Joana Isabel Reis Castro

Scopus Publications

Antimicrobial Resistance in the Food Chain: Bridging Knowledge Gaps for Effective Detection and Control
Emílio Gomes, Tomás Gonçalves Mesquita, Patrícia Serra, Daniela Araújo, Carina Almeida, António Machado, Ricardo Oliveira, Joana Castro
Antibiotics, 2026
Antimicrobial resistance (AMR) poses a critical global public health threat, with the food chain serving as a significant transmission route connecting animals, environment, and humans. This review adopts a One Health perspective to analyze the key drivers of AMR dissemination across animal agriculture, aquaculture and food processing. We evaluate detection methodologies, contrasting the regulatory gold standard of culture-based phenotypic testing with rapid molecular advancements, including Whole Genome Sequencing (WGS), metagenomics, and emerging CRISPR-Cas diagnostics. While molecular tools offer unprecedented speed and resolution, challenges such as matrix interference, the viable but non-culturable (VBNC) state, and the genotype-phenotype disconnect remain. Finally, integrated mitigation strategies are also described, ranging from on-farm antimicrobial stewardship and innovative biofilm control to consumer hygiene practices. It is essential to bridge the technical and regulatory gaps in AMR surveillance in order to develop effective interventions and ensure a safer food system.
Editorial: Perspectives in clinical infectious diseases: 2024/2025
Ricardo Oliveira, Lorena V. N. Oliveira, Sónia Silva, Joana Castro
Frontiers in Cellular and Infection Microbiology, 2026
Clinical infectious diseases remain a significant global health challenge, fueled by the ongoing emergence and re-emergence of pathogens, the rising threat of antimicrobial resistance, and the increasing complexity of populations dynamics. The interplay between microbial adaptation, host susceptibility, environmental pressures and healthcare system dynamics has profoundly reshaped the infectious disease landscape, making continuous innovation in diagnostics, therapeutics and surveillance essential.Over the past decade, breakthroughs in molecular biology, genomics, and data-driven approaches have reshaped how infectious diseases are detected, evaluated and managed. However, persistent challenges such as delayed diagnosis, limited treatment options for resistant pathogens and disparities in healthcare access continue to impact patient outcomes worldwide. These issues are further exacerbated by the burden of chronic comorbidities, ageing populations, and globalization.The Research Topic "Perspectives in Clinical Infectious Diseases: 2024/2025" brings together 25 contributions that collectively provide an overview of the current state of the field while also highlighting innovative directions and unresolved challenges. The included studies span a wide range of infectious diseases and clinical contexts, incorporating epidemiological analyses, diagnostic evaluations, therapeutic assessments, mechanistic insights, and case-based observations. This collection's integrative approach is a major strength, reflecting the inherently interdisciplinary nature of infectious disease research. Collectively, the articles emphasize the importance of combining clinical expertise with advances in microbiology, immunology, molecular diagnostics, and public health. This issue highlights the importance of a comprehensive approach that effectively bridges the gap between scientific research and clinical practice.Thus, the goal of this editorial is to summarize the key themes addressed in this Research Topic, focusing on the scientific and clinical contributions of the included articles, and to offer insights into future directions in the field. Together, these studies provide valuable perspectives on evolving challenges and opportunities, reinforcing the critical importance of continued innovation and collaboration to address the global burden of infectious diseases.The contributions included in this Research Topic collectively illustrate the rapid evolution and increasing complexity of clinical infectious diseases. Rather than representing isolated advances, these studies converge to reveal several overarching trends that are reshaping the field: the growing integration of precision diagnostics into clinical workflows, the persistent and escalating challenge of antimicrobial resistance, the critical influence of host factors on disease susceptibility and outcomes, and the expanding role of interdisciplinary and One Health approaches in understanding and controlling infectious diseases.Within this conceptual framework, the contributions can be categorized into four complementary and interrelated domains that mirror these emerging priorities: (i) infectious disease epidemiology and host susceptibility, (ii) antimicrobial resistance and therapeutic strategies, (iii) advances in diagnostics and clinical management, and (iv) emerging pathogens, rare infections, and One Health perspectives.A substantial number of contributions focus on the epidemiology of infectious diseases and the role of host-related risk factors in shaping susceptibility to and outcomes of these diseases. Liu and colleagues highlighted the increased risk of latent tuberculosis among patients with type 2 diabetes, reinforcing the importance of considering associated risk factors in the population (Liu et al., 2026). Similarly, Li et al. discussed the diagnostic and therapeutic challenges of latent tuberculosis in children with rheumatic diseases, emphasizing the importance of proactive management in immunologically vulnerable groups (Li et al., 2025).The influence of host-related factors in infection outcomes is further explored in studies addressing viral and systemic infections. Wang and colleagues showed the association between fatty liver disease and an increased risk of liver failure in patients with acute hepatitis B, illustrating how metabolic comorbidities influence disease progression and should be considered in early risks assessment and clinical management (Wang et al., 2026). Sun et al. investigated the impact of human papillomavirus infections on male reproductive health, highlighting the negative effects on fertility and the increased risk of transmission to partners, as well as the broader systemic consequences of viral infections. These findings emphasize that infectious diseases should not be viewed independently but rather understood within the broader context of overall patient health, including metabolic, immunological, and reproductive systems (Sun et al., 2025).In parallel, genetic susceptibility is addressed by Lv et al., who identify a novel loss-of-function variant in STAT1 associated with Mendelian susceptibility to mycobacterial disease (Lv et al., 2025). Hua et al. explored the HBV-miR-3 and cGAS-STING axis, identifying a novel immunomodulatory pathway that may represent a promising therapeutic target in chronic hepatitis B (Hua et al., 2025). These studies highlight the importance of host genetics in determining the risk of infection, disease progression, and clinical outcomes, and pave the way for a more personalized approach managing infectious diseases. However, the translation of these findings into routine clinical practice remains in its early stages.Together, these contributions emphasize that infections should not be understood in isolation, but instead as the result of complex interactions among pathogens, host biology, and underlying comorbidities.Antimicrobial resistance (AMR) emerges as a central issue across several studies, reflecting its global health relevance and the ongoing need for effective therapeutic strategies to