Use of organic acids to reduce Salmonella Typhimurium excretion in swine / Uso de ácidos orgânicos para reduzir a excreção de Salmonella Typhimurium em suínos

The use of antimicrobials as growth promoters and disease prevention is being constantly reduced in several animal production systems, including in the swine industry. Therefore, this study aimed to evaluate the effectiveness of using acidifiers to control Salmonella Typhimurium in 65-day-old pigs by detecting the pathogen in organs at euthanasia. For this, 24 piglets were divided into two experimental groups consisting of 12 piglets each. An untreated control group (G1) and a treatment group (G2) received a liquid organic acidifier in the drinking water for 10 days (D-5 to D5). Five days after the start of treatment (D0), all piglets were challenged with 106 CFU of Salmonella Typhimurium and assessed for 12 days (D12). Every three days (D3, D6, D9, and D12), three animals from each experimental group were euthanized and then submitted for necropsy. Samples from the intestines (ileum, cecum, mesenteric lymph nodes, and ileocolic lymph nodes), liver, spleen, and lungs were collected to isolate Salmonella. The results show that, numerically, Salmonella isolation in the organs of G2 was lower than in G1 and that the number of positive cecum samples in G1 (66.7%; 8/12) was statistically different from the number of positive models in G2 (16.7%; 2/12), with a reduction of 28.6% of the total cecum positive samples in the treated group compared to the control. Therefore, it was observed that the liquid organic acidifier product could reduce the colonization of organs by Salmonella Typhimurium. (AU)

Use of organic acids to reduce Salmonella Typhimurium excretion in swine / Uso de ácidos orgânicos para reduzir a excreção de Salmonella Typhimurium em suínos

The use of antimicrobials as growth promoters and disease prevention is being constantly reduced in several animal production systems, including in the swine industry. Therefore, this study aimed to evaluate the effectiveness of using acidifiers to control Salmonella Typhimurium in 65-day-old pigs by detecting the pathogen in organs at euthanasia. For this, 24 piglets were divided into two experimental groups consisting of 12 piglets each. An untreated control group (G1) and a treatment group (G2) received a liquid organic acidifier in the drinking water for 10 days (D-5 to D5). Five days after the start of treatment (D0), all piglets were challenged with 106 CFU of Salmonella Typhimurium and assessed for 12 days (D12). Every three days (D3, D6, D9, and D12), three animals from each experimental group were euthanized and then submitted for necropsy. Samples from the intestines (ileum, cecum, mesenteric lymph nodes, and ileocolic lymph nodes), liver, spleen, and lungs were collected to isolate Salmonella. The results show that, numerically, Salmonellaisolation in the organs of G2 was lower than in G1 and that the number of positive cecum samples in G1 (66.7%; 8/12) was statistically different from the number of positive models in G2 (16.7%; 2/12), with a reduction of 28.6% of the total cecum positive samples in the treated group compared to the control. Therefore, it was observed that the liquid organic acidifier product could reduce the colonization of organs by Salmonella Typhimurium.(AU)

O uso de antimicrobianos como promotores de crescimento e prevenção de doenças vem sendo constantemente reduzido em diversos sistemas de produção animal, inclusive na suinocultura. Portanto, o objetivo do presente estudo foi avaliar a eficácia do uso de acidificantes no controle de Salmonella Typhimurium em suínos de 65 dias de idade, detectando o patógeno em órgãos após a eutanásia. Para isso, 24 leitões foram divididos em dois grupos experimentais constituídos por 12 leitões cada. Um grupo controle não tratado (G1) e um grupo de tratamento (G2) que recebeu um acidificante orgânico líquido na água de beber por 10 dias (D-5 a D5). Cinco dias após o início do tratamento (D0), todos os animais foram inoculados oralmente com 106 UFC de Salmonella Typhimurium e avaliados por 12 dias (D12). A cada três dias (D3, D6, D9 e D12), três leitões de cada grupo experimental foram eutanasiados e posteriormente submetidos à necropsia. Amostras de intestino (íleo, ceco, linfonodos mesentéricos e linfonodos ileocólicos), fígado, baço e pulmões foram coletadas para o isolamento de Salmonella. Os resultados mostram que, numericamente, o isolamento de Salmonella nos órgãos do G2 foi inferior ao G1, e que o número de amostras positivas de ceco no G1 (66,7%; 8/12) foi estatisticamente diferente do número de amostras positivas no G2 (16,7%; 2/12), com redução de 28,6% do total de amostras positivas de ceco no grupo tratado em relação ao controle. Portanto, observou-se que o ácido orgânico líquido foi capaz de reduzir a colonização de órgãos por Salmonella Typhimurium.(AU)

Multilocus sequence typing, clinical characteristic, and drug resistance of Salmonella typhimurium in 93 children / 中华实用儿科临床杂志

Objective:To investigate the multilocus sequence typing (MLST), clinical manifestations, and drug resistance of Salmonella typhimurium in children, thus providing references for the diagnosis, treatment, and prevention of pediatric Salmonella typhimurium infection.Methods:The clinical data of patients with Salmonella typhimurium serotypes confirmed by stool or blood culture between November 2017 and October 2020 were retrospectively collected from Fujian Maternity and Child Health Hospital.MSLT and drug susceptibility test were performed on Salmonella typhimurium isolated from the samples. Kruskal- Wallis test, Chi- square test, and Fisher′ s exact probability method were employed for data analyses. Results:Salmonella typhimurium was cultured from clinical samples of 96 children, of which, 93 samples were effective, including 92 stool samples and 1 blood sample from 53 boys(56.99%) and 40 girls(43.01%). The median age of disease onset occurred at 12.0 (8.5-22.0) months, with peak months of onset ranging from July to October.According to MLST classification, 93 children were divided into ST34 classification( n=58), ST19 classification( n=22) and other classification( n=13). Respiratory symptoms were significant different among MLST classification, and ST34 type Salmonella typhimurium enteritis was more commonly accompanied by respiratory symptoms ( χ2=17.657, P<0.001; Cramer V=0.421, P<0.001). There were significant differences in the drug sensitivity to Ampicillin ( χ2=8.774, P=0.033), Piperacillin tazobactam ( χ2=6.713, P=0.022), Ciprofloxacin ( χ2=20.780, P<0.001), Sulfamethoxazole ( χ2=15.364, P=0.001), Ampicillin sulbactam ( χ2=17.626, P=0.001) and Levofloxacin ( χ2=25.648, P<0.001) among 3 groups.No significant difference was found in the sensitivity to Ceftriaxone ( χ2=1.027, P=0.621), Ceftazidime ( χ2=7.637, P=0.059), Cefepime ( χ2=6.099, P=0.116) and Cefoperazone/Sulbactam ( χ2=2.405, P=0.649). All MLST types were sensitive to Imipenem and Meropenem. Conclusions:Salmonella typhimurium infection mainly affects infants, with the peak months of onset ranging from July to October.MLST34 is the main serotype, accompanied respiratory symptoms.Antibiotics should be selected according to drug sensitivity, and third-generation cephalosporins can be selected empirically without drug susceptibility test results.

