Influence of the food matrix on dual-species biofilm of Listeria monocytogenes and Pseudomonas fluorescens

In the food industry, the formation of biofilms results in serious microbial recontamination problem. This work aimed to study Listeria sp. obtained from sliced products and food handling surfaces, for the preparation of food of animal origin to be consumed cold. Listeria monocytogenes, Listeria innocua and Listeria seeligeri were detected. In the evaluation of antibiotic susceptibility, the isolates were sensitive to ampicillin and penicillin, which were the main antibiotics used in treatment. The isolates showed the ability to form biofilm in microplate, in stainless steel and polypropylene surfaces, with a variation of sessile cells between 4.29±0.23 and 7.03±0.01 log10 CFU cm-2 in TSBYE. This ability was also observed in food matrix composed of UHT whole milk in mono-species cultivation and in associated cultivation of L. monocytogenes with Pseudomonas fluorescens. The application of sanitizers peracetic acid and sodium hypochlorite revealed efficiency in the eradication of adhered cells and biofilms formed in stainless steel surfaces. Therefore, Listeria sp. showed to be persistent and able to form biofilm in mono-species cultivation or associated with P. fluorescens, under different conditions. Taking these aspects into account, the need for proper hygiene in the food production process is highlighted in order to avoid risks to the consumers health.(AU)

EpCAM expression in canine malignant mammary tumors - immunohistochemical study

Background: Epithelial cell adhesion molecule (EpCAM) is a glycoprotein responsible for multiple cellular functions that includes cell-to-cell adhesion and signal transduction. Studies in human breast cancer demonstrated that its overexpression has been linked to a more aggressive histological phenotype. However, in canine mammary tumors, its use is still scarce and its function, remains unknown. The main goal of the present study was to characterize the expression of EpCAM in canine mammary malignant tumors and associate its expression with clinicopathological features. Materials, Methods & Results: Forty-eight canine mammary samples were analyzed in the present study and went through the Hematoxylin-Eosin (HE) routine technique for histopathological and immunohistochemical analysis. EpCAM expression was evaluated considering the total immunolabelling (TI) which consists of the addition of 2 parameters: proportion score (PS) and intensity score (IS). Thirteen (27.1%) cases showed weak expression, 9 (18.7%) were characterized by a moderate expression and 27 (54.2%) were classified as intense. EpCAM overexpression was described in 36 (72.9%) primary tumors and in 5 lymph node metastases (71.4%). Immunoreactivity was, mainly, characterized by a staining in the lateral cell membrane of luminal epithelial cells, both in primary tumors and lymph node metastases. The statistical analysis was performed using the SPSS Statistics, version 24. Clinicopathological features, such as histological grade (P = 0.027), intravascular emboli (P = 0.004) and lymph node metastases (P = 0.016) demonstrated to be associated with a more aggressive histological type. Intravascular emboli and lymph node metastases were also linked to a higher histological grade. Discussion: EpCAM overexpression has been described in the literature both in canine mammary tumors and breast cancer. While the results in breast cancer could be ambiguous, depending on the tumor subtype, in the present study our findings demonstrated that overexpression was present in more than 50% of the total cases evaluated. In addition, immunoreactivity was also in accordance with the literature that describes membranous staining as the most prevalent pattern type. It was not possible to correlate overexpression with histological type and histological grade. However, those clinicopathological features are essential to evaluate tumor prognosis. For instance, our results demonstrated that histological type was statistically significant with histological grade (P = 0.027), intravascular emboli (P = 0.004) and lymph node metastases (P = 0.016). Additionally, the Nottingham system was statistically significant with intravascular emboli (P = 0.033) and lymph node metastases (P = 0.006). These findings have also been described in similar studies using canine mammary tissues in bitches. When it comes to pattern of expression in primary tumors and lymph nodes metastases, our findings described that neoplastic cell presented the same phenotype in these 2 different locations. In addition, loss of expression was also described in cluster cells within the subcapsular area which reflects the idea that the expression of EpCAM molecule changes according to time, which could be translated into the multistep process of metastases. Finally, it demonstrates that the development of neoplasia is not something static, but rather dynamic.

Effect of Mauritia flexuosa L. leaf extract on Staphylococcus aureus and Staphylococcus haemolyticus biofilms adhered to stainless steel surface / Efeito do extrato de folhas de Mauritia flexuosa L. sobre biofilmes de Staphylococcus aureus e Staphylococcus haemolyticus aderidos a superfície de aço inoxidável

Staphylococcus spp. represents the main mastitis agents in ruminants and contaminants of milk due to their expressive capacity to make biofilms. The aims in this study was evaluate evaluated the antimicrobial activity of Mauritia flexuosa L. extracts against Staphylococcus spp. adhered to a stainless steel surface. Two isolates from cows with clinical mastitis were evaluated; one was identified as Staphylococcus aureus, and the other Staphylococcus haemolyticus. Additionally the ATCC 25923 strain, S. aureus from human was evaluated. The chemical profile obtained from gas chromatography revealed the presence of carbohydrates, organic acids, and flavonoids. The minimum bactericidal concentrations of the ethanolic extract (EE) and aqueous extract (AE) were 4.4 and 5.82 mg/mL, respectively. After EE treatment at 4.4 mg/mL for 2.5 min, total removal of mature biofilms grown on stainless steel coupons was observed (reduction by 3.85-4.81 log units). This extract from M. flexuosa shows potential as an effective sanitizer and may represent a natural alternative against Staphylococcus spp.

