Microbiology, Biochemistry, Genetics and Molecular Biology, Ecology, Evolution, Behavior and Systematics, Infectious Diseases
73
Scopus Publications
Scopus Publications
Isolation of Extracellular Vesicles From Gram-Negative Bacteria: A Solid Medium-Based Method Analysis Roberto G. M. Guardatti, Jhonatha R. Cordeiro‐Moura, Jhon J. Artunduaga Bonilla, Stefanie C. da Costa, Anderson S. Cabral, Bruna Montalvão, Flávia G. C. Reis, Leonardo Nimrichter, Márcio L. Rodrigues, Juliana Rizzo, Renata C. Picão Journal of Extracellular Biology, 2026 Extracellular vesicles (EVs) produced by gram‐negative bacteria have gained increased scientific interest over the last years due to a growing understanding of their roles in pathogenesis and biotechnological potential. However, isolating such nanoparticles is technically challenging. Here, we evaluated a rapid and reproducible protocol for EV isolation from bacteria grown on solid medium, specifically analyzing Gram‐negative species within the genera Acinetobacter , Aeromonas , Escherichia , Enterobacter , Klebsiella , and Pseudomonas . In addition to avoiding large culture volumes, the method was reproducible, relatively fast, compatible with multiple downstream analyses, including flow cytometry, TEM, and NTA, while yielding strain‐specific nanoparticle profiles with acceptable purity. Our results also show that Pseudomonas and Aeromonas strains were the highest and lowest EV producers, respectively. Although the method frequently co‐isolated filaments, clean isolates were also obtained. Comparison of EV isolation from cells grown on solid medium with those grown in liquid cultures showed that solid growth consistently yielded more nanoparticles. The methodology represents a valuable approach for bacterial EV isolation and may serve as an important initial step in EV research workflow.
Assessing antimicrobial resistance connectivity across One Health sectors Liguan Li, Bing Li, Xiaole Yin, Yu Xia, Yu Yang, Xiaoqing Xu, Tommy Tsan-Yuk Lam, Kenneth M. Y. Leung, Fujie Xu, Xuxiang Zhang, Jianhua Guo, Heike Schmitt, Gerard D. Wright, Janet Midega, Carlos Bezuidenhout, Renata C. Picão, Shaikh Z. Ahammad, Kornelia Smalla, Steven P. Djordjevic, Amy Pruden, Peter Vikesland, Dominic Frigon, Fiona Walsh, Thomas U. Berendonk, Gianluca Corno, Despo Fatta-Kassinos, Chang-Jun Cha, Nicholas Ashbolt, Guangguo Ying, Yi Luo, Yang Wang, Jianzhong Shen, Yu Zhang, Min Yang, Xiangdong Li, Baolan Hu, Lizhong Zhu, Yongning Wu, Song Tang, Biao Kan, Yong-Guan Zhu, Barth F. Smets, David Graham, Ryo Honda, Eddie Cytryn, Erica Donner, Per Halkjær Nielsen, Mark van Loosdrecht, Michael R. Gillings, William Gaze, Célia M. Manaia, Pedro J. J. Alvarez, Martin J. Blaser, Edward Topp, Tong Zhang Nature Water, 2025
Massive culture-based approach for the screening of AmpC, ESBL, and carbapenemase producers from rectal swabs Gabriel Taddeucci-Rocha, Victoria de Oliveira Costa, Sarah Vitória Martins da Silva, Jéssica Britto Gonçalves, Natalia Chilinque Zambão da Silva, Marcia Maiolino Garnica, Renata Cristina Picão Microbiology Spectrum, 2025 Gram-negative bacilli-producing beta-lactamases are major causes of difficult-to-treat infections, especially the AmpC, extended-spectrum beta-lactamases (ESBL), and carbapenemase types. Their spread within and outside hospital settings demands effective detection and monitoring in various environments, but current methods for this purpose often neglect important groups of beta-lactamases or are expensive and time consuming. We aimed to develop and test a massive culture approach to detect and differentiate between beta-lactamase producers from complex samples. The method includes enrichment on MacConkey agar supplemented with ceftriaxone to select for AmpC, ESBL, and carbapenemase producers, followed by replica plating under selective pressures (cefoxitin, cefepime, and imipenem) to differentiate them. The massive culture approach effectively differentiated strains producing different beta-lactamases in mixed cultures. In tests with rectal swabs, our method demonstrated 100% sensitivity, higher specificity, and greater accuracy for ESBL detection compared to the reference method. Additionally, it identified a broader spectrum of beta-lactamase producers, including AmpC and carbapenemase. The massive culture approach is a promising tool for detecting and differentiating gram-negative bacilli-producing beta-lactamases from rectal swabs. Due to the additional time required to produce results, this method is most suitable for central and research laboratories and enhances surveillance capabilities for antimicrobial resistance. IMPORTANCE The intestinal tract is a major reservoir of multidrug-resistant gram-negative bacilli, and surveillance of colonization is essential to understand resistance dissemination in both community and healthcare settings. However, standard culture-based methods typically target specific resistance mechanisms, often overlooking the coexistence of distinct beta-lactamase-producing strains within a single host. This limits our understanding of colonization dynamics and resistance evolution. To address this gap, we developed a culture-based approach that assesses the growth of strains under different selective pressures from a single rectal swab. By combining enrichment with replica plating, our method enables phenotypic discrimination between AmpC-, extended-spectrum beta-lactamases-, and carbapenemase-producing bacteria without depleting sample material across multiple media. Although validated for rectal swabs, the approach may be adapted for other complex samples such as urine, blood, soil, or water, expanding its utility in diverse clinical and environmental investigations. This strategy enhances the detection of diverse resistance profiles and supports a more comprehensive view of colonization and antimicrobial resistance dynamics.