Mutagenic Evaluation of Fluoxetine and Fluoxetine-Galactomannan Complex Through the Analysis of Chromosomal Aberrations in Human Peripheral Leukocytes and Salmonella typhimurium/Microssome Assay / Avaliação Mutagênica de Fluoxetina e Complexo Fluoxetina-Galactomanana Através da Análise de Aberrações Cromossômicas em Leucócitos Periféricos Humanos e em Salmonella typhimurium / Ensaio Microssômico

Objectives: The purpose of this study was to evaluate the mutagenic potential of fluoxetine and fluoxetine-galactomannan. Methods: Chromosomal aberration test and Salmonella typhimurium/microsome mutagenicity assay. Results: The results showed that fluoxetine (250 µg/mL) can cause chromosomal breaks of treated leukocytes and increase the frequency of reversion of the tester strains of S. typhimurium / microsome assay only at the highest concentration (5 mg/mL), while fluoxetine encapsulated in galactomannan did not cause these changes (leukocytes and S. typhimuriums strains). Conclusion: In summary, fluoxetine showed a mutagenic effect detectable only at high concentrations in both eukaryotic and prokaryotic models. Furthermore, the fluoxetine/galactomannan complex, in this first moment, prevented the mutagenicity attributed to fluoxetine, emphasizing that the present encapsulation process can be an alternative in preventing these effects in vitro.

Objetivos: avaliar o potencial mutagênico da fluoxetina e da fluoxetina-galactomanana. Métodos: Teste de aberração cromossômica e ensaio de mutagenicidade de Salmonella typhimurium /microssoma. Resultados: a fluoxetina (250 µg/mL) pode causar quebras cromossômicas de leucócitos tratados e aumentar a frequência de reversão das cepas testadoras de S. typhimurium /microssoma apenas na concentração mais alta (5 mg/mL), enquanto a fluoxetina encapsulada em galactomanano não causou essas alterações (leucócitos e cepas de S. typhimurium). Conclusão: a fluoxetina mostrou um efeito mutagênico detectável apenas em altas concentrações em modelos eucarióticos e procarióticos. Além disso, o complexo fluoxetina/galactomanan, neste primeiro momento, evitou a mutagenicidade atribuída à fluoxetina, ressaltando que o presente processo de encapsulamento pode ser uma alternativa na prevenção desses efeitos in vitro.

In Silico Modeling and Docking Analysis of CTX-M-5, Cefotaxime-Hydrolyzing ?-Lactamase from Human-Associated Salmonella Typhimurium

Objective: To computationally model the CTX-M-5 ?-lactamase and establish its structure, which is exclusively present in human-associated Salmonella. Methods: The CTX-M-5 aminoacid sequence (Uniprot ID:O65975) of Salmonella enterica subsp. enterica serovar typhimurium was retrieved from UniProt database and subjected to homology modeling using MODELLER 9v7. The homology models were duly validated using RAMPAGE tool by generating Ramachandran plots, ERRAT graphs, and ProSA score. DoGSiteScorer server and ConSurf server were used to detect the cavities, pockets, and clefts to identify conserved amino acid sites in the predicted model. Subsequently, the modeled structure was docked using CLC Drug Discovery Workbench against proven drugs and known inhibitors. Results: Obtained high-quality homology model with 91.7% of the residues in favorable regions in Ramachandran plot and qualified in other quality parameters. Docking studies resulted in a higher dock score for PNK (D-benzylpenicilloic acid) molecule when compared to other reported inhibitors. Conclusion: This in silico study suggests that the compound PNK could be an efficient ligand for CTX-M-5 ?-lactamase and serve as a potent inhibitor of CTX-M-5.

Capability of acidic electrolyzed water in the elimination of Salmonella Typhimurium and Escherichia coli in the chicken breast

Aims@#This study aimed to investigate the effect of acidic electrolyzed water (AEW) as pre-refrigeration and pre-freezing processing steps for chicken meat in regard to the behavior of S. Typhimurium and E. coli during storage.@*Methodology and results@#AEW (free available chlorine 30 ppm and pH 2.7) was tested against S. Typhimurium and E. coli in growth media (brain heart infusion broth) and by exposing inoculated chicken fillets. The in vitro study appointed 10 minutes as the straightening exposure time of fresh prepared AEW for S. Typhimurium and E. coli. The reduction effect of AEW was significant (p<0.05) for both S. Typhimurium and E. coli along the 8 days of refrigerated storage with a maximum reduction after 24 h of post-treatment reaching 23.3% (1.4 log CFU/g) and 32.43% (2.15 log CFU/g) for S. Typhimurium and E. coli, respectively. AEW resulted in a significant reduction (p<0.05) as a pre-freezing application for both microorganisms, where the maximum reductions of 20% (1.2 log CFU/g) and 31.84% (2.14 log CFU/g) for S. Typhimurium and E. coli, respectively, were reported at zero time (just after dipping). In exposed samples to AEW, S. Typhimurium could not be detected by the 6th week of frozen storage while E. coli continued detectable until till 10th week but with a reduced population of 30% compared to control.@*Conclusion, significance and impact of study@#The findings of the present study suggest the application of AEW as a pre-refrigeration and pre-freezing treatment for chicken products. AEW application significantly improved the safety of chicken products.

