iBB

Bacterial pathogens belonging to the Vibrio genus such as Vibrio parahaemolyticus, Vibrio alginolyticus and Vibrio vulnificus are some of the most common and widespread disease-causing agents in the aquaculture industry, inducing severe losses in fish and shellfish production worldwide. In a Review Article published in Frontiers in Microbiology, iBB researchers Gracinda Sanches Fernandes, Isabel-Sá Correia and Rodrigo Costa performed a meta-analysis of vibriosis outbreaks reported for the economically valuable, model aquaculture fish gilthead seabream (Sparus aurata) in the Mediterranean zone. The study reveals the acquisition of multiple antibiotic resistance traits among pathogenic Vibrio species during the past 25 years. An increase in human infections caused by Vibrio pathogens of fish – either via seafood ingestion or wound colonization - has been documented in the recent literature, and it is believed to result from the synergistic interaction between the expansion of intensive fish farming across coastal ecosystems and global warming. Finally, the authors critically examine the potential, and review cases of success, of alternative methods to antibiotics use to suppress vibrio pathogens in aquaculture, including vaccines, phage therapy and probiotics supplementation. Diverse bacterial symbionts of fish suppress vibrio pathogens in experimental trials, suggesting that the natural fish microbiome may be a reservoir of biological control agents of interest in applied biotechnology.

Helichrysum italicum is a halophyte shrub with bright yellow flowers with a strong curry-like aroma that have been used in the production of cosmetics and pharmaceuticals, for seasoning and flavoring of food, and as natural food preservatives. In a study  recently published in Food Research International, researchers of Universidade Nova de Lisboa, Polytechnic of Leiria, University of Coimbra, University of Lincoln, the Quadram Institute, and Carla C.C.R. de Carvalho (BERG-iBB), characterize the composition, bioactive compounds, and nutritive value of H. italicum flowers. The flowers were found to be a good source of minerals, omega-6 and omega-3 PUFAs, polyphenols, carotenoids, curcuminoids and other bioactive compounds with relevant antibacterial, antifungal and antioxidant properties.

Photo details: Helichrysum italicum, Bj.schoenmakers, CC0, via Wikimedia Commons.

The marine bacterial genus Aquimarina is a promising source of novel natural products. A new study published in Marine Drugs demonstrated widespread ability of Aquimarina species to inhibit growth of human-pathogenic microbes such as Candida glabrata and methicillin-resistant Staphylococcus aureus, as well as Vibrio and other marine bacteria relevant to aquaculture. Metabolomics and genomics analyses of Aquimarina strains indicated the presence of novel polyketides and peptides, including cyclic depsipeptide-related compounds. The study further showed that Aquimarina species possess low-abundance distributions across marine biotopes worldwide. It emphasizes the relevance of this member of the microbial rare biosphere as a promising source of novel natural products, supporting future efforts to isolate new bioactive compounds from Aquimarina. The work was developed within the SymbioReactor project (DGPM | Fundo Azul program) led by Prof Rodrigo Costa and Dr Tina Keller-Costa and was first-authored by PhD student Sandra Silva. It also includes former MSc student Patrícia Paula, Dr Dalila Mil-Homens, Prof Miguel Teixeira and Prof Arsénio Fialho from BSRG-iBB as coauthors.

A transatlantic endeavour involving researchers from BSRG-iBB, CQE (IST) and Université Laval and INRS (Canada) provided the first comprehensive assessment of prokaryotic communities in the late winter ice and its underlying water along a natural salinity gradient in Hudson Bay, a cryo-environment marking the transition between Subarctic and Arctic biomes. The study, published in Science of Total Environment, was led by Prof. Rodrigo Costa, first-authored by MIT-Portugal PhD student Lígia Coelho and co-authored by PhD student Joana Couceiro, Dr Tina Keller-Costa and Profs. Zita Martins and João Canário. The team found sharp shifts in community structure between the ice and underlying water samples at sampling sites with higher salinity, and high abundance of culturable, pigment-producing bacteria in ice. The study suggests that salinity, photosynthesis and dissolved organic matter are main drivers of prokaryotic community structure in the winter ice of Hudson Bay, the ecosystem with the fastest sea ice loss in the Canadian North.

The ocean is an excellent source for new biocatalysts due to the tremendous genetic diversity of marine microorganisms, and it may contribute to the development of sustainable industrial processes. In a recent paper published in Microorganisms, Carlos J.C. Rodrigues and Carla C.C.R. de Carvalho (iBB-BERG) used a marine bacterium for the conversion of benzaldehyde to benzyl alcohol, which is an important chemical employed as a precursor for producing esters for cosmetics and other industries. A stirred reactor, using a fed-batch approach, enabled a 1.5-fold increase in benzyl alcohol productivity when compared with batch mode. However, product accumulation in the reactor hindered the conversion. The use of a continuous flow reactor packed with immobilized cells enabled a 9.5-fold increase in productivity when compared with the fed-batch stirred reactor system.