address resistant infections.Wang and colleagues provided a comparative analysis of bloodstream infections caused by carbapenem-resistant Klebsiella pneumoniae and Escherichia coli in hematological patients (Wang et al., 2025). The authors addressed the severe clinical impact of resistant infections in immunocompromised populations while also highlighting potential therapeutic alternatives, such as tigecycline or ceftazidime-avibactam, which were associated with improved outcomes. Similarly, Prebianchi et al. characterized hospital-associated Methicillin-resistant Staphylococcus aureus (MRSA) genotypes responsible for community-onset infections, demonstrating the convergence of hospital and community reservoirs. In addition, they observed that deep tissue involvement, rather than specific bacterial genotypes, was the main predictor of severe outcomes and mortality (Bazanelli Prebianchi et al., 2025).Beyond resistance patterns, several studies in this Research Topic address the critical need for therapeutic optimization. Song et al. examined the prevalence and clinical impact of timely carbapenem administration, emphasizing the importance of pharmacokinetic/pharmacodynamic (PK/PD) target fulfillment in improving treatment efficacy (Song et al., 2025). Zhou and coworkers compared the effectiveness and safety of vancomycin versus linezolid in central nervous system infections, providing evidence to inform therapeutic decision-making, suggesting that, given their comparable effectiveness, treatment choice should be guided by patient-specific factors (Zhou et al., 2025b).Innovative and adjunctive therapeutic approaches are also highlighted in the context of other severe diseases. Sun et al. reported the successful use of blood purification therapy in a patient with severe falciparum malaria complicated by cytokine storm and multiorgan dysfunction syndrome, indicating the potential of adjunctive therapies in critical care settings (Sun et al., 2026).Collectively, these findings underscore the growing threat posed by resistant pathogens, particularly among vulnerable populations, and reinforce the importance of antimicrobial stewardship, early appropriate therapy, and PK/PD-guided treatment strategies. At the same time, they reveal both progress and persistent limitations in current therapeutic approaches, including the continued reliance on existing antimicrobial agents, many of which are decades old.Another key focus of this Research Topic is the advancement of diagnostic methodologies and their integration into clinical practice. Studies highlight the limitations of conventional culture-based techniques and the increasing clinical value of molecular approaches, reflecting a broader shift toward precision diagnostics in infectious diseases.Lin and coworkers (Lin et al., 2025) and Gao and colleagues (Gao et al., 2025) evaluated metagenomic next-generation sequencing (mNGS) in comparison with conventional culture methods for periprosthetic joint infection and severe pneumonia, respectively. These studies highlight the superior sensitivity and broader pathogen detection capabilities of mNGS, particularly in complex or culturenegative cases. Similarly, Miguélez Sánchez et al. assessed the diagnostic performance of broad-range PCR in bacterial peritonitis, reinforcing the value of molecular diagnostics in clinical microbiology (Miguélez Sánchez et al., 2025). Importantly, these approaches not only enhance diagnostic yield but also reveal the polymicrobial nature of infections that may otherwise remain undetected.In addition to advances in pathogen detection, several studies underline the expanding role of biomarkers in guiding clinical management and therapeutic decision-making. Chen et al. identified serum ferritin as a predictive low-cost biomarker for interferon therapy response in chronic hepatitis B (Chen et al., 2025), while Shi et al. evaluated serum apolipoprotein A1 as a prognostic marker in elderly patients with SARS-CoV-2 infection (Shi et al., 2025). Complementing these findings, Zhou and colleagues analyzed determinants of functional cure in interferon-treated chronic hepatitis B, demonstrating the prognostic value of HBsAg dynamics and clinical predictors in guiding therapeutic strategies (Zhou et al., 2025a). These biomarker-driven approaches, particularly in the context of viral infections, further emphasize the growing importance of measurable biological indicators in predicting disease progression and informing individualized care.The integration of diagnostics into clinical decision-making emerges as a critical determinant of impact. Kramme et al. demonstrated that combining rapid diagnostic testing with immediate infection consultation significantly increased the proportion of patients receiving targeted antibiotic therapy in septic intensive care settings (Kramme et al., 2025). These findings underscore that diagnostic innovation alone is insufficient unless it is effectively integrated into clinical workflows and supported by timely expertise.Taken together, these contributions illustrate a paradigm shift toward precision medicine in infectious diseases, where diagnostics, biomarkers, and clinical decision-making are increasingly integrated to optimize outcomes and patient care. However, the implementation of precision diagnostics and biomarker-guided strategies in routine practice remains uneven. Barriers related to cost, accessibility, data interpretation, and variability in biomarker performance across populations continue to limit widespread adoption, highlighting the need for further validation and standardization.The emergence of novel and rare pathogens, together with evolving epidemiological patterns among well-established organisms, remains a critical challenge in clinical infectious diseases. Collectively, the contributions in this section underscore the importance of continuous surveillance, advanced diagnostics, and integrative approaches to better understand pathogen dynamics across different contexts.Chen et al. provided a comprehensive analysis of candidemia over a three-year period, offering valuable epidemiological insights into fungal infections (Chen et al., 2026). In parallel with these findings, case reports by Lu et al. and Huang et al. describe infections caused by the fungal pathogen Trichosporon asahii (Lu et al., 2025) and the rare bacterial species Wohlfahrtiimonas (Huang et al., 2025), respectively, underscoring the clinical relevance of uncommon and opportunistic pathogens, particularly in immunocompromised individuals. These studies illustrate the expanding spectrum of infectious agents encountered in modern clinical practice and the challenges associated with their diagnosis and management.At the population level, Valenzuela et al. analyzed an emm1-dominant group A Streptococcus outbreak, situating it within a broader epidemiological context and demonstrating the importance of longitudinal surveillance in outbreak detection and characterization (Valenzuela et al., 2026). Similarly, Liu et al. described the prevalence of multiple respiratory pathogens among children using multiplex PCR, highlighting the complexity of pathogen circulation in community settings and the value of comprehensive surveillance strategies (Liu et al., 2025).A particularly important cross-cutting theme in this Research Topic is the adoption of One Health and genomic surveillance approaches. In this context, Gomes et al. emphasized the role of whole genome sequencing (WGS) in monitoring foodborne pathogens, aligning with the One Health framework (Gomes et al., 2025). This approach aligns with the One Health framework, which recognizes the interconnectedness