Antimicrobial resistance and molecular typing of Salmonella typhimurium in Wuxi city, 2011-2018 / 中华检验医学杂志

Objective:Analysis of the antibiotic resistance profiles and molecular typing of Salmonella typhimurium ( S. typhimurium) isolated in Wuxi city from 2011 to 2018. Methods:A total of 109 S. typhimurium isolates were detected from different types monitoring samples in Wuxi city from 2011 to 2018. Microbroth dilution method was used to test antimicrobial susceptibility of S. typhimurium for 17 antimicrobial agents. Pulsed field gel electrophoresis (PFGE) was used to conduct molecular typing. Statistical analysis was carried out using Chi-square test. Results:Tetracycline-resistance and ampicillin-resistance were most frequency in 109 S. typhimurium isolates, 69.72% (76/109) and 68.81% (75/109), respectively. For compound sulfamethoxazole, chloramphenicol, ciprofloxacin, ampicillin/sulbactam, azithromycin, nitrofurantoin, cefotaxime and aztreonam, the resistance rates were 23.85%(26/109), 22.02%(24/109), 11.93%(13/109), 4.59%(5/109), 3.67%(4/109), 3.67%(4/109), 0.92%(1/109), 0.92%(1/109), respectively. All isolates were susceptible to amikacin, ertapenem, imipenem, meropenem, ceftazidime and ceftazidime-avibactam. Azithromycin-resistance isolates were decreasing year by year gradually. The aztreonam and cefotaxime-resistant isolates were found since 2018, while chloramphenicol and compound sulfa-resistant isolates showed upward trend simultaneously. Conclusions:S. typhimurium in Wuxi city exhibited highly resistant to tetracycline and ampicillin. The significant variability existed between genotype and phenotype of S. typhimurium.

Antimicrobial activity and chemical characterization of Laurel essential oil / Atividade antimicrobiana e caracterização química do óleo essencial de Louro / Actividad antimicrobiana y caracterización química del aceite esencial de Laurel

The factors that arouse interest in the study of essential oils as biocidal agents are numerous, such as the fact that they have antibacterial, antifungal, insecticidal, antioxidant, anti-inflammatory and larvicidal properties. The objective of this work was to evaluate the antimicrobial activity, in vitro, of the laurel (Laurus nobilis L) essential oil on the growth of pathogenic bacteria Salmonella enterica serovar Typhimurium ATCC 14028 and Staphylococcus aureus ATCC 25923, at different exposure times, as well as to perform the chemical characterization. Twenty compounds were identified and quantified, representing 96.57% of the total composition. The class of oxygenated monoterpenes represented the majority class of the essential oil, with 1,8-cineol (33.8%) as the substance found in greater quantity, followed by linalool (17.79%). The third constituent in greater quantity was sabinene (12.23%), belonging to the group of monoterpene hydrocarbons. Terpinyl acetate (9.41%) was also considered to be quantitatively representative. Laurel essential oil showed bacteriostatic activity against S. Typhimurium ATCC 14028 and S. aureus ATCC 25923.(AU)

Os fatores que despertam interesse no estudo dos óleos essenciais como agentes biocidas são inúmeros, como o fato de possuírem propriedades antibacteriana, antifúngica, inseticida, antioxidante, antiinflamatória e larvicida. O objetivo deste trabalho foi avaliar a atividade antimicrobiana, in vitro, do óleo essencial de louro (Laurus nobilis L) sobre o crescimento das bactérias patogênicas Salmonella enterica sorovar Typhimurium ATCC 14028 e Staphylococcus aureus ATCC 25923, em diferentes tempos de exposição, assim como realizar a caracterização química do óleo. Vinte compostos foram identificados e quantificados, representando 96,57% da composição total. A classe dos monoterpenos oxigenados representou a classe majoritária do óleo essencial, sendo o 1,8-cineol (33,8%) a substância encontrada em maior quantidade, seguido do linalol (17,79%). O terceiro constituinte em maior quantidade foi o sabineno (12,23%), pertencente ao grupo dos hidrocarbonetos monoterpênicos. O acetato de terpinila (9,41%) também foi considerado quantitativamente representativo. O óleo essencial de louro apresentou atividade bacteriostática contra S. Typhimurium ATCC 14028 e S. aureus ATCC 25923.(AU)

Los factores que despiertan interés en el estudio de los aceites esenciales como agentes biocidas son innumerables, como el hecho de que tienen propiedades antibacterianas, anti fúngicas, insecticidas, antioxidantes, antiinflamatorias y larvicidas. El objetivo de este trabajo ha sido evaluar la actividad antimicrobiana, in vitro, del aceite esencial de laurel (Laurus nobilis L) sobre el crecimiento de bacterias patógenas Salmonella enterica serovar Typhimurium ATCC 14028 y Staphylococcus aureus ATCC 25923, en diferentes momentos de exposición, así como realizar la caracterización química del aceite. Se identificaron y cuantificaron veinte compuestos, que representan el 96,57% de la composición total. La clase de mono terpenos oxigenados representó la clase principal de aceite esencial, siendo el 1,8-cineol (33,8%) la sustancia que se encuentra en mayor cantidad, seguida del linalol (17,79%). El tercer constituyente en mayor cantidad fue el sabineno (12,23%), perteneciente al grupo de los hidrocarburos monoterpénicos. El acetato de terpinilo (9,41%) también se consideró cuantitativamente representativo. El aceite esencial de laurel mostró actividad bacteriostática contra S. Typhimurium ATCC 14028 y S. aureus ATCC 25923.(AU)