Understanding the behavior of human pathogens is both a challenge and a need, if we are to address infectious diseases with the best possible tools. Genome-scale metabolic models are a way to meet this task, which was just used for the second most prevalent Candida species in Europe, Candida parapsilosis. The construction and validation of a global stoichiometric model describing the whole metabolic network in this human pathogen, as well as its exploitation in drug target discovery, stemming from a collaboration between researchers from BSRG/iBB (Miguel Cacho Teixeira), ITQB-NOVA (Isabel Rocha) and CEB-UMinho (Óscar Dias), was just published in Genes.

Pathogenic Vibrio species are responsible for human and animal illness and one of the main causes of human infection is related to the ingestion of undercooked seafood. Due to their filter-feeding habit, marine invertebrates, such as clams, are known to be a natural reservoir of specific microbial communities. In a recently published study carried out by researchers of ESTM, Polytechnic of Leiria, and Carla C.C.R. de Carvalho (BERG-iBB), the profile of antimicrobial resistance and the presence of virulence genes in the bacterial isolates from clams were evaluated. The presence of the mobilisable colistin resistance gene mcr-1 in three Vibrio spp. isolates highlights a potential threat to public health.

Lactococcus lactis is a food-grade, and generally recognized as safe, bacterium, which making it ideal for producing plasmid DNA (pDNA) or recombinant proteins for industrial or pharmaceutical applications. A paper published in Microorganisms by Sofia Duarte and Gabriel Monteiro from BERG-iBB reviews the major findings from L. lactis transcriptome and proteome studies, with an overexpression of native or recombinant proteins. These studies provide important insights on how to engineer the plasmid vectors and/or the strains in order to achieve high pDNA or recombinant proteins yields, with high quality standards. 

Lateral Flow Assays (LFA) are point-of-care devices with impact in healthcare. In a paper published in Sensors and Actuators B, Miguel Prazeres and colleagues from the Indian Institute of Technology (Madras) propose a LFA for traumatic brain injury (TBI) diagnostics. The new LFA employs graphene oxide (GO) particles on nitrocellulose (NC) test strips to quantitatively detect the biomarker ubiquitin carboxyl-terminal hydrolase-L1 (UCH-L1). The incorporation of 80 ng of GO at test lines boosted fluorescence signals by 2–3 times by enhancing the immobilization of capture antibodies. With LOD and LOQ that match UCH-L1 threshold levels, the assay effectively analyzed mock plasma samples simulating TBI patients. In summary, the use of GO particles in the test zone of NC strips improves signal intensity, which could potentially enhance the accuracy and efficiency of LFA-based diagnosis.

The biopharmaceutical relevance of producing plasmid DNA at large scale has increased steadily over the years due to the development of a growing number of direct and indirect applications. Be it as biological drugs or as starting materials, plasmids are pervasive across the gene and cell therapy industry of today. In a commentary article in Cell & Gene Therapy Insights, Miguel Prazeres provides an overview of the uses of plasmids, discusses manufacturing challenges, and hints at what the future may bring.

Exploration of yeast diversity for the sustainable production of biofuels, in particular biodiesel, is gaining momentum in recent years. A BSRG-iBB review paper, just published in the Journal of Fungi (Special Issue: Yeasts for a Sustainable Circular Bio-Based Economy), provides an overview of lipid production by oleaginous yeasts focusing on yeast diversity, metabolism, and other microbiological issues related to the toxicity and tolerance to multiple challenging stresses limiting bioprocess performance. Examples gathered from the literature showing the potential of different oleaginous yeasts/process conditions to produce oils for biodiesel from agro-forestry and industrial organic residues are also provided. This publication has as first-author the PhD student Marta N. Mota of the IST PhD programme in Biotechnology and Biosciences and as corresponding author her thesis supervisor, Professor Isabel Sá-Correia. Paula Múgica, from BIOREF—Collaborative Laboratory for Biorefineries, also contributed to the publication during her stay at BSRG-iBB for scientific training and collaboration in the execution of the Move2LowC project.

DNA-origami biomanufacturing relies in many cases on the use of asymmetric PCR (aPCR) to generate 500-3500 base, single-stranded DNA (ssDNA) scaffolds that are then purified by agarose gel extraction, a technique that is laborious, not scalable and presents low recovery yields. In a paper just published in Separation and Purification Technology, Ana Rita Santos and Miguel Prazeres from BERG-iBB, in collaboration with Pedro Paulo from CQE, present alternative methods based on anion-exchange or multimodal chromatography that yield purified scaffolds from aPCR mixtures. The recovered scaffolds were further used to assemble DNA-origami nanostructures. The work indicates that chromatography could play an important role in scaling-up the purification of ssDNA scaffolds for the biomanufacturing DNA-origami nanostructures.