Genomic analysis of blaNDM-1-carrying-Pseudomonas aeruginosa ST2407 in the chromosome from Brazil Ivson Cassiano de Oliveira Santos, Melise Chaves Silveira, Daiana Cristina Silva Rodrigues, Bruna Ribeiro Sued-Karam, Bruno Rocha Pribul, Giovanna de Oliveira Santos, Jônathas Dias Nunes, Marcos Dornelas-Ribeiro, Gabriela Bergiante Kraychete, Renata Cristina Picão, Elizabeth Andrade Marques, Robson Souza Leão, Cláudio Marcos Rocha-de-Souza, Ana Paula D'Alincourt Carvalho-Assef Infection Genetics and Evolution, 2024 Pseudomonas aeruginosa , an opportunistic pathogen often found in Healthcare-associated infections (HAI), has shown increased resistance to carbapenems (imipenem, meropenem, doripenem), the primary treatment options. We've seen a rise in carbapenemase-producing P. aeruginosa in Brazil, including NDM-producers. This study characterises an isolate carrying bla NDM-1 from a patient's skin fragment in a Brazilian hospital. The whole genomic sequence (WGS) of P. aeruginosa CCBH26428 was extracted and sequenced using Illumina and minION platforms. The assembly used MinION results mapped with Illumina reads, and annotation was performed by the RAST server. Resistance genes and clonality were identified using the CABGen platform. Additional information was carried out by manual annotation using Geneious software and BLAST tool. The genomic analysis revealed a genome of 6.995.008 bp and G+C 65.9 %. P. aeruginosa CCBH26428 belongs to ST2407. The bla NDM gene, associated with ISA ba 125, was found in a 63.862 pb genomic region flanked by IS26 insertion sequences. This region also contained the repA of the plasmid incompatibility group IncC2 and other resistance genes, suggesting it is a possible “translocation unit”. Additionally, 17 resistance genes, mutations in OprD and GyrA, and several virulence genes were detected, potentially exacerbating the infection. This study is report a WGS analysis of P. aeruginosa carrying bla NDM-1 in Brazil, highlighting the role of IS26 in the acquisition and spread of resistance genes between plasmids and chromosomes. • The Whole Genomic Sequence (WGS) of P. aeruginosa CCBH26428, which carries the bla NDM-1 gene, aids in comprehending its spread. • The bla NDM-1 gene, along with other resistance genes has been inserted into the genome of P. aeruginosa ST2407. This insertion is flanked by IS26, forming a structure known as a pseudo-compound transposon (PCT). • Various determinants of resistance and virulence highlight the potential emergence of clonal dissemination in P. aeruginosa ST2407.
Comparative analysis of the antimicrobial resistance and virulence traits in ESBL-producing-Klebsiella pneumoniae ST307 strains colonizing the gastrointestinal tract and causing a fatal bloodstream infection in a leukemia patient Luana Boff, Humberlânia de Sousa Duarte, Gabriela Bergiante Kraychete, Gabriel Taddeucci-Rocha, Bianca Diniz Oliveira, Rodolpho Mattos Albano, Ana Paula D'Alincourt Carvalho-Assef, Silvana Vargas Superti, Ianick Souto Martins, Renata Cristina Picão Infection Genetics and Evolution, 2024 Klebsiella pneumoniae is an opportunistic pathogen that can colonize the gastrointestinal tract (GIT) of humans. The mechanisms underlying the successful translocation of this pathogen to cause extra-intestinal infections remain unknown, although virulence and antimicrobial resistance traits likely play significant roles in the establishment of infections. We investigated K. pneumoniae strains isolated from GIT colonization (strains Kp_FZcol-1, Kp_FZcol-2 and Kp_FZcro-1) and from a fatal bloodstream infection (strain Kp_HM-1) in a leukemia patient. All strains belonged to ST307, carried a transferable IncF plasmid containing the blaCTX-M-15 gene (pKPN3–307 TypeA-like plasmid) and showed a multidrug-resistance phenotype. Phylogenetic analysis demonstrated that Kp_HM-1 was more closely related to Kp_FZcro-1 than to the other colonizing strains. The Kp_FZcol-2 genome showed an 81 % coverage with the Kp_HM-1 246,730 bp plasmid (pKp_HM-1), lacking most of the plasmid's putative virulence genes. Searching public genomes with similar coverage, we observed the occurrence of this deletion in K. pneumoniae ST307 strains recovered from human colonization and infection in different countries. Our findings suggest that strains lacking the putative virulence genes found in the pKPN3–307 TypeA plasmid are still able to colonize and infect humans, highlighting the need to further investigate the role of these genes for the adaptation of K. pneumoniae ST307 in distinct human body sites.
