fine Chemistry
Catholic University of Portugal
Daniela Marlene da Silva Machado completed her PhD in Chemical and Biological Engineering in 2017 by University of Minho and BSc degree in Clinical Analysis and Public Health in 2010 by the Health Technology Superior School of Polytechnic Institute of Porto. Since May 2019, she is junior researcher in Centre of Biotechnology and Fine Chemistry of the Catholic University of Portugal. Works in the area(s) of Biological Sciences (with emphasis in Microbiology) and Engineering and Technology (with emphasis in Industrial Biotechnology). She had 2 research fellowships, a PhD fellowship ( from March 2013 to July 2017) working in epidemiologic profile of bacterial vaginosis and the search of novel strategies to treat this vaginal disorder and research fellowship in FCT project (from November 2017 to April 2019) targeting to develop microencapsulation technology applied to beneficial gut extreme oxygen-sensitive bacteria. Moreover, she has experience in teaching Microbiology classes at polytechnic and university scientific activities includes: jury member of Master thesis and member of Agência de Avaliação e Acreditação do Ensino Superior (A3ES), participating in diverse audits.
EDUCATION
Ph.D in Chemical and Biological Engineering, University of Minho
BSc in Clinical Analysis and Public Health, Health Technology Superior School of Polytechnic Institute of Porto
RESEARCH INTERESTS
Explore next generation probiotics as live microbial strategies to prevent oro-gastrointestinal infections
Scopus Publications
Scopus Publications
Daniela Machado, Mariana Fonseca, Rita Vedor, Sérgio Sousa, Joana Cristina Barbosa, and Ana Maria Gomes
MDPI AG
Akkermansia muciniphila is considered a next-generation probiotic to be incorporated in new food and pharmaceutical formulations. Effective delivery systems are required to ensure high probiotic viability and stability during product manufacture, shelf-life, and post-consumption, namely, throughout digestion. Hydrogelated matrices have demonstrated promising potential in this dominion. Hence, this work aimed to evaluate the effect of a calcium-alginate hydrogelated matrix on A. muciniphila viability during 28-days refrigerated aerobic storage and when exposed to simulated gastrointestinal conditions, in comparison with that of free cells. Akkermansia muciniphila was successfully encapsulated in the calcium-alginate matrix via extrusion (60% encapsulation yield). Furthermore, encapsulated A. muciniphila exhibited high stability (a loss in viability lower than 0.2 log-cycle) after 28-days of refrigerated aerobic storage, maintaining its viability around 108 CFU/g. Prominently, as the storage time increased, encapsulated A. muciniphila revealed higher viability and stability regarding in vitro gastrointestinal conditions than free cells. This suggests that this encapsulation method may attenuate the detrimental effects of prolonged aerobic storage with a subsequent gastrointestinal passage. In conclusion, encapsulation via extrusion using a calcium-alginate hydrogelated matrix seems to be a promising and adequate strategy for safeguarding A. muciniphila from adverse conditions encountered during refrigerated aerobic storage and when exposed to the gastrointestinal passage.
Rita Vedor, Daniela Machado, Joana Cristina Barbosa, Diana Almeida, and Ana Maria Gomes
Elsevier BV
Margarida Faustino, Daniela Machado, Dina Rodrigues, José Carlos Andrade, Ana Cristina Freitas, and Ana Maria Gomes
MDPI AG
Marine algae have been emerging as natural sources of bioactive compounds, such as soluble dietary fibers and peptides, presenting special interest as ingredients for functional foods. This study developed a cheese spread incorporating red seaweed Osmundea pinnatifida extract and subsequently characterized it in terms of nutritional, pH, and microbiological parameters and bioactivities including prebiotic, antidiabetic, antihypertensive, and antioxidant activities. This food was produced through incorporation of O. pinnatifida extract (3%), obtained via enzymatic extraction Viscozyme L in a matrix containing whey cheese (75%) and Greek-type yoghurt (22%). The product was then subjected to thermal processing and subsequently stored for 21 days at 4 °C. During storage, this food showed a high pH stability (variations lower than 0.2 units), the absence of microbial contamination and all tested bioactivities at the sampling timepoints 0 and 21 days. Indeed, it exerted prebiotic effects under Lactobacillus acidophilus LA-5® and Bifidobacterium animalis subsp. lactis BB-12®, increasing their viability to around 4 and 0.5 log CFU/g, respectively. In addition, it displayed antidiabetic (α-glucosidase inhibition: 5–9%), antihypertensive (ACE inhibition: 50–57%), and antioxidant (ABTS: 13–15%; DPPH: 3–5%; hydroxyl radical: 60–76%) activities. In summary, the cheese spread produced may be considered an innovative food with high potential to contribute toward healthier status and well-being of populations.
