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Given the urgent need for novel methods to control the spread of multidrug-resistant microorganisms, this study presents a green synthesis approach to produce silver nanoparticles (AgNPs) using the bark extract from Anadenanthera colubrina (Vell.) Brenan var. colubrina. The methodology included obtaining the extract and characterizing the AgNPs, which revealed antimicrobial activity against MDR bacteria. A. colubrina species is valued in indigenous and traditional medicine for its medicinal properties. Herein, it was employed to synthesize AgNPs with effective antibacterial activity (MIC = 19.53–78.12 μM) against clinical isolates from the ESKAPEE group, known for causing high hospitalization costs and mortality rates. Despite its complexity, AgNP synthesis is an affordable method with minimal environmental impacts and risks. Plant-synthesized AgNPs possess unique characteristics that affect their biological activity and cytotoxicity. In this work, A. colubrina bark extract resulted in the synthesis of nanoparticles measuring 75.62 nm in diameter, with a polydispersity index of 0.17 and an average zeta potential of −29 mV, as well as low toxicity for human erythrocytes, with a CC50 value in the range of 961 μM. This synthesis underscores its innovative potential owing to its low toxicity, suggesting applicability across several areas and paving the way for future research.

Abstract Foodborne diseases are a major challenge in the global food industry, especially those caused by multidrug-resistant (MDR) bacteria. Bacteria capable of biofilm formation, in addition to MDR strains, reduce the treatment efficacy, posing a significant threat to bacterial control. Bacteriophages, which are viruses that infect and kill bacteria, are considered a promising alternative in combating MDR bacteria, both in human medicine and animal production. Phage cocktails, comprising multiple phages, are commonly employed to broaden the host range and prevent or delay the development of phage resistance. There are numerous techniques and protocols available to evaluate the lytic activity of bacteriophages, with the most commonly used methods being Spot Test Assays, Efficiency of Plating (EOP), and infection assays in liquid culture. However, there is currently no standardization for which analyses should be employed and the possible differences among them in order to precisely determine the host range of phages and the composition of a cocktail. A preliminary selection using the Spot Test Assay resulted in four phages for subsequent evaluation against a panel of 36 Salmonella isolates of numerous serovars. Comparing EOP and infection assays in liquid culture revealed that EOP could underestimate the lytic activity of phages, directly influencing phage cocktail development. Moreover, the phage cocktail containing the four selected phages was able to control or remove biofilms formed by 66% (23/35) of the isolates, including those exhibiting low susceptibility to phages, according to EOP. Phages were characterized genomically, revealing the absence of genes associated with antibiotic resistance, virulence factors, or integrases. According to confocal laser scanning microscopy analysis, the biofilm maturation of one Salmonella isolate, which exhibited high susceptibility to phages in liquid culture and 96-well plates biofilm viability assays but had low values for EOP, was found to be inhibited and controlled by the phage cocktail. These observations indicate that phages could control and remove Salmonella biofilms throughout their growth and maturation process, despite their low EOP values. Moreover, using infection assays in liquid culture enables a more precise study of phage interactions for cocktail design timelessly and effortlessly. Hence, integrating strategies and techniques to comprehensively assess the host range and lytic activity of bacteriophages under different conditions can demonstrate more accurately the antibacterial potential of phage cocktails.

Pathogenic strains of Escherichia coli have acquired virulence factors, which confer an increased ability to cause a broad spectrum of enteric diseases and extraintestinal infections. The aim of this study was to analyze the antimicrobial resistance profile of and the presence of virulence-associated genes (VAGs) in E. coli fecal isolates from omnivores, vegetarians and vegans. A control group of 60 isolates from omnivores, as well as a study group with 41 isolates from vegetarians and 17 from vegans, were analyzed. Isolates from both groups showed a high rate of resistance to ampicillin, amoxicillin-clavulanic acid and nalidixic acid, and some of them were positive for the ESBL test (12% of isolates from vegetarians/vegans and 5% of isolates from omnivores). The most predominant VAGs detected in isolates from omnivores were fimH (70%), iutA (32%), fyuA (32%) and traT (32%), while among isolates from vegetarians or vegans, the most predominant were traT (62%), kpsMT k1 (28%) and iutA (22%). Most isolates from omnivores (55%) were positive for PAI I536, while most of those from vegetarians/vegans (59%) were positive for PAI IV536. Phylogenetic group A, composed of commensal non-pathogenic isolates that survive in the intestinal tract, was the most prevalent in both control and study groups. Some VAGs were found in only one of the groups, such as the pathogenicity island PAI III536, found in 12% of the isolates from omnivores, while the kpsMT III gene (15%) was detected only among isolates from vegetarians/vegans. Interestingly, this gene codes for a polysaccharide capsule found mainly in E. coli isolates causing intestinal infections, including EPEC, ETEC and EHEC. Finally, our results show that there were no advantages in vegetarian or vegan diets compared to the omnivorous diet, as in both groups we detected isolates harboring VAGs and displaying resistance to antibiotics, especially those most commonly used to treat urinary tract infections.