of human, animal, and environmental health, and underscores the need for interdisciplinary strategies in infectious disease control.In addition, Akula et al. raised important considerations regarding laboratory practices and experimental integrity, demonstrating that methodological factors can significantly influence both infectious disease research and the interpretation of immunological findings (Akula et al., 2025).Collectively, these studies highlight the need for robust surveillance systems, methodological rigor, and cross-sector collaboration. They further emphasize that addressing emerging and rare pathogens requires not only heightened clinical awareness but also integrated approaches that combine genomic technologies, epidemiological monitoring, and One Health principles.The Research Topic "Perspectives in Clinical Infectious Diseases: 2024/2025" provides a comprehensive and timely synthesis of current advances, ongoing challenges, and emerging directions in the field of clinical infectious diseases. The 25 contributions included in this collection taken together demonstrate that infectious diseases remain a highly dynamic and multifaceted domain, requiring continuous adaptation of diagnostic, therapeutic, and surveillance strategies.A central message from this Editorial is the expanding role of precision diagnostics and molecular technologies in transforming clinical practice. Advances such as mNGS and broad-range molecular assays are enabling faster, more accurate, and more comprehensive pathogen detection, particularly in complex and polymicrobial infections. However, their full clinical impact depends on effective integration into healthcare systems, underscoring the need to bridge technological innovation with clinical expertise, infrastructure, and standardized workflows.At the same time, the persistent global threat of antimicrobial resistance continues to challenge current treatment paradigms. The studies included in this Research Topic highlight the urgent need for strengthened antimicrobial stewardship, optimization of therapeutic strategies, and sustained investment in the development of new antimicrobial agents. The disproportionate burden of resistant infections in vulnerable populations further emphasizes the need for coordinated and globally aligned responses.Another key insight is the growing recognition of the influence of host-related factors, including comorbidities, immune status, and genetic predisposition, on infectious disease susceptibility and outcomes. Integrating these factors into clinical decision-making supports the movement toward more personalized and patient-centered approaches to care.Altogether, the contributions in this Topic highlight the importance of addressing both common and rare infections, as well as the continuous emergence of new pathogens and clinical scenarios. The inclusion of case reports, outbreak analyses, and epidemiological studies reflects the need for vigilance and adaptability in clinical practice, particularly in the context of immunocompromised patients and evolving microbial landscapes.Importantly, this Research Topic also reinforces the value of interdisciplinary collaboration and One Health approaches, integrating human, animal, and environmental health perspectives. Such strategies are essential for understanding transmission dynamics, improving surveillance systems, and enhancing preparedness for future infectious disease threats.Despite these advances, several limitations and gaps remain evident across the included studies. Many are retrospective in design, limiting causal inference, while others rely on single-center data, potentially restricting generalizability. In addition, the translation of advanced diagnostics and biomarker discoveries into routine clinical practice remains incomplete, often hindered by logistical, financial, and infrastructural barriers.Addressing these challenges will require well-designed prospective, multicenter, and interventional studies, alongside sustained investment and international collaboration. More broadly, continued progress in clinical infectious diseases will depend not only on technological and scientific innovation but also on effective integration, translation, and equitable accessibility, supported by close coordination among clinicians, microbiologists and immunologists, epidemiologists, and public health professionals.In conclusion, the contributions presented in this Research Topic reflect the current state of clinical infectious diseases and shed light on the path forward. By promoting interdisciplinary collaboration and embracing emerging technologies, the field is well-positioned to address the evolving challenges posed by infectious diseases and improve patient outcomes worldwide.
Evidence from a broad-range PNA probe links several Prevotella species to bacterial vaginosis
Taynara Mulinde, Lúcia G. V. Sousa, Joana Castro, Sheridan D. George, Christina A. Muzny, Nuno Cerca
Peerj, 2026
Background Bacterial vaginosis (BV) is the most prevalent vaginal infection among reproductive-age women. It is associated with multiple adverse health outcomes in women including adverse pregnancy outcomes, an increased risk of pelvic inflammatory disease, infertility, and an increased risk of HIV and other sexually transmitted infections. BV is characterized by an imbalance in the vaginal microbiota, namely a decrease in protective Lactobacillus species and an overgrowth of facultative and strict anaerobic bacteria, leading to the development of a polymicrobial biofilm. Despite extensive research, the etiology of BV remains unclear, and its pathophysiology is not fully understood. It has been hypothesized that P. bivia , in combination with Gardnerella spp., plays an important role in the early development of the BV biofilm. We previously developed a peptide nucleic acid (PNA) probe specifically targeting P. bivia to investigate its role as a potential early colonizer. However, our recent findings have raised doubts about the specificity of this association, suggesting a broader involvement of other Prevotella species in incident BV (iBV). Methods A new PNA probe targeting Prevotella spp. 23S rRNA was developed compared to the existing P. bivia -specific probe. This new probe was optimized in vitro through a variation of hybridization temperatures and times. Its performance was evaluated using a collection of 28 Prevotella strains representing 24 different species and 38 non- Prevotella spp. typically found in BV in order to assess its sensitivity and specificity. Both probes were tested on vaginal swab specimens from women with and without BV to assess the bacterial count and detection of Prevotella species. Results In vitro validation demonstrated that the new Prevotella spp. probe achieved a specificity of 100% and sensitivity of 96%. As expected, its broader detection allowed identification of a wider range of Prevotella spp. compared to the P. bivia -specific probe, which was intentionally restricted to a single species. Application to clinical specimens revealed that the new probe identified a significantly higher count of Prevotella spp. in 6/9 (66.6%) BV-positive specimens compared to the P. bivia -specific probe. In 2/9 (22.2%) healthy control specimens, greater Prevotella spp. detection was also observed. Conclusions Our findings suggest that the involvement of Prevotella spp. in BV extends beyond P. bivia , implicating a wider range of species which could be present in the polymicrobial BV biofilm. The broader specificity of this new Prevotella spp. probe provides a valuable tool for future research on the vaginal microbiome and the pathogenesis of BV.
