Effect of inoculants combining Lactobacillus buchneri strain LN40177 in different strata of the silo

The objective of this work was to evaluate the chemical composition and rumen disappearance rate of dry matter of corn silages with inoculants combining L. buchneri strain LN40177 in different strata of the silo. The experimental design was a 3x2 randomized complete block design, with three treatments: Control: corn silage without inoculant; 11CFT: corn silage with inoculant which combines L. buchneri strain LN40177 (1.1 x 1011 CFU g-1 ) with L. casei (1.1 x 1011 CFU g-1 ); and 11C33: corn silage with inoculant which combines L. buchneri strain LN40177 (1.1 x 1011 CFU g-1 ) with L. plantarum (1.1 x 1011 CFU g-1 ) and Enterococcus faecium (1 x 1010 CFU g-1 ), associated with two strata of the silo (lower and upper). The silage inoculated with 11C33 presented higher contents of crude protein and NDF and lower hemicellulose content in relation to the control treatment and 11CFT. The use of both inoculants resulted in silages with higher concentrations of soluble nutrients. Lower stratum silage had a higher rumen disappearance rate of dry matter compared to the upper stratum. In general, the combinations of L. buchneri promoted nutritional improvements in corn silage, but in presence of L. casei, there were more outstanding improvements.(AU)

Effect of inoculants combining Lactobacillus buchneri strain LN40177 in different strata of the silo

The objective of this work was to evaluate the chemical composition and rumen disappearance rate of dry matter of corn silages with inoculants combining L. buchneri strain LN40177 in different strata of the silo. The experimental design was a 3x2 randomized complete block design, with three treatments: Control: corn silage without inoculant; 11CFT: corn silage with inoculant which combines L. buchneri strain LN40177 (1.1 x 1011 CFU g-1 ) with L. casei (1.1 x 1011 CFU g-1 ); and 11C33: corn silage with inoculant which combines L. buchneri strain LN40177 (1.1 x 1011 CFU g-1 ) with L. plantarum (1.1 x 1011 CFU g-1 ) and Enterococcus faecium (1 x 1010 CFU g-1 ), associated with two strata of the silo (lower and upper). The silage inoculated with 11C33 presented higher contents of crude protein and NDF and lower hemicellulose content in relation to the control treatment and 11CFT. The use of both inoculants resulted in silages with higher concentrations of soluble nutrients. Lower stratum silage had a higher rumen disappearance rate of dry matter compared to the upper stratum. In general, the combinations of L. buchneri promoted nutritional improvements in corn silage, but in presence of L. casei, there were more outstanding improvements.

Frequency of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) in non-clinical Enterococcus faecalis and Enterococcus faecium strains / Frequência das repetições palindrômicas curtas agrupadas e regularmente interespaçadas (CRISPRs) em cepas não-clínicas de Enterococcus faecalis e Enterococcus faecium

The fidelity of the genomes is defended by mechanism known as Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) systems. Three Type II CRISPR systems (CRISPR1- cas, CRISPR2 and CRISPR3-cas) have been identified in enterococci isolates from clinical and environmental samples. The aim of this study was to observe the distribution of CRISPR1-cas, CRISPR2 and CRISPR3-cas in non-clinical strains of Enterococcus faecalis and Enterococcus faecium isolates from food and fecal samples, including wild marine animals. The presence of CRISPRs was evaluated by PCR in 120 enterococci strains, 67 E. faecalis and 53 E. faecium. It is the first report of the presence of the CRISPRs system in E. faecalis and E. faecium strains isolated from wild marine animal fecal samples. The results showed that in non-clinical strains, the CRISPRs were more frequently detected in E. faecalis than in E. faecium. And the frequencies of CRISPR1-cas and CRISPR2 were higher (60%) in E. faecalis strains isolated from animal feces, compared to food samples. Both strains showed low frequencies of CRISPR3-cas (8.95% and 1.88%). In conclusion, the differences in the habitats of enterococcal species may be related with the results observe in distribution of CRISPRs systems.(AU)