A joint collaborative effort coordinated by Prof Jörn Piel from the ETH Zürich (Switzerland) led to the identification of new antibiotic compounds from Aquimarina sp. strain Aq135 which was cultivated from the marine sponge Ircinia variabilis in the laboratory of Prof Rodrigo Costa from BSRG-iBB. Activity-guided isolation identified novel antibacterial peptides, named aquimarins, featuring a new scaffold with an unusual C-terminal amino group and chlorine moieties. Structure–activity relationship studies with these compounds showed that the synthetically more laborious chlorinations are not required for antibacterial activity but enhance cytotoxicity. In contrast, variants lacking the C-terminal amine were virtually inactive, suggesting diamines similar to the terminal aquimarin residue as candidate building blocks for new peptidomimetic antibiotics. The study was published in Angewandte Chemie.

L-asparaginases have an acknowledge role in food and pharmaceutical industries. Accordingly, efforts to produce highly efficient, selective and stable L-asparaginases are currently underway. One of the strategies used to fulfill this goal involves the production of recombinant. In a recent paper published in Marine Biotechnology, Pedro Fernandes (iBB-BERG) and colleagues from the University of Hormozgan and Tarbiat Modares University in Iran report the production of a novel marine recombinant L‑asparaginase and provide its biochemical and structural characterization. The results obtained suggest the potential of the thermostable recombinant enzyme obtained for practical applications in food and pharmaceutical areas.

Lupanine is used as a building block in the synthesis of sparteine, a chiral selector in drug synthesis. This alkaloid is found in wastewaters derived from the debittering process that makes lupin beans edible. In a recently published work, carried out by researchers of the Faculty of Sciences and Faculty of Pharmacy from the University of Lisbon, and Teresa Esteves, Frederico Ferreira, Flávio Ferreira and Ana Mota from BERG-iBB, a computational chemistry approach was taken to design molecularly imprinted polymers (MIPs) selecting itaconic acid, a biobased building block, as a functional monomer that can provide higher affinities for lupanine. In this work, lupanine was concentrated from lupin bean wastewater by nanofiltration, extracted with ethyl acetate, and purified using the synthesized MIP, which was able to selectively recognize lupanine and improve its purity to 88%, with 82% recovery of the alkaloid, showing the potential application of this strategy to render the industrial process more sustainable.

Understanding the mechano–biological coupling mechanisms of biomaterials for tissue engineering is of major importance to assure proper scaffold performance in situ. Therefore, it is of paramount importance to establish correlations between biomaterials, their processing conditions, and their mechanical behaviour, as well as their biological performance. In a collaborative work between CDRSP-Politécnico de Leiria and SCERG-iBB  (João C. Silva and Frederico Ferreira), it was possible to infer a correlation between the addition of different concentrations of graphene nanoparticles (GPN) in three-dimensional poly(ε-caprolactone) (PCL)-based scaffolds, their extrusion-based processing parameters, and the lamellar crystal orientation observed in the different scaffolds through small-angle X-ray scattering experiments. Moreover, in vitro cell culture studies performed at SCERG-iBB demonstrated the suitability and potential of these novel 3D PCL/GPN scaffolds for tissue engineering applications. The results of this study were just published in Polymers.

A new strategy to modify cellulose with the short antimicrobial hexapeptide MP196 (RWRWRW-NH2) is proposed by BERG and BSIRG researchers that uses fusions of Cys-terminated derivatives of MP196 and a carbohydrate binding module (CBM). CBM3-MP196-modified cellulose hydrogels displayed antibacterial activity that was significantly higher when compared with controls. This versatile concept offers a toolbox for the functionalization of different cellulose materials with a broad choice in peptides. the paper was published in Acta Biomaterialia. The work was funded by project CBM-X.

When an idea in a coffee break in an international meeting turns into a paper, Pedro Fonte from BERG-iBB in collaboration with researchers from UFVJM, Brazil just published a work in JSFA Reports performing an optimization of the brewing parameters on coffee extraction using a central composite rotatable design. The effects of extraction time, particle size of ground coffee, extraction temperature, coffee-to-water ratio, stirring on caffeine yield, and soluble solids on caffeine concentrations were studied. Optimized parameters showed 45 min sufficed to perform a cold extraction at 4°C and 24°C. The parameters selected for validation were 24°C, 30% coffee-to-water ratio, a stirring of 400 rpm resulting in 3.98 mg/ml of extracted caffeine, 11.20 °Brix, and 93.9% of caffeine yield. The smaller particle size (595 μm) displayed the higher caffeine extraction of about 4 mg/ml. This study reveals the high efficiency of cold brew extraction and its potential at the industrial scale, decreasing costs with energy and extraction time, and producing a coffee rich in caffeine and soluble solids.