Evaluation of antibacterial properties of Lactobacillus acidophilus and Lactobacillus gasseri on enterotoxigenic Escherichia coli associated with porcine post-weaning diarrhea (PWD) using Galleria mellonella model
D. Araújo, A.R. Silva, S. Silva, N.P. Mira, C. Almeida, J. Castro
Microbe Netherlands, 2025
Post-weaning diarrhea (PWD) in piglets, caused by enterotoxigenic Escherichia coli (ETEC), has been managed using antibiotics and zinc oxide. However, concerns regarding antibiotic resistance as well as the 2022 EU ban on zinc oxide, have prompted the development of sustainable alternatives. Growing evidence suggests that probiotics may play a promising role in controlling ETEC infections. In this study, we first evaluated the ability of three potential probiotics ( Lactobacillus gasseri, Lactobacillus acidophilus, and Limosilactobacillus reuteri ) to reduce ETEC infection in vitro . Subsequently, we used the Galleria mellonella model to further evaluate their impact in vivo , by testing two different approaches: co-infection (i.e. probiotic + ETEC) and prophylactic strategy (i.e. prior infection with probiotic for 4 h followed by ETEC infection). The survival rate and health index scores were assessed after 24, 48, and 72 h. In addition, quantitative reverse transcription polymerase chain reaction (RT-qPCR) was also performed to determine the transcript levels of genes encoding the G. mellonella antimicrobial peptides (AMPs), namely, lysozyme, inducible metalloproteinase inhibitor (IMPI), gallerimycin, and galliomycin, to infer the immune response to ETEC infection. Our results suggest that a co-infection strategy was ineffective in controlling ETEC infection under our experimental conditions. On the other hand, when using a prophylactic strategy, we observed significant differences between the treated larvae and the control. Overall, we observed that L. acidophilus significantly reduced infection by the ETEC strain SP11 ( p <0.05), while L. gasseri demonstrated anti-ETEC potential, specifically against the SP31 strain ( p <0.05). Differences in IMPI and galliomycin AMPs expression were also observed between treated and control conditions. However, these variations depended on the specific strains of both Lactobacillus and ETEC involved. In conclusion, selected Lactobacillus species showed promising potential as prophylactic alternatives for controlling ETEC infection, although efficacy was strain-dependent. • PWD in piglets is primarily caused by enterotoxigenic E. coli (ETEC). • Restrictions on zinc oxide and antibiotics raise the need for alternatives. • L. acidophilus and L. gasseri show strain-specific anti-ETEC effects. • Certain Lactobacillus strains improve survival and health in G. mellonella model. • Strain-specific probiotics may modulate immunity against ETEC infections.
Assessing host-adaptation of new sequence types of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA)
Ricardo Oliveira, Daniela Araújo, Joana Castro, Teresa Nogueira, Gonçalo Almeida, Nuno F. Azevedo, Carina Almeida
Microbial Pathogenesis, 2025
The importance of animals as reservoirs of human pathogens is widely recognised, with livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) being consistently reported. This study presents a phenotypic and genotypic characterization of five MRSA isolates from raw bovine milk in northern Portugal, using culturomics and whole genome sequencing (WGS), to assess their potential risk for human infection. Resistance to beta-lactams was confirmed by the presence of the mecA resistance gene as part of the SCC mec type V element (5C2&5), complemented by a resistance profile to other antibiotics characteristic of adaptation to animal hosts. Several virulence factors were identified, including some with potential human tropism, such clumping factors, gamma-hemolysins and staphylococcal complement inhibitors. All MRSA isolates gave rise to new sequence types (ST), newly added to the MLST database as ST8475, ST8894 and ST8895, which show a variation of 1 or 2 alleles compared to the recognised LA-MRSA ST398/CC398 lineage. A phylogenetic comparison of these MRSA isolates with other S. aureus isolates confirmed a close genetic relationship with the ST398/CC398 lineage. This finding further supports the inclusion of these new STs in this clonal complex. A pathogenicity in vivo test using the Galleria mellonella model, which has an innate immune system similar to humans, showed a high pathogenicity of the MRSA isolates, and an assessment of biofilm production showed a similar capacity for biomass formation among all isolates. These findings demonstrate a typical adaptation of the isolates to animal hosts, while retaining or acquiring traits associated with the ancestral human host of S. aureus. Such dual adaptation emphasizes their zoonotic potential and strengthens the relevance of LA-MRSA as a critical public health concern. • Methicillin-resistant Staphylococcus aureus (MRSA) from raw bovine milk were characterised by whole genome analysis, and in vitro and in vivo tests. • Resistance genes identified included mecA - SCC mec type Vc element and tetracyclines and phenicols genes in mobile elements. • Virulence factors-related genes with a potential human tropism were identified, including clfA /B , hlgA /B/C , and scn . • MRSA isolates belong to novel STs (ST8475, ST8894, ST8895) related to the European LA-MRSA predominant ST398/CC398 lineage. • They demonstrated pathogenicity in the in vivo Galleria mellonella model and the strong-moderate ability to form biofilms.