Bactérias do gênero Staphylococcus spp. são os principais agentes da mastite em ruminantes e contaminantes do leite devido à expressiva capacidade de formação de biofilmes. Neste estudo o objetivo foi avaliar a atividade antimicrobiana de extratos de Mauritia flexuosa L. (Buritizeiro) contra Staphylococcus spp. aderidos à superfície de aço inoxidável. Foram avaliados dois isolados de vacas com mastite clínica; o um isolado foi identificado como Staphylococcus aureus e o outro como Staphylococcus haemolyticus. Adicionalmente foi também avaliada a e a cepa S. aureus ATCC 25923 de origem humana. O perfil químico obtido por cromatografia gasosa revelou a presença de carboidratos, ácidos orgânicos e flavonóides. As concentrações bactericidas mínimas do extrato etanólico (EE) e do extrato aquoso (AE) foram 4,4 e 5,82 mg / mL, respectivamente. Após o tratamento com EE a 4,4 mg / mL por 2,5 min, foi observada remoção total de biofilmes maduros cultivados em cupons de aço inoxidável (redução de 3,85-4,81 unidades log). O EE de folhas de M. flexuosa apresenta potencial como um desinfetante eficaz e pode representar uma alternativa natural contra Staphylococcus spp.

Biofilm forming antimicrobial-multiresistant Staphylococcus aureus / Multirresistência antimicrobiana de Staphylococcus aureus formadores de biofilme

Staphylococcus aureus is one of the main agents isolated from bovine mastitis cases, characterized by lower cure rates compared to other pathogens causing this disease. This phenomenon is mainly explained by the multiresistance acquisition to antimicrobials and the ability of S. aureus to form biofilms on biotic and abiotic surfaces. In this work 15 samples of S. aureus isolated from the automated milking facility were analyzed regarding the resistance profile to antimicrobials, virulence factors (capsule production, hemolysin, and protease) and adhesion capacity under different temperatures (42±1°C, 36±1°C, 25±1°C, 9±1°C, and 3±1°C). All isolates showed methicillin-resistant (MRSA) characteristics and multidrug resistance profile to the antimicrobials tested (penicillin G, chloramphenicol, oxacillin, cephalexin, tetracycline, amoxicillin + clavulanic acid, sulfa + trimetropim, gentamicin, doxycycline, ceftiofur, neomycin, and vancomycin) with an IRMA index between 0.5 and 1.0. Five isolates were resistant to vancomycin (VRSA), two were resistant to all active principles, and the others to at least six of these drugs. Adhesion capacity and biofilm formation were found in 3 of the 5 evaluated temperatures, including the cooling conditions. Regarding the virulence factors, 86.7% of the isolates formed capsules, 60% revealed the presence of protease, 26.7% expressed the α-hemolysin factor, and 13.3% of them presented β-hemolysin. The fact that all isolates presented MRSA characteristics represents a potential risk to those exposed to this agent, and the formation of biofilm in liners even after the use of detergents and sanitization highlights the urgency of searching for alternatives for dispersion of the biofilm by S. aureusin the automated milking facility.

O Staphylococcus aureus é um dos principais agentes isolados de casos de mastite bovina, caracterizado por menores taxas de cura em comparação com outros patógenos desta enfermidade. Esse fenômeno é explicado principal-mente pela aquisição de resistência à antimicrobianos e a capacidade do S. aureus formar biofilmes em superficies bióti-cas e abióticas. Neste trabalho foram utilizadas 15 amostras de S. aureus isolados de ordenhadeira, analisados quanto ao perfil de resistência à antimicrobianos, fatores de virulência (produção de cápsula, hemolisina e protease) e capacidade de adesão sob diferentes temperaturas (42±1°C, 36±1°C, 25±1ºC, 9±1ºC e 3±1ºC). Todos os isolados apresentaram perfil de multirresistência aos antimicrobianos testados (penicilina G, cloranfenicol, oxacilina, cefalexina, tetraciclina, amoxicilina + ácido clavulônico, sulfa + trimetropim, gentamicina, doxiciclina, ceftiofur, neomicina e vancomicina) com índice IRMA entre 0,5 a 1,0. Duas cepas foram resistentes a todos os princípios ativos e as demais a pelo menos seis destes fármacos. Os isolados avaliados apresentaram característica de meticilina-resistentes (MRSA) e destes, 33,34% (5/15) foram resistentes à vancomicina (VRSA). Houve capacidade de adesão e formação de biofilmes em 3 das 5 tem-peraturas avaliadas, incluindo as temperaturas de refrigeração. Em relação aos fatores de virulência, 86,7% dos isolados formaram cápsula, 60% presença de protease, 26,7% expressaram o fator α-hemolisina e 13,3% β-hemolisina. O fato de todos isolados apresentarem característica MRSA representa um risco potencial aos expostos a esse agente. Já a for-mação de biofilmes em teteiras, mesmo após detergência e sanitização, destacam a urgência de alternativas de dispersão de biofilmes no ambiente de ordenha.