Daniela Machado, Melany Domingos, Joana Cristina Barbosa, Diana Almeida, José Carlos Andrade, Ana Cristina Freitas, and Ana Maria Gomes
MDPI AG
Faecalibacterium duncaniae is an intestinal commensal bacterium proposed as a next-generation probiotic due to its promising outcomes in the treatment and prevention of several human diseases, which demonstrate its multiple contributions to the host’s health. However, its strict anaerobic nature has created several hurdles in the development of functional foods, nutraceuticals, and biotherapeutic products. Herein, we explored freeze-dried formulations containing prebiotics, cryoprotectants, and antioxidant agents as a technological strategy to enhance the viability of F. duncaniae DSM 17677 upon aerobic storage and gastrointestinal tract conditions. Our results indicate that freeze-dried F. duncaniae in a matrix containing inulin, sucrose, cysteine, and riboflavin survived at levels higher than 106 CFU/g and around 105 CFU/g after 1 and 4 days of aerobic storage at room temperature, respectively. Thus, the freeze-dried formulation with inulin, sucrose, cysteine, and riboflavin presents as a protective strategy to improve F. duncaniae viability under aerobic environments. Nevertheless, incorporation of a suitable coating aimed at protecting F. duncaniae against the detrimental gastrointestinal passage effects is urgently required, given its high susceptibility to extreme acidic pH values and bile.
Diana Almeida, Daniela Machado, Sérgio Sousa, Catarina Leal Seabra, Joana Cristina Barbosa, José Carlos Andrade, Ana Maria Gomes, and Ana Cristina Freitas
Elsevier BV
Daniela Machado, Joana Cristina Barbosa, Diana Almeida, José Carlos Andrade, Ana Cristina Freitas, and Ana Maria Gomes
MDPI AG
Akkermansia muciniphila is a Gram-negative intestinal anaerobic bacterium recently proposed as a novel probiotic candidate to be incorporated in food and pharmaceutical forms. Despite its multiple health benefits, the data addressing its antimicrobial susceptibility profile remain scarce. However, the absence of acquired resistance in probiotic strains is a compulsory criterion for its approval in the qualified presumption of safety list. This study aimed at characterizing the A. muciniphila DSM 22959 strain’s antimicrobial susceptibility profile using phenotypic and in silico approaches. To establish the phenotypic antimicrobial susceptibility profile of this strain, minimum inhibitory concentrations of eight antimicrobials were determined using broth microdilution and E-test methods. Additionally, the A. muciniphila DSM 22959 genome was screened using available databases and bioinformatics tools to identify putative antimicrobial resistance genes (ARG), virulence factors (VF), genomic islands (GI), and mobile genetic elements (MGE). The same categorization was obtained for both phenotypic methods. Resistance phenotype was observed for gentamicin, kanamycin, streptomycin, and ciprofloxacin, which was supported by the genomic context. No evidence was found of horizontal acquisition or potential transferability of the identified ARG and VF. Thus, this study provides new insights regarding the phenotypic and genotypic antimicrobial susceptibility profiles of the probiotic candidate A. muciniphila DSM 22959.
Joana Barbosa, Diana Almeida, Daniela Machado, Sérgio Sousa, Ana Freitas, José Andrade, and Ana Gomes
MDPI AG
Akkermansia muciniphila is regarded as a promising next-generation probiotic or live biotherapeutic candidate. Effective delivery strategies must be developed to ensure high enough viability of the probiotic strain throughout its industrial formulation, distribution chain, shelf-life, and, ultimately, the host’s gastrointestinal tract, where it should exert its beneficial effect(s). Among the possible methodologies, spray-drying is considered industrially attractive regarding its costs, efficiency, and scalability, with the due parameter customization. In this study, spray-drying was explored as a one-step process to encapsulate A. muciniphila DSM 22959, testing the drying settings and three different dairy-based matrices. Microcapsule morphology and size was assessed, and viability throughout storage at 4 or 22 °C and simulated gastrointestinal passage was determined. Akkermansia muciniphila microencapsulation by spray-drying, using 10% skim milk and inlet/outlet temperatures of 150/65 °C, is effective in terms of viability stabilization, both during prolonged aerobic storage and exposure to simulated gastrointestinal passage. Akkermansia muciniphila viability was maintained at around 107 CFU/g up to 28 days at 4 °C under aerobic conditions with viability losses inferior to 1 log reduction. This methodology provides the necessary conditions to efficiently deliver the recommended dose of live A. muciniphila in the human gut as a live biotherapeutic product.