Caracterización molecular y fenotípica de aislamientos clínicos de Salmonella Typhimurium variante monofásica (1, 4, [5],12: i: -) recuperados en Colombia / Molecular and phenotypic characterization of Salmonella Typhimurium monophasic variant (1, 4, [5],12: i: -) from Colombian clinical isolates

Resumen: Introducción. La variante monofásica (1,4,[5],12:i:-) de Salmonella Typhimurium ocupa los primeros lugares en los programas de vigilancia de Salmonella a nivel mundial. En Colombia, Salmonella enterica variante monofásica alcanza el cuarto lugar en cuanto a los aislamientos clínicos recuperados por medio de la vigilancia por laboratorio del Grupo de Microbiología del Instituto Nacional de Salud, pero se desconoce si dichos aislamientos están relacionados con la variante monofásica de Typhimurium que circula a nivel global, y con sus características genéticas y fenotípicas. Objetivo. Caracterizar los aislamientos de Salmonella monofásica recuperados en Colombia entre el 2015 y el 2018 por el Grupo de Microbiología del Instituto Nacional de Salud. Materiales y métodos. Se analizaron 286 aislamientos clínicos de Salmonella enterica variante monofásica mediante PCR o secuenciación del genoma completo (Whole Genome Sequencing, WGS) para confirmar si correspondían a Salmonella Typhimurium variante monofásica, en tanto que, en 54 aislamientos, se determinó la estructura genética del operón que codifica la segunda fase flagelar y, en 23, se evaluó la motilidad, el crecimiento y la expresión de las proteínas de membrana externa. Resultados. El 61 % (n=174) de los aislamientos de Salmonella monofásica correspondió a Salmonella Typhimurium serovar monofásico. El 64,8 % (n=35/54) se relacionó con el clon europeo-español y, el 13 % (n=7/54), con el estadounidense. En dos aislamientos de orina se encontró una diferencia significativa en la motilidad y el crecimiento, así como ausencia de la porina OmpD en medio mínimo M9. Conclusiones. En el periodo de estudio, circuló en Colombia la variante monofásica de Salmonella Typhimurium relacionada con el clon europeo-español, y se registró ausencia total del operón fljAB. Los resultados evidenciaron cambios fenotípicos en los aislamientos provenientes de muestras de orina que sugieren adaptación en procesos invasivos.

Abstract: Introduction. The Salmonella Typhimurium monophasic variant (1,4,[5],12:i:-) is currently the most commonly detected variant in Salmonella surveillance programs worldwide. In Colombia, the Salmonella enterica monophasic variant is the fourth most common clinical isolate recovered through the laboratory surveillance of the Grupo de Microbiología from the Instituto Nacional de Salud; however, it is unknown whether these isolates are closely related to the monophasic Typhimurium variant, which circulates globally, and their genetic and phenotypic characteristics have not been reported. Objective. To characterize monophasic Salmonella enterica isolates identified in Colombia from 2015 to 2018 by the Instituto Nacional de Salud. Materials and methods. Two hundred eighty-six clinical isolates of the monophasic Salmonella enterica variant were analyzed by PCR or whole-genome sequencing to confirm whether they corresponded to the Salmonella Typhimurium monophasic variant while the genetic structure of the operon encoding the second flagellar phase was determined in 54 isolates. Motility, growth, and expression of the outer membrane proteins were evaluated in 23 isolates. Results. During the study period in Colombia, 61% (n=174) of Salmonella monophasic isolates belonged to Salmonella Typhimurium serovar monophasic (1,4,[5],12:i-). Of these, 64.8% (n=35/54) were related to the European/Spanish clone and 13% (n=7/54) to the U.S. clone. Two isolates recovered from urine samples showed differences in motility, growth, and the absence of the OmpD porin in M9 minimal medium. Conclusions. Most of the monophasic Salmonella Typhimurium variants that have circulated in Colombia since 2015 lacked the second phase of operon fljAB, which is related to the European/Spanish clone. The results evidenced phenotypic changes in urine samples suggesting bacterial adaptation in the case of these invasive samples.

Expressão e atividade de sistemas de dois componentes (TCS) de Salmonella enterica sorovar Typhimurium no contexto do metaboloma intestinal humano / Expression and activity of two-component systems of Salmonella enterica sorovar Typhimurium in the context of the human intestinal metabolome

A interação entre membros do microbioma intestinal humano, células hospedeiras e patógenos invasores pode ocorrer de diversas formas, sendo uma delas através de pequenas moléculas chamadas metabólitos. A percepção e resposta efetiva de um microrganismo às diferentes condições encontradas em seu ambiente, incluindo metabólitos produzidos por outros microrganismos, são fatores importantes para sua adaptação, sobrevivência e disseminação. Os sistemas de dois componentes (TCS) permitem a percepção e resposta a mudanças ambientais, regulando a expressão de genes específicos. Nosso grupo mostrou anteriormente que um extrato orgânico de fezes humanas (EF), bem como o ácido 3,4-dimetilbenzoico (3,4-DMB), encontrado no EF, inibe a capacidade de Salmonella enterica sorovar Typhimurium de invadir células hospedeiras. O presente trabalho propôs investigar o impacto do microbioma intestinal humano, bem como de pequenas moléculas produzidas por Clostridium citroniae (membro deste microbioma) na expressão e atividade dos genes de TCS de Salmonella. Os metabólitos de EF e de culturas puras de C. citroniae foram extraídos com acetato de etila e adicionados a meio de cultura. O pH do meio foi ajustado (~ 7,4) e a solução foi esterilizada por filtragem. Salmonella foi cultivada na presença ou ausência do EF e do extrato de C. citroniae, bem como do ácido 3,4-DMB, em condições aeróbias e anaeróbias, até alcançar o meio da fase logarítmica de crescimento. O RNA foi extraído para a realização de PCR em Tempo Real utilizando iniciadores direcionados a quase todos os TCS de Salmonella. Nossos resultados mostraram que vários genes de TCS envolvidos na virulência de Salmonella (SsrAB, EnvZ-OmpR, QseCB, PhoQP, TorSR, TtrRS) foram regulados diferencialmente por esses metabólitos, tanto em condições aeróbias quanto anaeróbias. EnvZ-OmpR, PhoPQ e SsrAB estão diretamente envolvidos na regulação das Ilhas de Patogenicidade 1 e 2 de Salmonella. QseCB é crucial para a detecção de quorum em Salmonella, de hormônios hospedeiros e para a regulação da motilidade (swimming). Vários outros TCS também foram regulados, incluindo TorSR e TtrRS, envolvidos na regulação da respiração anaeróbica de N-óxido de trimetilamina (TMAO) e tetrationato, respectivamente. Esses compostos são importantes para a sobrevivência de Salmonella no ambiente anaeróbico do intestino humano. Nossos resultados de avaliação de expressão gênica global de Salmonella cultivada na presença de ácido 3,4-DMB (aerobiose e anaerobiose) bem como na presença do EF em anaerobiose, mostraram que genes condificados em SPI-1 e SPI-2, SPI-4 e alguns genes do TCS foram reprimidos, enquanto genes marR, marB e marA foram ativadas nessas condições. Adicionalmente, comparamos nossos resultados de RNAseq, de Salmonella cultivada na presença do ácido 3,4-DMB em aerobiose, com resultados disponíveis da base de dados Salmonella Compendium. Ainda, a capacidade de Salmonella de adentrar e sobreviver dentro de células fagocíticas (macrófagos RAW 264.7) parece ser afetada pelas três condições testadas neste trabalho. Nossos resultados mostram que importantes vias de sinalização da virulência de Salmonella podem ser moduladas pelos metabólitos presentes no microbioma intestinal humano e abrem caminhos para novas pesquisas sobre a sinalização intercelular microbioma-patógeno no ambiente intestinal.