The demand for sustainable cosmetics leads to the search for natural and biotechnological ingredients. The present study reports the development of a multifunctional lip moisturizer containing levan (LEV) from Bacillus subtilis natto, sophorolipids (SOPs) from Starmerella bombicola and Citrus paradisi (OCP) essential oil, using a simplex-centroid experimental design. The formulations were evaluated physicochemically, pharmacotechnically and by DPPH assay. The optimized formulation was selected through the Response Surface Method, and the evaluation of its efficiency in lip hydration was carried out using the bioimpedance method and sensory analysis. The formulations showed pH compatibility with lips and remained stable after a centrifuge test and thermal stress. Spreadability varied between 415.3 and 1217.1 mm2, moisture retention was above 95% and antioxidant capacity was around 50% for all formulations. The optimized formulation, containing 0.4% LEV and 0.8% SOF, maintained the lip hydration already shown by the participants; 85% of them reported improvement in this aspect. For the first time, LEV and SOP were incorporated in lip moisturizers, which is an environmentally friendly product with marketing potential. Furthermore, the use of the Skin Analyzer Digital equipment, a low-cost and non-invasive technique, to evaluate the effectiveness of lip products is innovative; this methodology may help in the development of future cosmetology studies.

The prompt and accurate identification of the etiological agents of viral respiratory infections is a critical measure in mitigating outbreaks. In this study, we developed and clinically evaluated a novel melting-curve-based multiplex real-time PCR (M-m-qPCR) assay targeting the RNA-dependent RNA polymerase (RdRp) and nucleocapsid phosphoprotein N of SARS-CoV-2, the Matrix protein 2 of the Influenza A virus, the RdRp domain of the L protein from the Human Respiratory Syncytial Virus, and the polyprotein from Rhinovirus B genes. The analytical performance of the M-m-qPCR underwent assessment using in silico analysis and a panel of reference and clinical strains, encompassing viral, bacterial, and fungal pathogens, exhibiting 100% specificity. Moreover, the assay showed a detection limit of 10 copies per reaction for all targeted pathogens using the positive controls. To validate its applicability, the assay was further tested in simulated nasal fluid spiked with the viruses mentioned above, followed by validation on nasopharyngeal swabs collected from 811 individuals. Among them, 13.4% (109/811) tested positive for SARS-CoV-2, and 1.1% (9/811) tested positive for Influenza A. Notably, these results showed 100% concordance with those obtained using a commercial kit. Therefore, the M-m-qPCR exhibits great potential for the routine screening of these respiratory viral pathogens.

Escherichia coli is a key indicator of food hygiene, and its monitoring in meat samples points to the potential presence of antimicrobial-resistant strains capable of causing infections in humans, encompassing resistance profiles categorized as serious threats by the Centers for Disease Control and Prevention (CDC), such as Extended-Spectrum Beta-Lactamase (ESBL)—a problem with consequences for animal, human, and environmental health. The objective of the present work was to isolate and characterize ESBL-producing E. coli strains from poultry, pork, and beef meat samples, with a characterization of their virulence and antimicrobial resistance profiles. A total of 450 meat samples (150 chicken, 150 beef, and 150 pork) were obtained from supermarkets and subsequently cultured in medium supplemented with cefotaxime. The isolated colonies were characterized biochemically, followed by antibiogram testing using the disk diffusion technique. Further classification involved biofilm formation and the presence of antimicrobial resistance genes (blaCTX-M, AmpC-type, mcr-1, and fosA3), and virulence genes (eaeA, st, bfpA, lt, stx1, stx2, aggR, iss, ompT, hlyF, iutA, iroN, fyuA, cvaC, and hylA). Statistical analysis was performed via the likelihood-ratio test. In total, 168 strains were obtained, with 73% originating from chicken, 22% from pork, and 17% from beef samples. Notably, strains exhibited greater resistance to tetracycline (51%), ciprofloxacin (46%), and fosfomycin (38%), apart from β-lactams. The detection of antimicrobial resistance in food-isolated strains is noteworthy, underscoring the significance of antimicrobial resistance as a global concern. More than 90% of the strains were biofilm producers, and strains carrying many ExPEC genes were more likely to be biofilm formers (OR 2.42), which increases the problem since the microorganisms have a greater chance of environment persistence and genetic exchange. Regarding molecular characterization, bovine samples showed a higher prevalence of blaCTX-M-1 (OR 6.52), while chicken strains were more likely to carry the fosA3 gene (OR 2.43, CI 1.17–5.05) and presented between 6 to 8 ExPEC genes (OR 2.5, CI 1.33–5.01) compared to other meat samples. Concerning diarrheagenic E. coli genes, two strains harbored eae. It is important to highlight these strains, as they exhibited both biofilm-forming capacities and multidrug resistance (MDR), potentially enabling colonization in diverse environments and causing infections. In conclusion, this study underscores the presence of β-lactamase-producing E. coli strains, mainly in poultry samples, compared to beef and pork samples. Furthermore, all meat sample strains exhibited many virulence-associated extraintestinal genes, with some strains harboring diarrheagenic E. coli (DEC) genes.