Gardnerella species exhibit synergy in their ability to displace Lactobacillus crispatus adhered to HeLa cells
Ângela Lima, Joana Castro, Christina A. Muzny, Nuno Cerca
Peerj, 2025
Background Bacterial vaginosis (BV) is the most common vaginal infection in reproductive-age women. It is associated with adverse pregnancy complications, such as preterm birth and low birth weight, in addition to an increased risk of acquisition of HIV and sexually transmitted infections. BV is characterized by a vaginal dysbiosis, involving loss of protective Lactobacillus species (including L. crispatus ) and overgrowth of facultative and strict anaerobic bacteria, with Gardnerella species playing a predominant role. However, despite extensive research on BV pathogenesis, its etiology remains unclear, and the sequence of events leading to the displacement of lactobacilli by anaerobic bacteria in women has not yet been fully elucidated. Until 2019, all bacteria belonging to the Gardnerella genus were considered part of the species G. vaginalis . However, it is now recognized that different Gardnerella species exist, each with varying virulence potentials. Recent data have shown that multiple subgroups of Gardnerella spp. are frequently detected simultaneously in the vaginal microbiota of women with BV. With this in mind, we aimed to test the hypothesis that different combinations of known Gardnerella species, isolated from the vaginal microbiota of women with BV, have an enhanced ability to compete against Lactobacillus crispatus , pre-adhered to HeLa cells, thereby facilitating the early stages of BV development. Methods Adhesion assays of dual combinations of Gardnerella spp. ( G. vaginalis , G. leopoldii , G. swidsinskii , G. piotii ) were performed on an in vitro model of HeLa cells, covered with and without L. crispatus . Quantification of the species used in our assays was subsequently performed using quantitative polymerase chain reaction (qPCR). Results Our results revealed synergy between different Gardnerella spp., demonstrating their ability to overcome the presumed protective effect of L. crispatus , thereby creating favorable conditions for the development of a polymicrobial biofilm characteristic of BV. The combination of G. vaginalis and G. leopoldii showed the greatest synergistic effect on initial adhesion to HeLa cells while the combination of G. leopoldii and G. swidsinskii had the greatest ability to reduce L. crispatus colonization. Conclusions Although this in vitro study does not unequivocally prove that BV is initiated by the disruption of normal vaginal microbiota by Gardnerella spp., it strongly supports this possibility, contributing to a better understanding of BV etiology.
An Asset for Food Safety: The Knowledge Behind the Physiological Alterations Induced by ETEC Enterotoxins
Maria Margarida Barros, Ana Maria Campos, Joana Castro, Ricardo Oliveira, Daniela Araújo, Divanildo Outor-Monteiro, Carina Almeida
Foods, 2025
Foodborne pathogens represent a significant public health risk in both developed and developing countries. Among these pathogens, enterotoxigenic Escherichia coli (ETEC) is a major cause of diarrhea in humans and one of the leading causes of mortality in newly weaned pigs. The main sources of ETEC contamination include environments with poor hygiene and contaminated water, meat, cereals, and vegetables. Therefore, this review manuscript focuses on the pathogenesis of ETEC in humans and pigs. The main virulence factors responsible for ETEC-associated infections, such as colonization factors and toxins, will be described for both species, with particular emphasis on the toxins as well as, their classification and structural characterization. More specifically, this study will outline the main physiological alterations and adaptive mechanisms induced by these enterotoxins, namely heat-stable toxin (ST) and heat-labile toxin (LT), in the three most affected systems: the gastrointestinal system, the enteric nervous system (ENS), and the immune system. This set of findings provides a deeper insight into the pathogenesis of this relevant foodborne pathogen, which is crucial for empowering food scientists and stakeholders to more effectively mitigate associated risks. As such, it provides valuable understanding of toxin activity, serving as a means to raise awareness of food safety practices and strengthening risk communication, surveillance and intervention strategies, thereby ensuring consumer protection. Additionally, this knowledge enables the development of preventive strategies to reduce ETEC infections, thereby decreasing the need for clinical management among consumers exposed to this bacterium. Ultimately, it contributes to the preservation of public health, the reduction of antimicrobial use, and the lowering of antimicrobial resistance gene prevalence.
Amphiphilic dendrimer-assisted delivery of antisense nucleic acid mimics against E. coli
Mariana Gomes, Igor Resende, Yana Zamoshchak, Daniela Araújo, Joana Castro, Dinesh Dhumal, Ling Peng, Rita S. Santos, Nuno F. Azevedo
Journal of Controlled Release, 2025
The rise in antimicrobial resistance and the consequent ineffectiveness of conventional antibiotics emphasise the need for novel therapeutic strategies. Antisense nucleic acid mimics (NAMs) are emerging as promising precision therapeutic agents, inhibiting specific genes through hybridisation with selected nucleic acid targets. However, delivering NAMs into bacteria remains a significant challenge. This study explores the use of poly(amidoamine) (PAMAM) amphiphilic dendrimers (ADs) as delivery vehicles for NAMs targeting the essential acpP gene in Escherichia coli . Two ADs bearing primary amine or tertiary amine terminals, 1a and 1b, were tested for their ability to permeabilise the bacterial envelope, facilitate NAM internalisation, and enhance NAM-based antibacterial activity. Physicochemical characterisation studies, flow cytometry measurements, fluorescence and electron microscopy imaging, bacterial viability assays, and an in vivo toxicity assessment using a greater wax moth ( Galleria mellonella ) model were conducted. Both ADs acted as permeabilisers of the bacterial envelope and assisted in NAM internalisation and antibacterial activity. The most effective formulation, 1b combined with the peptide nucleic acid (PNA)-based NAM, achieved an 8 log 10 reduction in viable bacteria, with sustained activity up to 24 h against E. coli . In vivo, the most promising formulations showed no toxicity, with G. mellonella larvae maintaining overall health and no significant mortality detected for up to three days. These findings demonstrate that amphiphilic dendrimers can effectively deliver PNA-based NAMs, highlighting their potential as a novel strategy against antimicrobial-resistant pathogens. • Two amphiphilic dendrimers, 1a and 1b, were employed for the first time to deliver antisense nucleic acids, based on locked nucleic acid (LNA) and peptide nucleic acid (PNA), as a novel strategy against E. coli . • Both ADs were shown to act as envelope permeabilisers, with dose-dependent antibacterial activity. • While 1a and LNA showed better results regarding nucleic acid internalisation, 1b and PNA led to a more effective bacterial inhibition. • The PNA-based antisense sequence consistently outperformed the scrambled sequence, emphasising the bacterial specificity of this strategy. • The studied formulations showed no signs of toxicity in G. mellonella .