A fidelidade dos genomas é defendida por mecanismos conhecidos como sistemas de repetições palindrômicas curtas agrupadas e regularmente interespaçadas (CRISPRs). Três tipos de sistemas CRISPR II (CRISPR1-cas, CRISPR2 e CRISPR3-cas) têm sido identificados em cepas de enterococos isolados de amostras clínicas e ambientais. O objetivo deste estudo foi observar a distribuição dos CRISPR1-cas, CRISPR2 e CRISPR3-cas em cepas não-clínicas de Enterococcus faecalis e Enterococcus faecium isoladas de amostras alimentícias e fecais, incluindo animais marinhos selvagens. A presenca dos CRISPRs foi determinada por PCR em 120 cepas de enterococos, sendo 67 E. faecalis e 53 E. faecium. É o primeiro relato da presença do sistema CRISPRs nas estirpes E. faecalis e E. faecium isoladas de amostras fecais de animais marinhos selvagens. Os resultados mostraram que em cepas não-clínicas, os CRISPRs foram mais frequentemente detectados em E. faecalis do que em E. faecium. E as frequências de CRISPR1-cas e CRISPR2 foram maiores (60%) em cepas de E. faecalis isoladas de fezes de animais, quando comparadas à amostras de alimentos. Ambas as cepas apresentaram baixas freqüências de CRISPR3-cas (8,95% e 1,88%). Em conclusão, as diferenças nos habitats das espécies de enterococos podem estar relacionadas com os resultados observados na distribuição dos sistemas CRISPRs.(AU)

Frequency of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) in non-clinical Enterococcus faecalis and Enterococcus faecium strains

Abstract The fidelity of the genomes is defended by mechanism known as Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) systems. Three Type II CRISPR systems (CRISPR1- cas, CRISPR2 and CRISPR3-cas) have been identified in enterococci isolates from clinical and environmental samples. The aim of this study was to observe the distribution of CRISPR1-cas, CRISPR2 and CRISPR3-cas in non-clinical strains of Enterococcus faecalis and Enterococcus faecium isolates from food and fecal samples, including wild marine animals. The presence of CRISPRs was evaluated by PCR in 120 enterococci strains, 67 E. faecalis and 53 E. faecium. It is the first report of the presence of the CRISPRs system in E. faecalis and E. faecium strains isolated from wild marine animal fecal samples. The results showed that in non-clinical strains, the CRISPRs were more frequently detected in E. faecalis than in E. faecium. And the frequencies of CRISPR1-cas and CRISPR2 were higher (60%) in E. faecalis strains isolated from animal feces, compared to food samples. Both strains showed low frequencies of CRISPR3-cas (8.95% and 1.88%). In conclusion, the differences in the habitats of enterococcal species may be related with the results observe in distribution of CRISPRs systems.

Resumo A fidelidade dos genomas é defendida por mecanismos conhecidos como sistemas de repetições palindrômicas curtas agrupadas e regularmente interespaçadas (CRISPRs). Três tipos de sistemas CRISPR II (CRISPR1-cas, CRISPR2 e CRISPR3-cas) têm sido identificados em cepas de enterococos isolados de amostras clínicas e ambientais. O objetivo deste estudo foi observar a distribuição dos CRISPR1-cas, CRISPR2 e CRISPR3-cas em cepas não-clínicas de Enterococcus faecalis e Enterococcus faecium isoladas de amostras alimentícias e fecais, incluindo animais marinhos selvagens. A presenca dos CRISPRs foi determinada por PCR em 120 cepas de enterococos, sendo 67 E. faecalis e 53 E. faecium. É o primeiro relato da presença do sistema CRISPRs nas estirpes E. faecalis e E. faecium isoladas de amostras fecais de animais marinhos selvagens. Os resultados mostraram que em cepas não-clínicas, os CRISPRs foram mais frequentemente detectados em E. faecalis do que em E. faecium. E as frequências de CRISPR1-cas e CRISPR2 foram maiores (60%) em cepas de E. faecalis isoladas de fezes de animais, quando comparadas à amostras de alimentos. Ambas as cepas apresentaram baixas freqüências de CRISPR3-cas (8,95% e 1,88%). Em conclusão, as diferenças nos habitats das espécies de enterococos podem estar relacionadas com os resultados observados na distribuição dos sistemas CRISPRs.