Abstract During a microbiological and genomic surveillance study conducted to investigate the molecular epidemiology of extended-spectrum β-lactamase (ESBL)-producing Escherichia coli from community-acquired urinary tract infections (UTI) and commercial meat samples, in a Brazilian city with a high occurrence of infections by ESBL-producing bacteria, we have identified the presence of CTX-M (-2, -14, -15, -24, -27 and -55)-producing E. coli of international clones ST38, ST117, ST131 and ST354. The ST131 was more prevalent in human samples, and worryingly the high-risk ST131-C1-M27 was identified in human infections for the first time. We also detected CTX-M-55-producing E. coli ST117 from meat samples (i.e., chicken and pork) and human infections. Moreover, the clinically relevant CTX-M-24-positive E. coli ST354 clone was detected for the first time in human samples. In summary, our results highlight a potential of commercialized meat as a reservoir of high-priority E. coli lineages in the community, whereas the identification of E. coli ST131-C1-M27 indicates that novel pandemic clones have emerged in Brazil, constituting a public health issue.

Wound infections are feared complications due to their potential to increase healthcare costs and cause mortality since multidrug-resistant bacteria reduce treatment options. This study reports the development of a carbomer hydrogel containing biogenic silver nanoparticles (bioAgNPs) and its effectiveness in wound treatment. This hydrogel showed in vitro bactericidal activity after 2 h, according to the time–kill assay. It also reduced bacterial contamination in rat wounds without impairing their healing since the hydrogel hydrophilic groups provided hydration for the injured skin. The high number of inflammatory cells in the first days of the skin lesion and the greater degree of neovascularization one week after wound onset showed that the healing process occurred normally. Furthermore, the hydrogel-containing bioAgNPs did not cause toxic silver accumulation in the organs and blood of the rats. This study developed a bioAgNP hydrogel for the treatment of wounds; it has a potent antimicrobial action without interfering with cicatrization or causing silver bioaccumulation. This formulation is effective against bacteria that commonly cause wound infections, such as Pseudomonas aeruginosa and Staphylococcus aureus, and for which new antimicrobials are urgently needed, according to the World Health Organization’s warning.

Silver nanoparticles have been extensively researched and have several applications such as antimicrobial and anticancer, and can be used in the agricultural, biomedical, pharmaceutical, textile area, among others, but its toxicity is still little understood. Nanoparticles can measure between 1 – 100 nm and the most currently studied is the silver nanoparticle (AgNP). The main methods of synthesis are the chemical and the biogenic pathway, or green, which is less environmentally polluting, more environmentally friendly, and simpler, although standardization is more complex. Morphological and physical-chemical characteristics differ according to the synthesis method and, consequently, present different degrees of toxicity. Nanotoxicology studies the toxicity of nanoparticles on living organisms and scientists seek to know about the physical-chemical properties and their influence on interaction with the environment. It is known that there are several parameters that influence toxicity, such as dose, particle size, shape, morphology, surface chemistry, agglomeration/aggregation state, synthesis method, cell type and organism in which it is tested. Therefore, this article aims to address the main ways of synthesis of AgNPs, discuss the advantages and disadvantages of each method, the parameters that influence toxicity and examples of studies.

Pathogenic bacteria resistant to conventional antibiotics represent a global challenge and justify the need for new antimicrobials capable of combating bacterial multidrug resistance. This study describes the development of a topical hydrogel in a formulation composed of cellulose, hyaluronic acid (HA), and silver nanoparticles (AgNPs) against strains of Pseudomonas aeruginosa. AgNPs as an antimicrobial agent were synthesized by a new method based on green chemistry, using arginine as a reducing agent and potassium hydroxide as a carrier. Scanning electron microscopy showed the formation of a composite between cellulose and HA in a three-dimensional network of cellulose fibrils, with thickening of the fibrils and filling of spaces by HA with the presence of pores. Ultraviolet-visible spectroscopy (UV-vis) and particle size distribution for dynamic light scattering (DLS) confirmed the formation of AgNPs with peak absorption at ~430 nm and 57.88 nm. AgNPs dispersion showed a minimum inhibitory concentration (MIC) of 1.5 µg/mL. The time–kill assay showed that after 3 h of exposure to the hydrogel containing AgNPs, there were no viable cells, corresponding to a bactericidal efficacy of 99.999% in the 95% confidence level. We obtained a hydrogel that is easy to apply, with sustained release and bactericidal properties against strains of Pseudomonas aeruginosa at low concentrations of the agent.

American tegumentary leishmaniasis, a zoonotic disease caused by the Leishmania genus, poses significant challenges in treatment, including administration difficulty, low efficacy, and parasite resistance. Novel compounds or associations offer alternative therapies, and natural products such as oregano essential oil (OEO), extracted from Origanum vulgare, have been extensively researched due to biological effects, including antibacterial, antifungal, and antiparasitic properties. Silver nanoparticles (AgNp), a nanomaterial with compelling antimicrobial and antiparasitic activity, have been shown to exhibit potent leishmanicidal properties. We evaluated the in vitro effect of OEO and AgNp-Bio association on L. amazonensis and the death mechanisms of the parasite involved. Our results demonstrated a synergistic antileishmanial effect of OEO + AgNp on promastigote forms and L. amazonensis-infected macrophages, which induced morphological and ultrastructural changes in promastigotes. Subsequently, we investigated the mechanisms underlying parasite death and showed an increase in NO, ROS, mitochondrial depolarization, accumulation of lipid-storage bodies, autophagic vacuoles, phosphatidylserine exposure, and damage to the plasma membrane. Moreover, the association resulted in a reduction in the percentage of infected cells and the number of amastigotes per macrophage. In conclusion, our findings establish that OEO + AgNp elicits a late apoptosis-like mechanism to combat promastigote forms and promotes ROS and NO production in infected macrophages to target intracellular amastigote forms.