The interaction between members of the human gut microbiome, host cells and invading pathogens often occurs through small molecules, also called metabolites. The perception and effective response of a microorganism to the different conditions found in its environment, including metabolites produced by other microbes, is important for its adaptation, survival and dissemination. Two-component systems (TCS) allow the perception and response to environmental changes by regulating the expression of specific genes. Our group previously showed that organic extracts of human feces (EF) as well as the specific metabolite 3,4-dimethylbenzoic acid (3,4-DMB) found within the EF, inhibit the ability of Salmonella enterica sorovar Typhimurium to invade host cells. In the present work, we investigated the impact of the human gut microbiome as well as small molecules produced by Clostridium citroniae (a member of this microbiome) on the expression and activity of Salmonella TCS genes. Metabolites (from feces or C. citroniae cultures) were extracted using ethyl acetate and added to culture medium. The pH of the medium was adjusted (~7.4), and the solution was filter sterilized. Salmonella was grown in the presence or absence of the organic extracts as well as 3,4-DMB acid under aerobic and anaerobic conditions until it reached mid-log growth. RNA was then extracted for Real-time PCR using primers targeting almost all Salmonella TCS. Our results showed that several TCS involved in Salmonella virulence (SsrAB, EnvZ-OmpR, QseCB, PhoQP, TorSR, TtrRS) were differentially regulated by these metabolites both in aerobic and anaerobic conditions. EnvZ-OmpR, PhoPQ, and SsrAB are directly involved in the regulation of Salmonella Pathogenicity Islands 1 and 2. QseCB is crucial for Salmonella =quorum sensing, sensing of host hormones and regulation of swimming motility. Several other TCS were also regulated, including TorSR and TtrRS, which are involved in the anaerobic respiration of trimethylamine N-oxide (TMAO) and tetrathionate, respectively. These compounds are important for Salmonella survival in the anaerobic environment of the human gut. Our results of the evaluation of global Salmonella gene expression grown in the presence of 3,4-DMB acid (aerobiosis and anaerobiosis) as well as in the presence of EF in anaerobiosis, showed that genes encoded in SPI-1 and SPI-2, SPI-4 and some TCS genes have been repressed, while multiple drug resistance genes, as well marR, marB and marA genes have been activated under these conditions. Besides, we compared our results of RNAseq, Salmonella was grown in the presence of 3,4-DMB acid in aerobiosis, with results available from the Salmonella Compendium database. Also, Salmonella's ability to enter and survive within phagocytic cells (macrophages RAW 264.7) appears to be affected by the three conditions tested in this work. Our results show that important Salmonella virulence signalling pathways can be modulated by the metabolites present in the human intestinal microbiome and open the way for further research on the microbiome-pathogen intercellular signalling in the intestinal environment.

Anti-inflammatory actions and Salmonella typhimurium-bacteraemia clearance by methanol extract of Curcuma longa Linn. (Turmeric)

Aims@#Rhizome of turmeric is known to possess therapeutic activities and has been used in medical practice as an anti-diabetic, hypolipidemic, hepatoprotective, anti-diarrheal, and anti-asthma agent. This study was designed to investigate the anti-inflammatory and antimicrobial activities of Curcuma longa. @*Methodology and results@#Rhizomes of Curcuma longa (Turmeric) purchased from markets in Ado-Ekiti, Nigeria, were analysed for anti-inflammatory and anti-microbial properties, as well as phytochemical constituents. The in vitro anti-inflammatory activities of the C. longa methanol extract (CLME) were evaluated by albumin denaturation, proteinase inhibitory activity, membrane stabilization, and anti-lipoxygenase activity, at different concentrations using Aspirin, Diclofenac sodium and Indomethacin as standard drugs. The in vitro antimicrobial activities of CLME were carried out on five pathogenic microbes namely Escherichia coli ATCC 29929, Staphylococcus aureus ATCC 29293, Salmonella typhimurium ATCC 14028, Klebsiella pneumoniae ATCC 4252 and Candida albicans ATCC 10231, using both agar well diffusion and broth dilution techniques. A S. typhimurium infected rat model was used for in vivo antimicrobial studies. Phytochemical analyses showed that C. longa rhizomes contain high concentrations of alkaloids, flavonoid and saponins, with moderate levels of phenols, tannin and ferric reducing antioxidant power. CLME was found to be rich in alkaloids, tannins, phenols, steroids, saponins, terpenoids, flavonoids and reducing sugars. CLME showed potent anti-inflammatory activities, and the results compared favourably with the standard anti-inflammatory drugs used. C. longa methanol extract significantly inhibited albumin denaturation and proteinase activity, stabilized membrane of red blood cell from haemolysis in heat and hypotonic conditions, as well inhibited lipoxygenase activity; all of which are associated with inflammatory processes. CLME was found to possess high in vitro antimicrobial activities against the five microorganisms tested. Rats orally infected with S. typhimurium, demonstrated bacteraemia five days post infection, with a total clearance of bacteraemia within 3-5 days following oral administration of CLME. The infected rats treated with CLME equally showed significant improvement in some haematological indices compared to infected rats that were not treated with CLME. @*Conclusion, significance and impact of study@#The results also showed that methanol extract of C. longa rhizome effectively cured with S. typhimurium infected rats. The overall results suggest that Curcuma longa is a potential source of anti-inflammatory and antimicrobial agents.