Frequency of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) in non-clinical Enterococcus faecalis and Enterococcus faecium strains

Abstract The fidelity of the genomes is defended by mechanism known as Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) systems. Three Type II CRISPR systems (CRISPR1- cas, CRISPR2 and CRISPR3-cas) have been identified in enterococci isolates from clinical and environmental samples. The aim of this study was to observe the distribution of CRISPR1-cas, CRISPR2 and CRISPR3-cas in non-clinical strains of Enterococcus faecalis and Enterococcus faecium isolates from food and fecal samples, including wild marine animals. The presence of CRISPRs was evaluated by PCR in 120 enterococci strains, 67 E. faecalis and 53 E. faecium. It is the first report of the presence of the CRISPRs system in E. faecalis and E. faecium strains isolated from wild marine animal fecal samples. The results showed that in non-clinical strains, the CRISPRs were more frequently detected in E. faecalis than in E. faecium. And the frequencies of CRISPR1-cas and CRISPR2 were higher (60%) in E. faecalis strains isolated from animal feces, compared to food samples. Both strains showed low frequencies of CRISPR3-cas (8.95% and 1.88%). In conclusion, the differences in the habitats of enterococcal species may be related with the results observe in distribution of CRISPRs systems.

Resumo A fidelidade dos genomas é defendida por mecanismos conhecidos como sistemas de repetições palindrômicas curtas agrupadas e regularmente interespaçadas (CRISPRs). Três tipos de sistemas CRISPR II (CRISPR1-cas, CRISPR2 e CRISPR3-cas) têm sido identificados em cepas de enterococos isolados de amostras clínicas e ambientais. O objetivo deste estudo foi observar a distribuição dos CRISPR1-cas, CRISPR2 e CRISPR3-cas em cepas não-clínicas de Enterococcus faecalis e Enterococcus faecium isoladas de amostras alimentícias e fecais, incluindo animais marinhos selvagens. A presenca dos CRISPRs foi determinada por PCR em 120 cepas de enterococos, sendo 67 E. faecalis e 53 E. faecium. É o primeiro relato da presença do sistema CRISPRs nas estirpes E. faecalis e E. faecium isoladas de amostras fecais de animais marinhos selvagens. Os resultados mostraram que em cepas não-clínicas, os CRISPRs foram mais frequentemente detectados em E. faecalis do que em E. faecium. E as frequências de CRISPR1-cas e CRISPR2 foram maiores (60%) em cepas de E. faecalis isoladas de fezes de animais, quando comparadas à amostras de alimentos. Ambas as cepas apresentaram baixas freqüências de CRISPR3-cas (8,95% e 1,88%). Em conclusão, as diferenças nos habitats das espécies de enterococos podem estar relacionadas com os resultados observados na distribuição dos sistemas CRISPRs.

EFEITO ANTI-INFLAMATÓRIO DE Enterococcus faecium 137v ISOLADO DO QUEIJO DE COALHO SOBRE O DESENVOLVIMENTO DO CÂNCER COLORRETAL EM MODELO ANIMAL

Dentre as bactérias ácido láticas encontradas no microambiente do queijo de Coalho está a Enterococcus faecium uma importante espécie que também forma parte integrante da microbiota humana saudável. A aplicação tecnológica de Enterococcus mais amplamente explorada é a sua atuação como bactéria probiótica, cuja função a partir do seu consumo em quantidades adequadas, é reforçar o sistema imunológico do hospedeiro. Tendo em vista que o câncer colorretal está em constante crescimento no mundo por fatores relacionados à alimentação, pesquisas científicas que busquem alternativas eficazes, acessíveis, adaptáveis ao produtor e ao consumidor são de grande relevância. O objetivo desta tese foi utilizar uma bactéria ácido lática Enterococcus faecium 137v (EF137v) isolada da microbiota do queijo de Coalho já avaliada como probiótica para atuar como promotora de prevenção e adjuvante terapêutico para o câncer colorretal em modelo animal. Para isso esta pesquisa avaliou a administração de EF137v em ratos Wistar induzidos pelo carcinógeno 1,2-dimetilhidrazina-DMH ao câncer colorretal. Os grupos experimentais foram distribuídos em Grupo 1 (controle negativo), Grupo 2 (controle positivo, recebendo penas DMH), Grupo 3 (Recebeu apenas EF137v), Grupo 4 e 5 (consumiram EF137v antes e após indução do câncer por DMH). Grupo 6 e 7 (consumiram EF137v após indução do câncer), sendo Grupo 4 e 7 receberam o antineoplásico Vincristina (VCR). Em primeiro momento, os animais foram acompanhados através da ultrassonografia abdominal, a fim de observar os efeitos da administração oral da EF137v sobre os diversos grupos experimentais na saúde sistêmica dos ratos Wistar, bem como observações de possíveis alterações no peso e níveis séricos desses animais. Em segundo momento, a histopatologia, imunohistoquímica para VEGF-A e apoptose foram analisadas. Em terceiro momento, foi avaliado se EF137v pode minimizar o efeito do câncer colorretal, induzido DMH no fígado. Sendo realizadas análises bioquímicas da função hepática, histológicas, morfométricas e imunohistoquímicas. Os ratos apresentaram peso corporal estável, sem mudanças bruscas no decorrer do experimento. Triglicerídeos apresentaram-se altos após o câncer na maioria dos grupos tratados. Bilirrubina, Alanina Amino Transferase e Fosfatase Alcalina não apresentaram alterações significativas. A mucosa intestinal foi preservada, fraca marcação para VEGF-A e índice atenuado de apoptose foram observadas nos grupos tratados com EF137v. Níveis baixos de glicogênio foram observados no grupo controle positivo (Grupo 2) que recebeu apenas DMH quando comparado aos demais grupos. Forte marcação para IL1- e TNF- foi identificada no grupo que recebeu o EF137v. Coclui-se que EF137v aumenta a proteção do intestino e fígado durante o desenvolvimento do câncer colerratal e o procedimento ultrassonográfico pode ser uma ferramenta adicional para o acompanhar alterações importantes no intestino em modelo animal.