Considering the worrying emergence of multidrug resistance, including in animal husbandry and especially in food-producing animals, the need to detect antimicrobial resistance strains in poultry environments is relevant, mainly considering a One Health approach. Thus, this study aimed to conduct longitudinal monitoring of antimicrobial resistance in broiler chicken farms, with an emphasis on evaluating the frequency of resistance to fosfomycin and β-lactams. Escherichia coli was isolated from broiler chicken farms (cloacal swabs, meconium, poultry feed, water, poultry litter, and Alphitobius diaperinus) in northern Paraná from 2019 to 2020 during three periods: the first period (1st days of life), the second period (20th to 25th days of life), and third period (40th to 42nd days of life). Antibiogram tests and the detection of phenotypic extended-spectrum β-lactamase (ESBL) were performed, and they were confirmed by seaching for genes from the blaCTX–M group. The other resistance genes searched were mcr-1 and fosA3. Some ESBL blaCTX–M–1 group strains were selected for ESBL identification by sequencing and enterobacterial repetitive intergenic consensus-polymerase chain reaction analysis. To determine the transferability of the blaCTX–M–1– and fosA3-carrying plasmids, strains were subjected to conjugation experiments. A total of 507 E. coli were analyzed: 360 from cloacal swabs, 24 from meconium samples, 3 from poultry feed samples, 18 from water samples, 69 from poultry litter samples, and 33 from A. diaperinus samples. Among the strain isolate, 80% (406/507) were multidrug-resistant (MDR), and 51% (260/507) were ESBL-positive, with the blaCTX–M–1 group being the most frequent. For the fosA3 gene, 68% (344/507) of the strains isolated were positive, deserves to be highlighted E. coli isolated from day-old chickens (OR 6.34, CI 2.34–17.17), when compared with strains isolated from other origins (poultry litter, A. diaperinus, water, and poultry feed). This work alerts us to the high frequency of the fosA3 gene correlated with the CTX-M-1 group (OR 3.57, CI 95% 2.7–4.72, p < 0.05), especially the blaCTX–M–55 gene, in broiler chickens. This profile was observed mainly in day-old chicken, with a high percentage of E. coli that were MDR. The findings emphasize the importance of conducting longitudinal monitoring to detect the primary risk points during poultry production.

The present study aimed to evaluate the prevalence of antimicrobial resistance and clonal relationships in Proteus mirabilis isolated from chicken meat, beef, pork, and community-acquired urinary tract infections (UTI-CA). Chicken meat isolates showed the highest multidrug resistance (MDR), followed by those from pork and UTI-CA, whereas beef had relatively few MDR strains. All sources had strains that carried blaCTX-M-65, whereas blaCTX-M-2 and blaCMY-2 were only detected in chicken meat and UTI-CA isolates. This indicates that chicken meat should be considered an important risk factor for the spread of P. mirabilis carrying ESBL and AmpC. Furthermore, ESBL/AmpC producing strains were resistant to a greater number of antimicrobials and possessed more resistance genes than non-producing strains. In addition, the antimicrobial resistance genes qnrD, aac(6′)-Ib-cr, sul1, sul2, fosA3, cmlA, and floR were also found. Molecular typing showed a genetic similarity between chicken meat and UTI-CA isolates, including some strains with 100% similarity, indicating that chicken can be a source of P. mirabilis causing UTI-CA. It was concluded that meat, especially chicken meat, can be an important source of dissemination of multidrug-resistant P. mirabilis in the community.

Antimicrobial resistance is a significant public health problem, and the use of nanotechnology to control bacterial growth has increased, including multidrug-resistant ones. In this context, nanoparticles of different elements, mainly metals, such as silver, gold, and iron, have been extensively researched due to their efficient antimicrobial activity. Another element that stands out is selenium, which nanoparticles have high bioactivity and antibacterial action, although few studies report its use as a metallic nanoparticle. Therefore, the objective of this study was to perform the synthesis of selenium nanoparticles (SeNPs) from extracts of three plants: Allium cepa (onion), Malpighia emarginata (acerola), and Gymnanthemum amygdalinum (boldo). The synthesized nanoparticles were characterized by Dynamic Light Scattering (DLS), scanning electron microscopy (SEM), Fourier-Transform Infrared Spectroscopy (FTIR) and X-Ray Diffraction (DRX). Antibacterial activity was evaluated by microdilution in broth, followed by the time-kill curve analysis. The bacterial strains tested were Streptococcus agalactiae, Staphylococcus aureus, methicillin-resistant S. aureus, Pseudomonas aeruginosa, and Escherichia coli. Hemolytic activity was also determined. The synthesized SeNPs had a size between 245 and 321 ​nm and mostly spherical morphology. Antimicrobial activity against all Gram-positive bacteria tested was observed, with minimal inhibitory concentrations ranging from 6.125 to 98 ​μg/ml, but without action in the tested Gram-negative bacteria. Low hemolytic capacity was observed, with CC50 ranging from 0.82 to 2.1 ​mg/ml. The antimicrobial activity and low hemolytic concentration indicate the possibility of use against Gram-positive bacteria, including multidrug-resistant ones, opening a wide variety of options for their applications.