In Vitro Investigation of the Antimicrobial Properties of Gerês Propolis in Bacteria Isolated from Companion Animals and Safety Profile Characterization Using the Galleria mellonella Model
Rafael Rodrigues, Rui Almeida, Soraia C. V. Rodrigues, Joana Castro, Ricardo Oliveira, Nuno Mendes, Carina Almeida, Sónia Silva, Daniela Araújo, Cristina Almeida-Aguiar
Pathogens, 2025
Propolis, also known as bee glue, is a natural resinous mixture produced by Western honeybees and has long been recognized for its potential therapeutic properties. Recent research has focused on its diverse bioactivities, particularly its antimicrobial effects against a broad spectrum of microorganisms, including human but also animal pathogens. However, further investigation is required to fully elucidate the pharmacological properties and potential toxicity of propolis to support its broader application. This study investigated the antimicrobial efficacy and safety of an ethanol extract of Portuguese propolis collected from the Gerês region (G23.EE). The antimicrobial activity was assessed in vitro against both Gram-positive and Gram-negative bacteria associated with infections in companion animals, using the agar dilution method. To evaluate potential toxicity, the extract was administered via injection and topical application in an in vivo Galleria mellonella larval model, with health parameters monitored over a 96 hours period. The in vitro results showed that G23.EE was more effective against Gram-positive bacteria, including Staphylococcus spp. (e.g., S. felis, S. hominis, S. simulans, and S. pseudintermedius; MIC = 0.5 mg/mL) and Enterococcus faecium (MIC = 1.5 mg/mL), than against Gram-negative bacteria, such as Escherichia coli and Klebsiella oxytoca (MIC > 8.0 mg/mL). No significant adverse effects were observed in G. mellonella larvae following injection or brushing with propolis extract concentrations up to 8.0 mg/mL. Overall, these findings suggest that Portuguese propolis from Gêres is a promising, safe, and effective natural antimicrobial agent for targeting Gram-positive bacterial infections in companion animals.
Students' summer internships in a research centre: The impact on scientific literacy and the choice of a career in the STEM fields
Elisa Saraiva, Sónia Silva, Joana Castro, Daniela Araújo, Carina Almeida, Maria Manuel Azevedo
Heliyon, 2025
Science communication is crucial for engaging society in real-life contexts. In this line of thought, research centres play a pivotal role in promoting dynamic and interactive communication that enables science to be disseminated. This study constitutes a qualitative research based on a case study involving middle school students, researchers and teachers, which took place in a research centre in Portugal that provided a two-week internship experience for students. The results confirm a significant increase in students' knowledge, attitudes, and 21st-century skills. The student's participation in the internship inspires them to pursue further education, in general, and in the sciences in particular. Unfortunately, such opportunities are uncommon for middle school students, so, extracurricular activities that provide authentic science experiences outside the regular school curriculum are of utmost value.
NADES-in-Oil Emulsions Enriched with Essential Oils for Cosmetic Application
David S. Freitas, Diana Rocha, Joana Santos, Jennifer Noro, Tânia D. Tavares, Marta O. Teixeira, Daniela Araújo, Joana Castro, Carina Almeida, Sónia Silva, Helena P. Felgueiras, Artur Ribeiro, Armando Venâncio, Artur Cavaco-Paulo, Carla Silva
Processes, 2025
Advances in whole genome sequencing for foodborne pathogens: implications for clinical infectious disease surveillance and public health
Emílio Gomes, Daniela Araújo, Teresa Nogueira, Ricardo Oliveira, Sónia Silva, Lorena V. N. Oliveira, Nuno F. Azevedo, Carina Almeida, Joana Castro
Frontiers in Cellular and Infection Microbiology, 2025
Exploring the Spectrum of Biofilm Complexity
Daniela Araújo, Ricardo Oliveira, Ana Maria Campos, Maria Margarida Barros, Andreia S. Azevedo, Ana Barbosa, Laura Cerqueira, Nuno F. Azevedo, Sónia Silva, Carina Almeida, Joana Castro
Microbial Biofilm Dynamics Contemporary Approaches Models and Analytical Tools, 2025
Laboratory Models for Biofilm Studies
Joana Castro, Ricardo Oliveira, Ana Maria Campos, Maria Margarida Barros, Andreia S. Azevedo, Ana Barbosa, Laura Cerqueira, Nuno F. Azevedo, Sónia Silva, Carina Almeida, Daniela Araújo
Microbial Biofilm Dynamics Contemporary Approaches Models and Analytical Tools, 2025
Multidrug-resistant Klebsiella pneumoniae and Klebsiella variicola isolated from patients in Portuguese hospitals: Genomic and phenotypic characterization
Joana Castro, Daniela Araújo, Hugo Oliveira, Liliana Fernandes, Ricardo Oliveira, Erick Brinks, Gyu-Sung Cho, Charles Franz, Maria José Saavedra, Sónia Silva, Carina Almeida
Microbe Netherlands, 2024
Developing translucent emulsions using sustainable NADES-based extracts