Construction and characterization of △sseK2 mutant strain of Salmonella typhimurium SL1344 / 中华微生物学和免疫学杂志

Objective To analyze the biological characteristics of a mutant strain of Salmonella ty-phimurium SL1344 with sseK2-deletion (SL1344△sseK2) in order to provide reference for further study of safe and effective live vaccines. Methods The mutant strain SL1344△sseK2 with a deletion of 1047 bp in sseK2 gene was constructed through a two-step allelic exchange using recombinant suicide plasmid. Its com-plemented strain, SL1344C△sseK2, was also constructed. Biological and immunological characteristics of the mutant strain were detected. Results PCR, double-enzyme digestion and sequencing analysis showed that the mutant strain SL1344△sseK2 and the complemented strain SL1344C△sseK2 were successfully con-structed. The serotype of the mutant strain was 1,4,[5],12:i:1,2, identical to the parent strain SL1344. In addition, the mutant strain showed no significant change in biochemical characteristics or growth rate and was genetically stable in vitro. Compared with the parent strain SL1344, the virulence of SL1344△sseK2 was attenuated in BALB/ c mice. The median lethal dose of SL1344△sseK2 for 6-week-old BALB/ c mice was 3. 44×108 colony-forming units (CFU), which was 1620 times lower than that of SL1344. Oral immuniza-tion with SL1344△sseK2 protected 62. 5％ of the mice against challenge with wild Salmonella typhimurium strains on 17 d after vaccination. The levels of serum IgG antibody peaked on 14 d after immunization. No significant difference in biological characteristics was observed between the complemented and the parent strains, indicating that the mutant strain was basically complemented to the wild-type strain.Conclusions The mutant strain SL1344△sseK2 was constructed successfully and genetically stable with sig-nificantly attenuated virulence and good immunogenicity. This study suggested that sseK2 gene played an im-portant role in regulating the virulence of SL1344, which might provide reference for further study of its func-tion and for assessing its potential as a candidate live attenuated vaccine.

Molecular typing, virulence genes and drug resistance of Salmonella typhimurium in Longyan city / 中华微生物学和免疫学杂志

Objective@#To analyze the molecular typing characteristics by pulsed field gel electrophoresis (PFGE), drug resistance and virulence genes of Salmonella typhimurium in Longyan city in order to provide reference for the prevention and control.@*Methods@#A total of 79 Salmonella typhimurium strains were isolated from sporadic cases of diarrhea and food poisoning and raw poultry meat samples during 2010 to 2017. PFGE was performed to measure the minimum inhibitory concentrations (MIC) of 15 commonly used drugs against them. PCR was used to detect nine virulence genes (sopB, invA, sifA, sscA, sseE, spvB, pefA, spvR, spvC) in 55 strains.@*Results@#The 79 Salmonella typhimurium strains belonged to 61 PFGE types. There were 10, three and four strains of P1, P3 and P21 types, respectively. Seven P1 type strains were isolate from one food poisoning event. According to the 85% classification standard, 79 Salmonella typhimurium strains could be divided into five predominant gene clusters (G1-G5). Drug susceptibility test showed that the 79 strains had the highest resistance rate to ampicillin (88.61%), followed by that to tetracycline (87.34%) and streptomycin (73.41%). Multidrug resistant bacteria resistant to three or more antibacterial drugs accounted for 84.81% (67/79). All of the 55 strains carried invA, sopB, sseE and sscA genes. The other five genes, sifA, spvC, spvB, spvR and pefA, were detected in 54, 31. 10, 11 and 12 strains, respectively. There were 76.4% (42/55) of the strains carrying five or six virulence genes and all were positive for invA, sopB, sseE, sscA and sifA, and negative for spvB, spvR and pefA. The strains carrying all of the nine virulence genes accounted for 18.2% (10/55).@*Conclusions@#Salmonella typhimurium strains isolated in Longyan city had a diverse PFGE type. P1, P3 and P21 types were the three predominant PFGE types. In the food poisoning event, PFGE molecular typing could quickly alert the outbreak and traceability of Salmonella typhimurium. Attention should be paid to the multidrug resistance in Salmonella typhimurium. Monitoring of multidrug-resistant strains and supervision on antibacterial drug usage should be strengthened. Salmonella typhimurium had high virulence as it carried many virulence genes.