Green light for improving our understanding of AMR spread Gabriela B Kraychete, Raquel R Bonelli, Renata C Picão Lancet Microbe, 2024 The discovery and use of antibiotics brought about a groundbreaking transformation in the treatment of bacterial infections, substantially reducing the number of deaths associated with microorganism-related diseases in humans and animals. However, the emergence and spread of antimicrobial resistance (AMR) impacts the effectiveness of these drugs and threatens many achievements of modern medicine, justifying the urgent need to prevent its spread.1,2 The consistent detection of clinically relevant antimicrobial resistance genes and their bacterial hosts in humans, animals, and the environment suggests that One Health approaches are essential for tackling AMR.
Toward a Universal Unit for Quantification of Antibiotic Resistance Genes in Environmental Samples Xiaole Yin, Xi Chen, Xiao-Tao Jiang, Ying Yang, Bing Li, Marcus Ho-Hin Shum, Tommy T. Y. Lam, Gabriel M. Leung, Joan Rose, Concepcion Sanchez-Cid, Timothy M. Vogel, Fiona Walsh, Thomas U. Berendonk, Janet Midega, Chibuzor Uchea, Dominic Frigon, Gerard D. Wright, Carlos Bezuidenhout, Renata C. Picão, Shaikh Z. Ahammad, Per Halkjær Nielsen, Philip Hugenholtz, Nicholas J. Ashbolt, Gianluca Corno, Despo Fatta-Kassinos, Helmut Bürgmann, Heike Schmitt, Chang-Jun Cha, Amy Pruden, Kornelia Smalla, Eddie Cytryn, Yu Zhang, Min Yang, Yong-Guan Zhu, Arnaud Dechesne, Barth F. Smets, David W. Graham, Michael R. Gillings, William H. Gaze, Célia M. Manaia, Mark C. M. van Loosdrecht, Pedro J. J. Alvarez, Martin J. Blaser, James M. Tiedje, Edward Topp, Tong Zhang Environmental Science and Technology, 2023
Diversity of clonal types of Klebsiella pneumoniae causing infections in intensive care neonatal patients in a large urban setting Livia Helena Justo-da-Silva, Andrea Nunes De-Azeredo, Arnaldo Costa Bueno, Lara Feital Montezzi, Maria Beatriz Gerardin Poirot Leobons, Maria Silvana Alves, Patrícia de Souza Inhaquite, Rosana Rangel Santos, Valéria Brígido Carvalho Girão, Antônio José Ledo Alves da Cunha, Carmem Lucia Pessoa-Silva, Renata Cristina Picão, Cristina Barroso Hofer, Guilherme Santoro-Lopes, Lee Woodland Riley, Beatriz Meurer Moreira Brazilian Journal of Microbiology, 2019
Staphylococcus saprophyticus recovered from humans, food, and recreational waters in Rio de Janeiro, Brazil Viviane Santos de Sousa, Ana Paula de Souza da-Silva, Leif Sorenson, Raphael Paiva Paschoal, Renata Fernandes Rabello, Eloiza Helena Campana, Márcia Soares Pinheiro, Lyssa Oliveira Ferreira dos Santos, Natacha Martins, Ana Carolina Nunes Botelho, Renata Cristina Picão, Sérgio Eduardo Longo Fracalanzza, Lee Woodland Riley, George Sensabaugh, Beatriz Meurer Moreira International Journal of Microbiology, 2017
Occurrence of carbapenemase-producing bacteria in coastal recreational waters Lara Feital Montezzi, Eloiza Helena Campana, Laís Lisboa Corrêa, Livia Helena Justo, Raphael Paiva Paschoal, Isabel Lemos Vieira Dias da Silva, Maria do Carmo Maciel Souza, Marcia Drolshagen, Renata Cristina Picão International Journal of Antimicrobial Agents, 2015
Low prevalence of blaoxa-143 in private hospitals in Brazil Jéssica S. Werneck, Renata C. Picão, Raquel Girardello, Rodrigo Cayô, Vitor Marguti, Líbera Dalla-Costa, Ana C. Gales, Charline S. Antonio, Patrícia R. Neves, Micheli Medeiros, Elsa M. Mamizuka, Maria R. Elmor de Araújo, Nilton Lincopan Antimicrobial Agents and Chemotherapy, 2011