David S. Freitas, Diana Rocha, Joana Santos, Jennifer Noro, Tânia D. Tavares, Marta O. Teixeira, Daniela Araújo, Joana Castro, Sónia Silva, Helena P. Felgueiras, Artur Ribeiro, Armando Venâncio, Artur Cavaco-Paulo, Carla Silva
Journal of Molecular Liquids, 2024
Probiotics, prebiotics, and synbiotics as antimicrobials: A one-health perspective
Advances in Health and Disease, 2024
Evaluation of Simultaneous Growth of Escherichia coli O157:H7, Salmonella spp., and Listeria monocytogenes in Ground Beef Samples in Different Growth Media
José Mário Sousa, Ana Barbosa, Daniela Araújo, Joana Castro, Nuno Filipe Azevedo, Laura Cerqueira, Carina Almeida
Foods, 2024
Antimicrobial resistance patterns of Staphylococcus spp. isolated from clinical specimens of companion animals in Northern Portugal, 2021–2023
D. Araújo, R. Oliveira, B.L. Silva, J. Castro, C. Ramos, F. Matos, C. Almeida, S. Silva
Veterinary Journal, 2024
A New Peptide Nucleic Acid Fluorescence In Situ Hybridization Probe for the Specific Detection of Salmonella Species in Food Matrices
Mário Sousa, Rui Rocha, Daniela Araújo, Joana Castro, Ana Barbosa, Nuno F. Azevedo, Laura Cerqueira, Carina Almeida
Foodborne Pathogens and Disease, 2024
Molecular characterization and virulence profile of Klebsiella pneumoniae and Klebsiella oxytoca isolated from ill cats and dogs in Portugal
Joana Castro, Ricardo Oliveira, Liliana Fernandes, Isabel Carvalho, Hugo Oliveira, Erik Brinks, Gyu-Sung Cho, Charles Franz, Carina Almeida, Sónia Silva, Daniela Araújo
Veterinary Microbiology, 2024
Emerging Approaches for Mitigating Biofilm-Formation-Associated Infections in Farm, Wild, and Companion Animals
Daniela Araújo, Ana Rita Silva, Rúben Fernandes, Patrícia Serra, Maria Margarida Barros, Ana Maria Campos, Ricardo Oliveira, Sónia Silva, Carina Almeida, Joana Castro
Pathogens, 2024
The Role of Flagellum and Flagellum-Based Motility on Salmonella Enteritidis and Escherichia coli Biofilm Formation
Diana Vilas Boas, Joana Castro, Daniela Araújo, Franklin L. Nóbrega, Charles W. Keevil, Nuno F. Azevedo, Maria João Vieira, Carina Almeida
Microorganisms, 2024
Application of DNA aptamers to block enterotoxigenic Escherichia coli toxicity in a Galleria mellonella larval model
Maria Margarida Barros, Joana Castro, Daniela Araújo, Ricardo Oliveira, Ana Maria Campos, Sónia Silva, Divanildo Outor-Monteiro, Carina Almeida
Frontiers in Chemistry, 2024
Exploring the prevalence and antibiotic resistance profile of Klebsiella pneumoniae and Klebsiella oxytoca isolated from clinically ill companion animals from North of Portugal
D. Araújo, J. Castro, F. Matos, R. Oliveira, C. Ramos, C. Almeida, S. Silva
Research in Veterinary Science, 2023
Swine Colibacillosis: Global Epidemiologic and Antimicrobial Scenario
Maria Margarida Barros, Joana Castro, Daniela Araújo, Ana Maria Campos, Ricardo Oliveira, Sónia Silva, Divanildo Outor-Monteiro, Carina Almeida
Antibiotics, 2023
Effect of phage vB_EcoM_FJ1 on the reduction of ETEC O9:H9 infection in a neonatal pig cell line
Alice Ferreira, Daniela Silva, Carina Almeida, Maria Elisa Rodrigues, Sónia Silva, Joana Castro, Dalila Mil-Homens, Isidro García-Meniño, Azucena Mora, Mariana Henriques, Ana Oliveira
Veterinary Research, 2023
Synergistic effects of carvacrol, α-terpinene, γ-terpinene, ρ-cymene and linalool against Gardnerella species
Lúcia G. V. Sousa, Joana Castro, Carlos Cavaleiro, Lígia Salgueiro, Mariana Tomás, Rita Palmeira-Oliveira, José Martinez-Oliveira, Nuno Cerca
Scientific Reports, 2022
Gardnerella Vaginalis Dominates Multi-Species Biofilms in both Pre-Conditioned and Competitive In Vitro Biofilm Formation Models
Aliona S. Rosca, Joana Castro, Ângela França, Mario Vaneechoutte, Nuno Cerca
Microbial Ecology, 2022
Exploring the Antibiotic Resistance Profile of Clinical Klebsiella pneumoniae Isolates in Portugal
Ricardo Oliveira, Joana Castro, Sónia Silva, Hugo Oliveira, Maria José Saavedra, Nuno Filipe Azevedo, Carina Almeida
Antibiotics, 2022
Swine enteric colibacillosis: Current treatment avenues and future directions
Joana Castro, Maria Margarida Barros, Daniela Araújo, Ana Maria Campos, Ricardo Oliveira, Sónia Silva, Carina Almeida
Frontiers in Veterinary Science, 2022
Bacteriocin Production by Escherichia coli during Biofilm Development
Hanna Fokt, Sara Cleto, Hugo Oliveira, Daniela Araújo, Joana Castro, Nuno Cerca, Maria João Vieira, Carina Almeida
Foods, 2022
In vitro interactions within a biofilm containing three species found in bacterial vaginosis (BV) support the higher antimicrobial tolerance associated with BV recurrence
Aliona S Rosca, Joana Castro, Lúcia G V Sousa, Angela França, Mario Vaneechoutte, Nuno Cerca
Journal of Antimicrobial Chemotherapy, 2022
Six Bacterial Vaginosis-Associated Species Can Form an In Vitro and Ex Vivo Polymicrobial Biofilm That Is Susceptible to Thymbra capitata Essential Oil
Aliona S. Rosca, Joana Castro, Lúcia G. V. Sousa, Angela França, Carlos Cavaleiro, Lígia Salgueiro, Nuno Cerca
Frontiers in Cellular and Infection Microbiology, 2022
Exploiting the Anti-Biofilm Effect of the Engineered Phage Endolysin PM-477 to Disrupt In Vitro Single-and Dual-Species Biofilms of Vaginal Pathogens Associated with Bacterial Vaginosis