Pseudomonas aeruginosa possui grande importância para a saúde pública por estar presente no ambiente e nas infecções clínicas. O objetivo deste trabalho foi avaliar fenotipicamente os fatores de virulência motilidade, protease e ramnolipídeos em isolados clínicos e ambientais de P. aeruginosa. Vinte e cinco isolados clínicos e dez isolados ambientais foram analisados através de ensaios fenotípicos e categorizados em cepas não móveis, fracamente, moderadamente e altamente móveis; e produtores ou não de protease e ramnolipídeos. Em todos os ensaios, os isolados foram testados em triplicata em três dias diferentes. Os isolados ambientais produziram fatores de virulência como a motilidade (Swimming e Twitching) e Ramnolipídeos significativamente maior que os isolados clínicos. Este estudo nos alerta para o elevado nível de patogenicidade das cepas de P. aeruginosa, principalmente ambientais. A melhor compreensão da motilidade e produção de ramnolipídeos, fatores que estão diretamente associados à formação de biofilmes, pode favorecer estudos complementares às pesquisas visando o controle de bactérias patogênicas.

The increase in multidrug-resistant microorganisms represents a global threat requiring the development novel strategies to fight bacterial infection. This study aimed to assess the effect of silver nanoparticles (bio-AgNPs) on bacterial growth, biofilm formation, production of virulence factors, and expression of genes related to the quorum-sensing (QS) system of P. aeruginosa PAO1 and PA14. Biofilm formation and virulence assays were performed with bio-AgNPs. RT-qPCR was carried out to determine the effect of bio-AgNPs on the QS regulatory genes lasI, lasR, rhlI, rhlR, pqsA, and mvfR. Bio-AgNPs had an MIC value of 62.50 μM, for both strains. Phenotypic and genotypic assays were carried out using sub-MIC values. Experimental results showed that treatment with sub-MICs of bio-AgNPs reduced (p < 0.05) the motility and rhamnolipids and elastase production in P. aeruginosa PAO1. In PA14, bio-AgNPs stimulated swarming and twitching motilities as well as biofilm formation and elastase and pyocyanin production. Bio-AgNP treatment increased (p < 0.05) the expression of QS genes in PAO1 and PA14. Despite the different phenotypic behaviors in both strains, both showed an increase in the expression of QS genes. Demonstrating that the bio-AgNPs acted in the induction of regulation. The possible mechanism underlying the action of bio-AgNPs involves the induction of the rhl and/or pqs system of PAO1 and of the las and/or pqs system of PA14. These results suggest that exposure to low concentrations of bio-AgNPs may promote the expression of QS regulatory genes in P. aeruginosa, consequently inducing the production of virulence factors such as elastase, pyocyanin, and biofilms.

Biogenic silver nanoparticles (bio-AgNPs) are increasingly recognized as an antibiofilm and antivirulence strategy against P. aeruginosa, a bacterium that causes chronic infections in immunocompromised and cystic fibrosis patients. This study aimed to investigate the effects of subinhibitory concentrations of bio-AgNPs on motility and biofilm formation in P. aeruginosa. Bio-AgNPs were synthesized via reduction of ionic silver catalyzed by cell-free culture filtrate from Fusarium oxysporum. A total of 17 P. aeruginosa isolates and strains were evaluated for swarming, swimming, and twitching motility in the presence and absence (control) of bio-AgNPs, including 10 clinical isolates from patients with and without cystic fibrosis, 5 environmental isolates obtained from the public water supply system, and 2 reference strains (PAO1 and PA14). Isolates were identified by biochemical and molecular methods. Minimum inhibitory concentrations (MICs) were determined by the broth microdilution method. Swarming, swimming, and twitching motility assays were performed in Petri dishes. Biofilm formation capacity was assessed quantitatively by the crystal violet method. MIC values ranged from 15.62 to 62.50 µM. The results showed that subinhibitory concentrations of bio-AgNPs (½ MIC, 7.81–31.25 µM) significantly increased (p < 0.05) swarming, swimming, and twitching motility in 40.0, 40.0, and 46.7% of isolates, respectively. Subinhibitory bio-AgNP treatment enhanced (p < 0.05) biofilm formation capacity in PA14 and a cystic fibrosis isolate (P11). It is concluded that subinhibitory concentrations of bio-AgNPs increased biofilm formation and swarming, swimming, and twitching motility in PA14 and some P. aeruginosa isolates. These virulence factors are directly involved with quorum-sensing systems. Further research should investigate the effects of AgNPs on P. aeruginosa quorum sensing to help elucidate their mechanism of action at subinhibitory concentrations.