Among the acid lactic bacteria found in microenvironment of Coalho cheese is Enterococcus faecium, an important species that also forms an integral part of the healthy human microbiota. The most widely explored technological application of Enterococcus is its performance as a probiotic bacterium, whose function from its consumption in adequate quantities, is to strengthen the host's immune system. Bearing in mind that colorectal cancer is constantly growing in the world due to factors related to food, scientific research that seeks effective, accessible alternatives, adaptable to producers and consumers is of great relevance. The objective of this thesis was to use a acid lactic bacteria Enterococcus faecium 137v (EF137v) extracted from the Coalho cheese microbiota with probiotic characteristics to act as a preventive promoter and therapeutic adjuvant for colorectal cancer in an animal model. For this, this research evaluated the administration of EF137v in Wistar rats induced by the carcinogen 1,2-dimethylhydrazine-DMH to colorectal cancer. The experimental groups were divided into Group 1 (negative control), Group 2 (positive control, receiving DMH penalties), Group 3 (Received only EF137v), Group 4 and 5 (consumed EF137v before and after cancer induction by DMH). Group 6 and 7 (consumed EF137v after cancer induction), and Group 4 and 7 received the antineoplastic Vincristine (VCR). At first, the animals were followed up through abdominal ultrasound, in order to observe the effects of oral administration of EF137v on the various experimental groups on the systemic health of Wistar rats, as well as observations of possible changes in the weight and serum levels of these animals. Second, histopathology, immunohistochemistry for VEGF-A and apoptosis were analyzed. Thirdly, it was assessed whether EF137v can minimize the effect of colorectal cancer, induced DMH in the liver. Biochemical analyzes of liver function, histological, morphometric and immunohistochemistry are performed. The rats had a stable body weight, with no sudden changes during the experiment. Triglycerides were high after cancer in most treated groups. Bilirubin, Alanine Amino Transferase and Alkaline Phosphatase showed no significant changes. The intestinal mucosa was preserved, weak labeling for VEGF-A and attenuated apoptosis index were observed in the groups treated with EF137v. Low glycogen levels were observed in the positive control group (Group 2) that received only DMH when compared to the other groups. Strong labeling for IL1- and TNF- was identified in the group that received EF137v. It is concluded that EF137v increases the protection of the intestine and liver during the development of cholesterol cancer and the ultrasound procedure can be an additional tool to accompany important changes in the intestine in an animal model.