Joana Castro, Lúcia G. V. Sousa, Ângela França, Lenka Podpera Tisakova, Lorenzo Corsini, Nuno Cerca
Antibiotics, 2022
Crystal Violet Staining Alone Is Not Adequate to Assess Synergism or Antagonism in Multi-Species Biofilms of Bacteria Associated With Bacterial Vaginosis
Joana Castro, Ângela Lima, Lúcia G. V. Sousa, Aliona S. Rosca, Christina A. Muzny, Nuno Cerca
Frontiers in Cellular and Infection Microbiology, 2022
A New PNA-FISH Probe Targeting Fannyhessea vaginae
Lúcia G. V. Sousa, Joana Castro, Angela França, Carina Almeida, Christina A. Muzny, Nuno Cerca
Frontiers in Cellular and Infection Microbiology, 2021
Atopobium vaginae and prevotella bivia are able to incorporate and influence gene expression in a pre-formed gardnerella vaginalis biofilm
Joana Castro, Aliona S. Rosca, Christina A. Muzny, Nuno Cerca
Pathogens, 2021
Essential Oils as Potential Antibiofilm Agents: Insights into the Key Role of Thymbra capitata to Fight Biofilm-Associated Infections
Volatile Oils Production Composition and Uses, 2021
Gardnerella vaginalis Enhances Atopobium vaginae Viability in an in vitro Model
Joana Castro, Aliona S. Rosca, Piet Cools, Mario Vaneechoutte, Nuno Cerca
Frontiers in Cellular and Infection Microbiology, 2020
Genetic Heterogeneity and Taxonomic Diversity among Gardnerella Species
Joana Castro, Kimberly K. Jefferson, Nuno Cerca
Trends in Microbiology, 2020
Evaluation of different culture media to support in vitro growth and biofilm formation of bacterial vaginosis-associated anaerobes
Aliona S. Rosca, Joana Castro, Nuno Cerca
Peerj, 2020
Gardnerella and vaginal health: The truth is out there
Aliona S Rosca, Joana Castro, Lúcia G V Sousa, Nuno Cerca
FEMS Microbiology Reviews, 2019
Unveiling the role of Gardnerella vaginalis in polymicrobial Bacterial Vaginosis biofilms: the impact of other vaginal pathogens living as neighbors
Joana Castro, Daniela Machado, Nuno Cerca
Isme Journal, 2019
Innate immune components affect growth and virulence traits of bacterial-vaginosis-associated and non-bacterial-vaginosis-associated Gardnerella vaginalis strains similarly
Joana Castro, Kimberly K Jefferson, Nuno Cerca
Pathogens and Disease, 2019
Lactobacillus crispatus represses vaginolysin expression by BV associated Gardnerella vaginalis and reduces cell cytotoxicity
Joana Castro, Ana Paula Martins, Maria Elisa Rodrigues, Nuno Cerca
Anaerobe, 2018
Comparative transcriptomic analysis of Gardnerella vaginalis biofilms vs. planktonic cultures using RNA-seq
Joana Castro, Angela França, Katie R. Bradwell, Myrna G. Serrano, Kimberly K. Jefferson, Nuno Cerca
Npj Biofilms and Microbiomes, 2017
Prevalence of bacterial vaginosis in Portuguese pregnant women and vaginal colonization by Gardnerella vaginalis
Daniela Machado, Joana Castro, José Martinez-de-Oliveira, Cristina Nogueira-Silva, Nuno Cerca
Peerj, 2017
Escherichia coli and Enterococcus faecalis are able to incorporate and enhance a pre-formed Gardnerella vaginalis biofilm
Joana Castro, Daniela Machado, Nuno Cerca
Pathogens and Disease, 2016
Bacterial vaginosis biofilms: Challenges to current therapies and emerging solutions
Daniela Machado, Joana Castro, Ana Palmeira-de-Oliveira, José Martinez-de-Oliveira, Nuno Cerca
Frontiers in Microbiology, 2016
BV and non-BV associated Gardnerella vaginalis establish similar synergistic interactions with other BV-associated microorganisms in dual-species biofilms
Joana Castro, Nuno Cerca
Anaerobe, 2015
Using an in-vitro biofilm model to assess the virulence potential of Bacterial Vaginosis or non-Bacterial Vaginosis Gardnerella vaginalis isolates
Joana Castro, Patrícia Alves, Cármen Sousa, Tatiana Cereija, Ângela França, Kimberly K. Jefferson, Nuno Cerca
Scientific Reports, 2015
Diagnosis of bacterial vaginosis by a new multiplex peptide nucleic acid fluorescence in situ hybridization method
António Machado, Joana Castro, Tatiana Cereija, Carina Almeida, Nuno Cerca
Peerj, 2015
Comparative genomics of Lactobacillus crispatus suggests novel mechanisms for the competitive exclusion of Gardnerella vaginalis
Teija Ojala, Matti Kankainen, Joana Castro, Nuno Cerca, Sanna Edelman, Benita Westerlund-Wikström, Lars Paulin, Liisa Holm, Petri Auvinen
BMC Genomics, 2014
Gardnerella vaginalis Outcompetes 29 other bacterial species isolated from patients with bacterial vaginosis, using in an in vitro biofilm formation model
Patrícia Alves, Joana Castro, Cármen Sousa, Tatiana B. Cereija, Nuno Cerca
Journal of Infectious Diseases, 2014
Reciprocal interference between Lactobacillus spp. and Gardnerella vaginalis on initial adherence to epithelial cells
Joana Castro, Ana Henriques, António Machado, Mariana Henriques, Kimberly K. Jefferson, Nuno Cerca
International Journal of Medical Sciences, 2013
Influence of anaerobic conditions on vaginal microbiota recovery from bacterial vaginosis patients
Tatiana B Cereija, Joana Castro, Patrícia Alves, Nuno Cerca
Sexually Transmitted Infections, 2013

Joana Isabel Reis Castro

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