A patogenia de Salmonella em peixes é desconhecida e diversas são as formas de contaminação do pescado por este microrganismo, sem relatos que esclareçam sua função como agente etiológico ou como pertencente à microbiota destes animais. Em diversos países esta bactéria é responsável por milhões de casos de Salmonelose e as salmonelas paratíficas são as principais causadoras de doenças associadas ao consumo de peixes e crustáceos. Vários fatores podem influenciar na segurança dos alimentos da aquicultura, como o tipo das espécies cultivadas, localização dos criadouros, práticas de cultivo e manejo, preparo e consumo do pescado. Além disso, a escassez de saneamento básico ou disposição de rejeitos animais e humanos de forma inapropriada contribuem para a poluição desses ambientes. A contaminação do ecossistema aquático por Salmonella torna o ambiente uma fonte de disseminação desta bactéria. A crescente expansão da aquicultura como fonte de alimentos reforça a importância de um sistema de controle em todas as etapas necessárias para a produção e processamento de pescados. Através de práticas de manejo como o uso de temperaturas adequadas nas criações, uso controlado de antimicrobianos e adoção de medidas higiênico-sanitárias durante a produção e comercialização, é possível evitar a disseminação de enterobactérias na aquicultura. Apesar de ainda ser desconhecida sua importância para a sanidade do pescado, medidas de controle da qualidade da água e ração, higiene dos tanques e procedência de alevinos devem ser consideradas para prevenção de Salmonella em peixes uma vez que esta bactéria é um importante patógeno veiculado por alimentos.

This study discussed the use of antimicrobials in the commercial chicken production system and the possible factors influencing the presence of Extended-spectrum β-lactamase (ESBL)/AmpC producers strains in the broiler production chain. The aim of this study was to perform longitudinal monitoring of ESBL-producing and fosfomycin-resistant Escherichia coli from poultry farms in southern Brazil (Paraná and Rio Grande do Sul states) and determine the possible critical points that may be reservoirs for these strains. Samples of poultry litter, cloacal swabs, poultry feed, water, and beetles (Alphitobius sp.) were collected during three distinct samplings. Phenotypic and genotypic tests were performed for characterization of antimicrobial resistant strains. A total of 117 strains were isolated and 78 (66%) were positive for ESBL production. The poultry litter presented ESBL positive strains in all three sampled periods, whereas the cloacal swab presented positive strains only from the second period. The poultry litter represents a significant risk factor mainly at the beginning poultry production (odds ratio 6.43, 95% confidence interval 1–41.21, p < 0.05). All beetles presented ESBL positive strains. The predominant gene was blaCTX–M group 2, which occurred in approximately 55% of the ESBL-producing E. coli. The cit gene was found in approximately 13% of the ESBL-producing E. coli as AmpC type determinants. A total of 19 out of 26 fosfomycin-resistant strains showed the fosA3 gene, all of which produced ESBL. The correlation between fosA3 and blaCTX–M group 1 (blaCTX–M55) genes was significant among ESBL-producing E. coli isolated from Paraná (OR 3.66, 95% CI 1.9–9.68) and these genetic determinants can be transmitted by conjugation to broiler chicken microbiota strains. Our data revealed that poultry litter and beetles were critical points during poultry production and the presence of fosfomycin-resistant strains indicate the possibility of risks associated with the use of this antimicrobial during production. Furthermore, the genetic determinants encoding CTX-M and fosA3 enzymes can be transferred to E. coli strains from broiler chicken microbiota, thereby creating a risk to public health.

A presença de microrganismos patogênicos em produtos cárneos pode levar a doenças de origem alimentar e perdas econômicas aos seus produtores. Pequenas indústrias da região de Londrina, Paraná, produzem embutidos que são comercializados em feiras livres, pequenos mercados, bares e restaurantes da cidade. Embora essas indústrias sejam fiscalizadas pelo Serviço de Inspeção Municipal de Londrina, não existem dados sobre microrganismos patogênicos presentes nesses produtos. Os objetivos deste estudo foram pesquisar a presença de Salmonella spp. em amostras de linguiças produzidas e comercializadas na região de Londrina, Paraná, e identificar os genes de virulência eae, bfp, stx1, stx2, hlyA, ipaH, elt, est, aggR, aap e AA probe das cepas de E. coli isoladas dessas amostras. Quarenta e seis amostras de três tipos de linguiça (suína fresca, toscana e calabresa) produzidas por quatro diferentes produtores (marcas A, B, C e D) foram analisadas. Salmonella spp. foi isolada em 13 (28.3%) amostras e E. coli em 33 (71.3%) amostras. Das 13 amostras contaminadas com Salmonella spp., sete (53.8%) foram da marca A. A contaminação por Salmonella spp. foi maior nas amostras de linguiça toscana (8/17 – 41.7%) quando comparada com a contaminação encontrada nas amostras de linguiça suína (4/22 – 18.2%), independente da marca analisada. E. coli foi isolada de 12 das 13 amostras contaminadas com Salmonella spp. Uma amostra de linguiça calabresa estava contaminada com E. coli enteropatogênica clássica atípica do sorotipo O108:H9, que apresentava genes eae e hlyA. Os resultados obtidos sugerem contaminação da planta de processamento e ou das matérias primas utilizadas na fabricação das linguiças. Uma fiscalização mais adequada das indústrias se torna necessária para que as Boas Práticas de Fabricação sejam atendidas e, consequentemente, prevenida a contaminação do produto por bactérias patogênicas.