Daniela Machado, Joana Cristina Barbosa, Melany Domingos, Diana Almeida, José Carlos Andrade, Ana Cristina Freitas, and Ana Maria Gomes
Elsevier BV
Faecalibacterium prausnitzii, a resident anaerobic bacterium commonly found in healthy gut microbiota, has been proposed as a next generation probiotic with high potential for application in food matrices and pharmaceutical formulations. Despite its recognized health benefits, detailed information regarding its antimicrobial susceptibility profile is still lacking. However, this information is crucial to determine its safety, since the absence of acquired antimicrobial resistance is required to qualify a probiotic candidate as safe for human and animal consumption. Herein, the antimicrobial susceptibility profile of F. prausnitzii DSM 17677 strain was evaluated by integrating both phenotypic and in silico data. Phenotypic antimicrobial susceptibility was evaluated by determining minimum inhibitory concentrations of 9 antimicrobials using broth microdilution and E-test® methods. Also, the whole genome of F. prausnitzii DSM 17677 was analysed, using several databases and bioinformatics tools, to identify possible antibiotic resistance genes (ARG), genomic islands (GI) and mobile genetic elements (MGE). With exception of erythromycin, the same classification (susceptible or resistant) was obtained in both broth microdilution and E-test® methods. Phenotypic resistance to ampicillin, gentamycin, kanamycin and streptomycin were detected, which was supported by the genomic context. Other ARG were also identified but they seem not to be expressed under the tested conditions. F. prausnitzii DSM 17677 genome contains 24 annotated genes putatively involved in resistance against the following classes of antimicrobials: aminoglycosides (such as gentamycin, kanamycin and streptomycin), macrolides (such as erythromycin), tetracyclines and lincosamides. The presence of putative ARG conferring resistance to β-lactams could only be detected using a broader homology search. The majority of these genes are not encoded within GI or MGE and no plasmids were reported for this strain. Despite the fact that most genes are related with general resistance mechanisms, a streptomycin-specific ARG poses the only potential concern identified. This specific ARG is encoded within a GI and a MGE, meaning that it could have been laterally acquired and might be transferred to other bacteria present in the same environment. Thus, our findings provide relevant insights regarding the phenotypic and genotypic antimicrobial resistance profiles of the probiotic candidate F. prausnitzii DSM 17677.
Glenise Bierhalz Voss, Daniela Machado, Joana Cristina Barbosa, Débora A. Campos, Ana Maria Gomes, and Manuela Pintado
Elsevier
Joana Cristina Barbosa, Daniela Machado, Diana Almeida, José Carlos Andrade, Adriano Brandelli, Ana Maria Gomes, and Ana Cristina Freitas
Elsevier
Joana Cristina Barbosa, Daniela Machado, Diana Almeida, José Carlos Andrade, Adriano Brandelli, Ana Maria Gomes, and Ana Cristina Freitas
Elsevier
Diana Almeida, Daniela Machado, José Carlos Andrade, Sónia Mendo, Ana Maria Gomes, and Ana Cristina Freitas
Informa UK Limited
Abstract In recent years, scientific community has been gathering increasingly more insight on the dynamics that are at play in metabolic and inflammatory disorders. These rapidly growing conditions are reaching epidemic proportions, bringing clinicians and researcher’s new challenges. The specific roles and modulating properties that beneficial/probiotic bacteria hold in the context of the gut ecosystem seem to be key to avert these inflammatory and diet-related disorders. Faecalibacterium prausnitzii, Akkermansia muciniphila and Eubacterium hallii have been identified as candidates for next generation probiotics (NGPs) with exciting potential for the prevention and treatment of such of dysbiosis-associated diseases. The challenges of these non-conventional native gut bacteria lie mainly on their extreme sensitivity to O2 traces. If these strains are to be used successfully in food, supplements or drugs they need to be stable and active in humans. In the present review, we present an overall perspective of the most updated scientific literature on the newly called NGPs through the 5W1H (What, Why, Who, Where, When, and How) method, an innovative and attractive problem-solving approach that provides the reader an effective understanding of the issue at hand.