Comparison of three diagnostic methods for Salmonella enterica serovars detection in chicken rinse / Comparação de três métodos diagnósticos para detecção de Salmonella enterica em lavados de carcaça de frango

Salmonella detection is a key point in food safety testing, because of the frequent association of this pathogen with food poisoning in humans. The standard bacteriological tests currently used for Salmonella-detection are time-consuming; therefore, there is a need to develop alternative methods to accelerate the detection. In order to accelerate Salmonella diagnosis, we used the immunomagnetic separation assay associated with bacteriophage P22 for the rapid detection of the following Salmonella serovars in chicken rinses of drumsticks, artificially contaminated with 5, 10, and 100 CFU/25mL of bacteria: Salmonella enterica subsp. enterica serovar Heidelberg (S. Heidelberg), Salmonella enterica subsp. enterica serovar Enteritidis (S. Enteritidis) and Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium). The efficiency of the technique, represented by the time required for detection of positive and negative samples, was compared with that of the standard diagnostic tests used for this pathogen, the bacteriological assay and the polymerase chain reaction (PCR)-based test. This study confirmed the ability of the bacteriophage-associated immunomagnetic separation assay to identify 99.6% of Salmonella-positive samples of the three serovars tested. In contrast, the bacteriological assay and PCR-based test detected 95.1% and 98.5% of the Salmonella-positive samples respectively.(AU)

A detecção de Salmonella é um ponto crucial para a segurança alimentar, devido a frequente associação deste patógeno com infecções alimentares em humanos. O método padrão para detecção de Salmonella é o bacteriológico, mas o tempo requerido para o processamento das amostras e o diagnóstico final é longo, por isso existe a necessidade de desenvolvimento de métodos alternativos que visem acelerar esta etapa. Para isto utilizamos a separação imunomagnética associada ao bacteriófago P22 como técnica de detecção rápida para os seguintes sorovares de Salmonella: Salmonella enterica subsp. enterica sorovar Heidelberg (S. Heidelberg), Salmonella enterica subsp. enterica sorovar Enteritidis (S. Enteritidis) e Salmonella enterica subsp. enterica sorovar Typhimurium (S. Typhimurium), os quais foram inoculados artificialmente em lavados de sobre-coxas de frango nas seguintes concentrações: 5, 10 e 100 UFC/25mL. A eficiência da técnica, representada pelo tempo requerido para detecção de amostras positivas ou negativas, foi comparado com os testes rotineiramente utilizados para detecção de Salmonella, o exame bacteriológico e a reação em cadeia da polimerase (PCR). Este estudo confirmou a capacidade do teste de separação imunomagnética associado a bacteriófago, o qual identificou 99,6% das amostras positivas para Salmonella, dos três sorovares testados. Já o bacteriológico e PCR identificaram respectivamente 95,1% e 98,5% das amostras positivas.(AU)

Serotyping and PFGE Type of Salmonella Isolates in Yunnan Province / 昆明医科大学学报

Objective To study the serotypes and molecular characteristics of foodborne Salmonella in Yunnan Province using pulsed field gel electrophoresis (PFGE) and to establish PFGE fingerprint database. Methods This study was carried out on the basis of the Serological typing of 322 strains of foodborne Salmonella which was isolated from Yunnan national foodborne disease surveillance from 2013 to 2016. The clustering analysis was conducted on 148 strains of Salmonella DNA restriction enzyme map by using the software of BioNumerics. Fingerprint database was established through the comparison of clustering analysis correlation on bacterial strain. Results The serotype of 322 strains of Salmonella mainly included A, B, C, D, E, F, G and other 7 groups,among which Salmonella typhimurium was the major type, accounting for 11.4% (37/322) . Cluster analysis was applied using BioNumerics software in 148 strains of Salmonella DNA restriction enzyme map. According to the different number and the different positions, electrophoresis strips were divided into 102 different PFGE patterns (Figure 1) , which was categorized into 39 clusters if 90% of the strips was similar. Conclusion Foodborne salmonella molecular classification is complex. Salmonella typhimurium is the major type. PFGE belt type presents diversity.

Serotype distribution characteristics and molecular typing of Salmonella in Yunnan Province from 2015 to 2017 / 中华微生物学和免疫学杂志

Objective To study the serotype distribution characteristics and genotypes of Salmo-nella strains isolated in Yunnan Province from 2015 to 2017. Methods Automatic microbiological identifi-cation system and mass spectrometer were used to identify Salmonella strains. Their serotypes were detected using the White-Kauffmann-Le Minor (WKL) scheme based on serological detection. Genotyping was car-ried out by referring to the molecular typing method of Salmonella serotype pulse field gel electrophoresis (PFGE) in PulsenetChina. Cluster analysis was performed with Bionumerics (7.6). Results A total of 408 strains of Salmonella were detected in food and patients in Yunnan Province form 2015 to 2017,belong-ing to 70 serotypes. Thirty-four Salmonella derby strains were detected in food,accounting for 19.10% of all Salmonella strains detected in food. Among the Salmonella strains detected in patients,71 were Salmonella enteritis and 67 were Salmonella typhimurium,accounting for 30.34% and 27.63%, respectively. Results of PFGE revealed that Salmonella derby and Salmonella typhimurium were polymorphic,and Salmonella en-teritis had obvious advantages PFGE band patterns. No obvious time or geographical aggregation was found in the PFGE bands of the three Salmonella species. Conclusion Seventy Salmonella serotypes had been iden-tified in Yunnan Province by 2017. Salmonella derby was the predominant serotype detected in food, while Salmonella enteritidis and Salmonella typhimurium were the predominant serotypes in patients. These three Salmonella species caused sporadic infections in Yunnan Province.

Salicylic Acid Reduces OmpF Expression, Rendering Salmonella enterica Serovar Typhimurium More Resistant to Cephalosporin Antibiotics / 전남의대학술지

Salmonella enterica serovar Typhimurium is one of the most important bacterial pathogens causing diarrhea. The resistance of S. typhimurium to antimicrobial agents, which has recently been isolated from patients, is causing serious problems. We investigated the effects of salicylic acid (Sal) and acetyl salicylate (AcSal) on the susceptibility of S. typhimurium to cephalosporin antibiotics, which are known to increase resistance to cephalosporin and quinolone antibiotics. The MIC of cephalosporin antibiotics was higher than that of the media without Sal. The rate of accumulation of ethidium bromide (EtBr) in the bacteria by the outer membrane protein (Omp) was not different from that of the bacteria cultured in the medium containing Sal. However, Carbonyl cyanide-m-chlorophenylhydrazone (CCCP), an inhibitor of bacterial efflux pumps, significantly reduced the rate of accumulation of EtBr in bacteria cultured on Sal containing medium. In the medium containing CCCP, the MIC of the antimicrobial agent tended to decrease as compared with the control. In addition, the MIC of the bacteria treated with CCCP and Sal was higher than that of the antimicrobial agent against the CCCP treated experimental bacteria. These results suggest that Sal decreases the expression of OmpF in the Omp of S. typhimurium and reduces the permeability of cephalosporin antibiotics to bacteria, which may induce tolerance to cephalosporin antibiotics.