Salmonella spp. is one of the main agents responsible for foodborne infection in humans, and products of poultry origin are the most common infection sources. Studies have shown the occurrence of antimicrobials resistant Salmonella spp. in animal products. The Extended Spectrum ?-Lactamase (ESBL) are enzymes that confer to bacteria the ability to hydrolyze cephalosporin with an oximino side chain and monobactams. This study aimed to investigate antimicrobial resistance profile, identify phenotypes and genotypes for multiple drug resistance (MDR) and that produce ESBL from isolates of Salmonella spp. in the broiler production chain. We used samples of Salmonella spp. (n=11) isolates from poultry, poultry products and poultry-source environment from the state of Maranhão - Brazil. The isolates of Salmonella spp. assessed showed genotypical and phenotypical characteristics of MDR. The results show that 72.72% (08/11) of the strains presented the phenotypic profile for ESBL production. The isolates showed positivity to at least 13.64% (03/22) of the genes studied and the highest frequencies were observed in genes sul1 (73%), dfrA12 (55%), blaCTX-M (55%), tetA, tetB and tetC, with 45%. In conclusion, the strains of Salmonella spp. isolates present genotypic and phenotypic characteristics for MDR and ESBL production, demonstrating the dissemination risk of these microorganisms through the food chain.

Pathogenic Escherichia coli found in humans and poultry carcasses harbor similar virulence and resistance genes. The present study aimed to analyze the distribution of extraintestinal pathogenic E. coli (ExPEC) virulence factors (VF), blaCTX−M groups, fosA3, and mcr-1 genes in E. coli isolated from commercialized chicken carcasses in southern Brazil and to evaluate their pathogenic risk. A total of 409 E. coli strains were isolated and characterized for genes encoding virulence factors described in ExPEC. Results of antimicrobial susceptibility testing confirmed that the strains were resistant to β-lactams, fosfomycin, colistin, and others resistance groups. The highest prevalence of VFs was observed in isolates belonging to the CTX-M groups, especially the CTX-M-2 group, when compared to those in other susceptible strains or strains with different mechanisms of resistance. Furthermore, ESBL strains were found to be 1.40 times more likely to contain three to five ExPEC virulence genes than non-ESBL strains. Our findings revealed the successful conjugation between ESBL-producing E. coli isolated from chicken carcass and the E. coli recipient strain J53, which suggested that genetic determinants encoding CTX-M enzymes may have originated from animals and could be transmitted to humans via food chain. In summary, chicken meat is a potential reservoir of MDR E. coli strains harboring resistance and virulence genes that could pose serious risks to human public health.

Bacterial resistance to conventional antibiotics has become a clinical and public health problem, making therapeutic decisions more challenging. Plant compounds and nanodrugs have been proposed as potential antimicrobial alternatives. Studies have shown that oregano (Origanum vulgare) essential oil (OEO) and silver nanoparticles have potent antibacterial activity, also against multidrug-resistant strains; however, the strong organoleptic characteristics of OEO and the development of resistance to these metal nanoparticles can limit their use. This study evaluated the antibacterial effect of a two-drug combination of biologically synthesized silver nanoparticles (bio-AgNP), produced by Fusarium oxysporum, and OEO against Gram-positive and Gram-negative bacteria, including multidrug-resistant strains. OEO and bio-AgNP showed bactericidal effects against all seventeen strains tested, with minimal inhibitory concentrations (MIC) ranging from 0.298 to 1.193 mg/mL and 62.5 to 250 µM, respectively. Time-kill curves indicated that OEO acted rapidly (within 10 min), while the metallic nanoparticles took 4 h to kill Gram-negative bacteria and 24 h to kill Gram-positive bacteria. The combination of the two compounds resulted in a synergistic or additive effect, reducing their MIC values and reducing the time of action compared to bio-AgNP used alone, i.e., 20 min for Gram-negative bacteria and 7 h for Gram-positive bacteria. Scanning electron microscopy (SEM) revealed similar morphological alterations in Staphylococcus aureus (non-methicillin-resistant S. aureus, non-MRSA) cells exposed to three different treatments (OEO, bio-AgNP and combination of the two), which appeared cell surface blebbing. Individual and combined treatments showed reduction in cell density and decrease in exopolysaccharide matrix compared to untreated bacterial cells. It indicated that this composition have an antimicrobial activity against S. aureus by disrupting cells. Both compounds showed very low hemolytic activity, especially at MIC levels. This study describes for the first time the synergistic and additive interaction between OEO and bio-AgNP produced by F. oxysporum against multidrug-resistant bacteria, such as MRSA, and β-lactamase- and carbapenemase-producing Escherichia coli and Acinetobacter baumannii strains. These results indicated that this combination can be an alternative in the control of infections with few or no treatment options.