Healthcare-associated vancomycin resistant Enterococcus faecium infections in the Mansoura University Hospitals intensive care units, Egypt

Vancomycin resistant Enterococcus faecium (VREF) ia an emerging and challenging nosocomial pathogen. This study aimed to determine the prevalence, risk factors and clonal relationships between different VREF isolates in the intensive care units (ICUs) of the university hospitals in our geographic location. This prospective study was conducted from July, 2012 until September, 2013 on 781 patients who were admitted to the ICUs of the Mansoura University Hospitals (MUHs), and fulfilled the healthcare-associated infection (HAI) criteria. Susceptibility testing was determined using the disk diffusion method. The clonal relationships were evaluated with pulsed field gel electrophoresis (PFGE). Out of 52 E. faecium isolates, 12 (23.1%) were vancomycin resistant. The significant risk factors for the VREF infections were: transfer to the ICU from a ward, renal failure, an extended ICU stay and use of third-generation cephalosporins, gentamicin, or ciprofloxacin. PFGE with the 12 isolates showed 9 different patterns; 3 belonged to the same pulsotype and another 2 carried a second pulsotypes. The similar pulsotypes isolates were isolated from ICUs of one hospital (EICUs); however, all of the isolates from the other ICUs had different patterns. Infection control policy, in conjunction with antibiotic stewardship, is important to combat VREF transmission in these high-risk patients..(AU)

Safety, beneficial and technological properties of Enterococcus faecium isolated from Brazilian cheeses

This study aimed to characterize the safety and technological properties of Enterococcus faecium strains isolated from Brazilian Coalho cheeses. High levels of co-aggregation were observed between Enterococcus faecium strains EM485 and EM925 and both Escherichia coli and Clostridium perfringens. Both strains presented low levels of hydrophobicity. E. faecium EM485 and EM925 were both able to grow in the presence of 0.5% of the sodium salts of taurocholic acid (TC), taurodeoxycholic acid (TDC), glycocholic acid (GC), and glycodeoxycholic acid (GDC), although they showed the ability to deconjugate only GDC and TDC. Both strains showed good survival when exposed to conditions simulating the gastro intestinal tract (GIT). When tested for the presence of virulence genes, only tyrosine decarboxylase and vancomycin B generated positive PCR results.(AU)

Effect of ingestion of soy yogurt on intestinal parameters of rats fed on a beef-based animal diet

The aim of this study was to investigate whether the ingestion of soy yogurt fermented with Enterococcus faecium CRL 183 would modify the intestinal count of enterococci, fecal pH and ammonia content in rats fed on a diet containing red meat. The rats were placed in 4 groups: for 60 days, group I was given a standard casein-based rodent feed and groups II-IV, the beef-based feed. From day 30, groups III-IV also received the following products: III) soy yogurt; IV) suspension of E. faecium CRL 183. At the start and on days 30 and 60, feces were collected for the determination of pH, ammonia content, count of enterococci and identification of their species. On day 60, rats were sacrificed and their colons also removed for count of enterococci and identification of their species. Rats that ingested soy yogurt showed no significant change (P 0.05) in fecal counts of Enterococcus spp., but, this rat group showed a higher count of E. faecium than rats that ingested suspension of E. faecium CRL 183. The ingestion of soy yogurt and E. faecium culture caused a significant rise (P 0.05) in fecal pH and ammonia content. Our results suggest that consumption of soy yogurt fermented with E. faecium CRL 183 and L. helveticus subsp. jugurti could change the species of Enterococcus spp. present in the feces and colon of rats fed on a beef-based diet. However, the fermented soy product and the pure culture of E. faecium CRL 183 also induced undesirable effects such as the increase of fecal pH and ammonia content in the feces of rats fed on a beef-based diet.

Antimicrobial activity of Enterococcus faecium FAIR-E 198 against gram-positive pathogens

This study investigated the antimicrobial activity of Enterococcus faecium FAIR-E 198 against Bacillus cereus, Listeria monocytogenes and Staphylococcus aureus. Using the critical-dilution method, the bacteriocin produced by E. faecium FAIR-E 198 inhibited all L. monocytogenes strains evaluated (1,600 to 19,200 AU mL-1). However, none of the B. cereus and S. aureus strains investigated were inhibited. The maximum activity of this bacteriocin (800 AU mL-1) was observed in MRS broth, while the activity in milk was 100 AU mL-1. In the co-cultivation test in milk, B. cereus K1-B041 was reduced to below the detection limit (1.00 log CFU mL-1) after 48 h. E. faecium reduced the initial L. monocytogenes Scott A population by 1 log CFU mL-1 after 3 h at 35ºC. However, the pathogen regained growth, reaching 3.68 log CFU mL-1 after 48 h. E. faecium did not influence the growth of S. aureus ATCC 27154 during the 48 h of co-cultivation. Therefore, it can be concluded that the effectiveness of the antimicrobial activity of E. faecium FAIR-E 198 is strictly related to the species and strain of the target microorganism and to the culture medium.