Antibacterial and antibiofilm activity of carvacrol against Salmonella enterica serotype Typhimurium

ABSTRACT The present study evaluated the antibacterial and antibiofilm activity of carvacrol against Salmonella Typhimurium. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined and the time-kill curve and scanning electron microscopy (SEM) were performed to evaluate antibacterial activity. Antibiofilm activity was evaluated by quantifying total biomass using crystal violet assay, and metabolic activity was determined using MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay. The action of carvacrol against preformed biofilm on polypropylene and stainless steel was also evaluated by colony counting and SEM. The MIC and MBC was 312 µg mL-1. Carvacrol at MIC and 2 x MIC eliminated cells after 6 and 1 h of treatment, respectively, as exhibited in the time-kill curve. The greatest reduction in biofilm biomass and metabolic activity was 1,719 OD550 and 0,089 OD550 respectively, both at 4 x MIC of carvacrol. In carvacrol treated biofilms of S. Typhimurium on polypropylene, a reduction of 5.12 log was observed with 4 x MIC, while on stainless steel, carvacrol at 4 x MIC reduced bacterial counts by 5 log. The results showed that carvacrol exhibits antibacterial activity and can be used as an alternative for the control of S. Typhimurium biofilms.

Pathologic and microbiologic aspects of pet psittacine infected by Escherichia coli and Salmonella Typhimurium / Aspectos patológicos e microbiológicos de Psittaciformes de companhia infectados por Escherichia coli e Salmonella Typhimurium

The role of Escherichia coli in healthy microbiota of psittacine is controversial, and the presence of Salmonella sp. indicates possible disease. Therefore, this study aimed to identify the presence of E. coli and Salmonella spp. in a psittacine pet that died in Fortaleza, Brazil, correlating pathogenicity aspects of the isolates through the evaluation of lesions and antimicrobial susceptibility. Psittacine pets sent to the Laboratory of Ornithological Studies, State University of Ceará, that died in 2014 and 2015 were necropsied. Fragments of liver, kidneys, intestine, lung, heart, spleen and brain were collected for microbiological and histopathological analyses. Scores were attributed to lesions and isolated strains submitted to antimicrobial susceptibility test. From the seventy necropsied birds, nineteen were positive for E. coli and one for Salmonella Typhimurium. Congestive lesions and lymphoplasmocitic inflammatory infiltrate were observed varying from light to moderate and were the main findings. In the analyzed strains, multidrug resistance against different groups of antibiotics was observed. In conclusion, according to the results, E. coli strains and the Salmonella Typhimurium isolate produced significant lesions in the psittacine pets, and multidrug resistance may hinder treatments with antibiotics used in avian pet medicine.(AU)

A participação de Escherichia coli na microbiota saudável de Psicittaciformes e a de Salmonella spp. já indica possível doença. O objetivo deste estudo foi pesquisar a presença de E. coli e Salmonella spp. em psittaciformes de companhia na cidade de Fortaleza/Ceará, traçando os aspectos de patogenicidade destas cepas através das lesões e da sensibilidade antimicrobiana. Foram necropsiados os psittaciformes de companhia encaminhados ao Laboratório de Estudos Ornitológicos da Universidade Estadual do Ceará durante o período de 2014 a 2015. No momento da necropsia foram coletados fragmentos de fígado, rins, intestino, pulmão, coração, baço e encéfalo para posterior processamento microbiológico e histopatológico. As lesões foram graduadas e as cepas isoladas submetidas a antibiograma. Das setenta aves necropsiadas, dezenove foram positivas para E. coli e apenas uma para Salmonella Typhimurium. As lesões de congestão e infiltrado inflamatório linfoplasmocitário variaram de leve a moderado, e foram as principais lesões encontradas. Nas cepas analisadas foi constatada multiresistência a diferentes grupos de antibióticos testados. De acordo com os achados, pode-se concluir que os isolados de E. coli e Salmonella Typhimurium produziram lesões significativas em psittaciformes em Fortaleza, Brasil, e a multirresistência pode dificultar o tratamento com antibióticos usados na clínica de aves de companhia.(AU)

A Salmonella Typhimurium outbreak linked to Vietnamese bread rolls in South Western Sydney, Australia, 2015

Introduction: In September 2015, the South Western Sydney (SWS) Public Health Unit was notified of a cluster of Salmonella Typhimurium (STm) cases with a common multiple-locus variable-number tandem repeats analysis (MLVA) pattern. An investigation was conducted to identify a source and contain the outbreak. Methods: The cluster was initially identified through routine geographic information system cluster scanning applied to the New South Wales Notifiable Conditions Management System. Additional cases were identified through a complaint to local council about a bakery. The bakery was inspected and 48 environmental and food swabs were collected for analysis. Results: A total of 26 suspected cases were identified, of which 14 were interviewed. STm MLVA type 3-16-9-11-523 was identified in 19 of 26 case stool specimens. Most cases (12/14) consumed bread rolls containing pork or chicken with chicken liver pâté and raw egg mayonnaise filling. Five cases identified a common bakery exposure. Environmental and food samples from the bakery isolated STm with an identical MLVA pattern. Discussion: An STm cluster in SWS was investigated and found to be linked to Vietnamese bread rolls containing pork or chicken with chicken liver pâté and raw egg mayonnaise filling. Confirmation of a distinct MLVA pattern among STm isolates from clinical, food and environmental samples provided evidence to establish an epidemiological link between the cases and the implicated premises and informed public health action to contain the outbreak.