The aim of this research was to evaluate the effects of spray-dried plasma (SDP) on the performance, hematological profile, diarrhea frequency and economic viability of this ingredient in diets for light piglets at weaning, with 21 days of age. The trial was done using 72 pigs, 48 castrated male and 24 female (PenArLan), with an initial age of 21 days. The experimental design consisted of randomized blocks, with 4 treatments and 6 repetitions (each pen with two males and one female was considered a repetition). The treatments were: T1 - heavy weight weaned pigs, without supplementation; T2 - light weight weaned pigs, supplemented with 10g/animal/day of SDP; T3 - light weight weaned pigs, supplemented with 20g/animal/day of SDP; T4- light weight weaned pigs, without supplementation. The SDP were fed daily directly at the feeder three times per day. At 35 and 70 days old piglets were subjected to blood collection, and 8 animals in each treatment were used. Daily evaluation was made of diarrhea frequency. There was difference to feed:gain (P < 0,05) considering the period of 21 to 49 days of age and total period (21-70 days of age), with advantage to T3. The initial value for plasmatic urea was higher (P < 0,01) to pigs of T1 but the final titles were similar. There were no differences (P > 0,05) on frequency of diarrhea. There were differences in feed cost with advantage to T3 in the period of 21 to 35; 36 to 49 and 21 to 49 days of age. In total period (21 to 70 days of age) T3 showed the best index of economic efficiency.O objetivo desse trabalho foi avaliar o desempenho, perfil hematológico, freqüência de diarréia e potencial econômico de uso do plasma sanguíneo desidratado (PSD) na dieta de leitões desmamados com baixo peso, aos 21 dias de idade. Foram utilizados 72 leitões, sendo 48 machos castrados e 24 fêmeas (PenArLan) com idade média inicial de 21 dias. O delineamento experimental foi de blocos ao acaso, com 4 tratamentos e 6 repetições por tratamento (cada baia com dois machos e uma fêmea foi considerada uma repetição). Os tratamentos foram: T1 - animais pesados ao desmame, sem suplementação; T2 - animais leves ao desmame, suplementados com 10g/animal/dia de PSD; T3 - animais leves ao desmame, suplementados com 20g/animal/dia de PSD; T4 - animais leves ao desmame, sem suplementação. O PSD (10 e 20g/animal/dia) foi fornecido diretamente no cocho três vezes ao dia. Aos 35 e 70 dias de idade os leitões foram submetidos a coleta de sangue, sendo utilizado 8 animais de cada tratamento. Diariamente foi realizada a avaliação da freqüência de diarréia. Foi verificada diferença para conversão alimentar (P < 0,05) no período de tratamento compreendido entre 21 a 49 dias de idade e para o período experimental total (21 a 70 dias de idade) com vantagem para o T3. Os valores iniciais de uréia plasmática apresentaram diferença (P < 0,01) com vantagem para o T1, no entanto os valores finais mostraram-se semelhantes. Não houve diferença (P > 0,05) para freqüência de diarréia. Houve um melhor resultado na análise econômica, apontando vantagem para o T3 em relação ao custo da ração nos períodos compreendidos entre 21 a 35 dias, 36 a 49 dias e entre 21 a 49 dias de idade. Considerando todo o período experimental (21 a 70 dias de idade) o T3 apresentou melhor índice de eficiência econômica e de custo médio.

A proteína Iss (increased serum survival) é uma importante característica de resistência ao sistema complemento da Escherichia coli patogênica para aves (APEC). Os objetivos deste trabalho foram clonar e verificar a diversidade da seqüência do gene iss de APEC e caracterizar a proteína Iss recombinante. O gene iss de 309 bp foi amplificado por PCR, clonado e expresso na E. coli BL21 (DE3) utilizando o vetor pET SUMO. O gene iss da APEC9 foi classificado como iss tipo 1 pela diferenciação entre 3 tipos de iss alelos. A proteína Iss foi expressa pela indução com IPTG, purificada em coluna com resina ligada ao íon níquel e utilizada na imunização de galinhas poedeiras. Anticorpos da classe IgY anti rIss reagiram com a proteina rIss, a qual apresentou massa molecular de 22 kDa, correspondendo 11kDa da Iss e 11 kDa da proteína SUMO. The Iss (Increased serum survival) protein is an important characteristic of resistance to complement system of avian pathogenic Escherichia coli (APEC). The objectives of this work were to cloning and verify the sequence diversity of iss gene from APEC and characterize the recombinant Iss protein. The iss gene of 309 bp was amplified by PCR, cloned and expressed in E. coli BL21 (DE3) using the pET SUMO vector. The iss gene from APEC9 strain was classified as iss type 1 by differentiation of the three iss gene allele types. The protein was expressed by induction of IPTG and purified in resin charged with the nickel ion. Antibodies IgY anti rIss reacted with rIss showing a molecular mass of 22 kDa, corresponding 11KDa of Iss protein and 11 KDa SUMO protein.