José Carlos Andrade, Diana Almeida, Melany Domingos, Catarina Leal Seabra, Daniela Machado, Ana Cristina Freitas, and Ana Maria Gomes
Frontiers Media SA
In the last years several human commensals have emerged from the gut microbiota studies as potential probiotics or therapeutic agents. Strains of human gut inhabitants such as Akkermansia, Bacteroides, or Faecalibacterium have shown several interesting bioactivities and are thus currently being considered as food supplements or as live biotherapeutics, as is already the case with other human commensals such as bifidobacteria. The large-scale use of these bacteria will pose many challenges and drawbacks mainly because they are quite sensitive to oxygen and/or very difficult to cultivate. This review highlights the properties of some of the most promising human commensals bacteria and summarizes the most up-to-date knowledge on their potential health effects. A comprehensive outlook on the potential strategies currently employed and/or available to produce, stabilize, and deliver these microorganisms is also presented.
Daniela Machado, Diana Almeida, Catarina L. Seabra, José Carlos Andrade, Ana Maria Gomes, and Ana Cristina Freitas
Elsevier BV
Data regarding Akkermansia muciniphila viability under stress remains scarce despite its beneficial potential. Therefore, the main goal was to assess A. muciniphila culturability when exposed to different temperatures, atmospheres and gastrointestinal simulated conditions. Cultivable cell numbers A. muciniphila remain high after refrigerated and room temperatures oxygen exposure, and gastrointestinal passage.
Daniela Machado, Diana Almeida, Catarina Leal Seabra, José Carlos Andrade, Ana Maria Gomes, and Ana Cristina Freitas
Springer International Publishing
Nanotechnology is a fast-rising industry not defined by a single field of research, but as the convergence of disciplines, such as chemistry, biology, physics, mathematics, and engineering, which exploits the benefits of nanoscale dimensions and characteristics for application in the macroworld. Current applications vary widely from nanorobotic industry to simple household items. However, the combination of such phenomena with probiotic science, another emerging and potentially promising area for the prevention and treatment of several human gastrointestinal and extraintestinal disorders using beneficial microorganisms, gives birth to “nanoprobiotics,” a field that focuses on the application of nanoscience into the probiotic-related world. In this chapter, we will navigate through the basic nanotech and probiotic knowledge and the current technologies employed with success for probiotic delivery and, ultimately, discuss what possibilities lie ahead in the nanoprobiotic future.
Joana Castro, Daniela Machado, and Nuno Cerca
Springer Science and Business Media LLC
Bacterial vaginosis (BV) is characterized by a highly structured polymicrobial biofilm, which is strongly adhered to the vaginal epithelium and primarily consists of the bacterium Gardnerella vaginalis. However, despite the presence of other BV-associated bacteria, little is known regarding the impact of other species on BV development. To gain insight into BV progress, we analyzed the ecological interactions between G. vaginalis and 15 BV-associated microorganisms using a dual-species biofilm model. Bacterial populations were quantified using a validated peptide nucleic acid fluorescence in situ hybridization approach. Furthermore, biofilm structure was analyzed by confocal laser scanning microscopy. In addition, bacterial coaggregation ability was determined as well as the expression of key virulence genes. Remarkably, our results revealed distinct biofilm structures between each bacterial consortium, leading to at least three unique dual-species biofilm morphotypes. Furthermore, our transcriptomic findings seem to indicate that Enterococcus faecalis and Actinomyces neuii had a higher impact on the enhancement of G. vaginalis virulence, while the other tested species had a lower or no impact on G. vaginalis virulence. This study casts a new light on how BV-associated species can modulate the virulence aspects of G. vaginalis, contributing to a better understanding of the development of BV-associated biofilms.
Daniela Machado, Carlos Gaspar, Ana Palmeira-de-Oliveira, Carlos Cavaleiro, Lígia Salgueiro, José Martinez-de-Oliveira, and Nuno Cerca
Future Medicine Ltd
AIM
To evaluate the antibacterial activity of Thymbra capitata essential oil and its main compound, carvacrol, against Gardnerella vaginalis grown planktonically and as biofilms, and its effect of vaginal lactobacilli.
MATERIALS & METHODS
Minimal inhibitory concentration, minimal lethal concentration determination and flow cytometry analysis were used to assess the antibacterial effect against planktonic cells. Antibiofilm activity was measured through quantification of biomass and visualization of biofilm structure by confocal laser scanning microscopy.
RESULTS
T. capitata essential oil and carvacrol exhibited a potent antibacterial activity against G. vaginalis cells. Antibiofilm activity was more evident with the essential oil than carvacrol. Furthermore, vaginal lactobacilli were significantly more tolerant to the essential oil.
CONCLUSION
T. capitata essential oil stands up as a promising therapeutic agent against G. vaginalis biofilm-related infections.
Daniela Machado, Joana Castro, José Martinez-de-Oliveira, Cristina Nogueira-Silva, and Nuno Cerca
PeerJ
BackgroundWe aimed to determine the prevalence of vaginal colonization byGardnerella vaginalisand of bacterial vaginosis (BV) in Portuguese pregnant women, and to identify risk factors for BV andG. vaginaliscolonization in pregnancy.MethodsA cross-sectional study was conducted among pregnant women aged ≥ 18 years who were attending in two public hospitals of the Northwest region of Portugal. Epidemiological data was collected by anonymous questionnaire. BV was diagnosed by Nugent criteria andG. vaginalispresence was identified by polymerase chain reaction. Crude associations between the study variables and BV orG. vaginaliscolonization were quantified by odds ratios (ORs) and their 95% confidence intervals (CIs).ResultsThe prevalences of BV and ofG. vaginaliscolonization among Portuguese pregnant women were 3.88% and 67.48%, respectively. Previous preterm delivery and colonization byG. vaginaliswere factors with very high OR, but only statistically significant for a 90% CI. Conversely, higher rates ofG. vaginaliscolonization were found in women with basic educational level (OR = 2.77, 95% CI [1.33–5.78]), during the second trimester of pregnancy (OR = 6.12, 95% CI [1.80–20.85]) and with BV flora (OR = 8.73, 95% CI [0.50–153.60]).DiscussionDespite the lower number of women with BV, prevalence ratios and association with risk factors were similar to recent European studies. However, the percentage of healthy women colonized byG. vaginaliswas significantly higher than many previous studies, confirming thatG. vaginaliscolonization does not always lead to BV development.
Joana Castro, Daniela Machado, and Nuno Cerca
Oxford University Press (OUP)
Gardnerella vaginalis is the most frequent microorganism found in bacterial vaginosis (BV), while Escherichia coli and Enterococcus faecalis are amongst the most frequent pathogens found in urinary tract infections (UTIs). This study aimed to evaluate possible interactions between UTIs pathogens and G. vaginalis using an in vitro dual-species biofilm model. Our results showed that dual-species biofilms reached significantly higher bacterial concentration than monospecies biofilms. Moreover, visualization of dual-populations species in the biofilms, using the epifluorescence microscopy, revealed that all of the urogenital pathogens coexisted with G. vaginalis. In conclusion, our work demonstrates that uropathogens can incorporate into mature BV biofilms.
Daniela Machado, Joana Castro, Ana Palmeira-de-Oliveira, José Martinez-de-Oliveira, and Nuno Cerca
Frontiers Media SA
Bacterial vaginosis (BV) is the most common genital tract infection in women during their reproductive years and it has been associated with serious health complications, such as preterm delivery and acquisition or transmission of several sexually transmitted agents. BV is characterized by a reduction of beneficial lactobacilli and a significant increase in number of anaerobic bacteria, including Gardnerella vaginalis, Atopobium vaginae, Mobiluncus spp., Bacteroides spp. and Prevotella spp.. Being polymicrobial in nature, BV etiology remains unclear. However, it is certain that BV involves the presence of a thick vaginal multi-species biofilm, where G. vaginalis is the predominant species. Similar to what happens in many other biofilm-related infections, standard antibiotics, like metronidazole, are unable to fully eradicate the vaginal biofilm, which can explain the high recurrence rates of BV. Furthermore, antibiotic therapy can also cause a negative impact on the healthy vaginal microflora. These issues sparked the interest in developing alternative therapeutic strategies. This review provides a quick synopsis of the currently approved and available antibiotics for BV treatment while presenting an overview of novel strategies that are being explored for the treatment of this disorder, with special focus on natural compounds that are able to overcome biofilm-associated antibiotic resistance.
Daniela Machado, Ana Palmeira-de-Oliveira, and Nuno Cerca
Elsevier BV
Bacterial vaginosis is the leading vaginal disorder in women in reproductive age. Although bacterial vaginosis is related with presence of a biofilm composed predominantly by Gardnerella vaginalis, there has not been a detailed information addressing the environmental conditions that influence the biofilm formation of this bacterial species. Here, we evaluated the influence of some common culture conditions on G. vaginalis biofilm formation, namely inoculum concentration, incubation period, feeding conditions and culture medium composition. Our results showed that culture conditions strongly influenced G. vaginalis biofilm formation and that biofilm formation was enhanced when starting the culture with a higher inoculum, using a fed-batch system and supplementing the growth medium with maltose.