Biofilm forming antimicrobial-multiresistant Staphylococcus aureus / Multirresistência antimicrobiana de Staphylococcus aureus formadores de biofilme

Staphylococcus aureus is one of the main agents isolated from bovine mastitis cases, characterized by lower cure rates compared to other pathogens causing this disease. This phenomenon is mainly explained by the multiresistance acquisition to antimicrobials and the ability of S. aureus to form biofilms on biotic and abiotic surfaces. In this work 15 samples of S. aureus isolated from the automated milking facility were analyzed regarding the resistance profile to antimicrobials, virulence factors (capsule production, hemolysin, and protease) and adhesion capacity under different temperatures (42±1°C, 36±1°C, 25±1°C, 9±1°C, and 3±1°C). All isolates showed methicillin-resistant (MRSA) characteristics and multidrug resistance profile to the antimicrobials tested (penicillin G, chloramphenicol, oxacillin, cephalexin, tetracycline, amoxicillin + clavulanic acid, sulfa + trimetropim, gentamicin, doxycycline, ceftiofur, neomycin, and vancomycin) with an IRMA index between 0.5 and 1.0. Five isolates were resistant to vancomycin (VRSA), two were resistant to all active principles, and the others to at least six of these drugs. Adhesion capacity and biofilm formation were found in 3 of the 5 evaluated temperatures, including the cooling conditions. Regarding the virulence factors, 86.7% of the isolates formed capsules, 60% revealed the presence of protease, 26.7% expressed the α-hemolysin factor, and 13.3% of them presented β-hemolysin. The fact that all isolates presented MRSA characteristics represents a potential risk to those exposed to this agent, and the formation of biofilm in liners even after the use of detergents and sanitization highlights the urgency of searching for alternatives for dispersion of the biofilm by S. aureusin the automated milking facility.(AU)

O Staphylococcus aureus é um dos principais agentes isolados de casos de mastite bovina, caracterizado por menores taxas de cura em comparação com outros patógenos desta enfermidade. Esse fenômeno é explicado principal-mente pela aquisição de resistência à antimicrobianos e a capacidade do S. aureus formar biofilmes em superficies bióti-cas e abióticas. Neste trabalho foram utilizadas 15 amostras de S. aureus isolados de ordenhadeira, analisados quanto ao perfil de resistência à antimicrobianos, fatores de virulência (produção de cápsula, hemolisina e protease) e capacidade de adesão sob diferentes temperaturas (42±1°C, 36±1°C, 25±1ºC, 9±1ºC e 3±1ºC). Todos os isolados apresentaram perfil de multirresistência aos antimicrobianos testados (penicilina G, cloranfenicol, oxacilina, cefalexina, tetraciclina, amoxicilina + ácido clavulônico, sulfa + trimetropim, gentamicina, doxiciclina, ceftiofur, neomicina e vancomicina) com índice IRMA entre 0,5 a 1,0. Duas cepas foram resistentes a todos os princípios ativos e as demais a pelo menos seis destes fármacos. Os isolados avaliados apresentaram característica de meticilina-resistentes (MRSA) e destes, 33,34% (5/15) foram resistentes à vancomicina (VRSA). Houve capacidade de adesão e formação de biofilmes em 3 das 5 tem-peraturas avaliadas, incluindo as temperaturas de refrigeração. Em relação aos fatores de virulência, 86,7% dos isolados formaram cápsula, 60% presença de protease, 26,7% expressaram o fator α-hemolisina e 13,3% β-hemolisina. O fato de todos isolados apresentarem característica MRSA representa um risco potencial aos expostos a esse agente. Já a for-mação de biofilmes em teteiras, mesmo após detergência e sanitização, destacam a urgência de alternativas de dispersão de biofilmes no ambiente de ordenha.(AU)

Produção de biofilme e susceptibilidade in vitro a desinfetantes de Staphylococcus coagulase positiva isolados de leite bovino / Biofilm production and in vitro susceptibility to disinfectants of Staphylococcus coagulase positive isolated from bovine milk

Background: Mastitis is one of the diseases that cause damage to dairy cattle, and coagulase positive Staphylococcus (CPS),are its main etiological agents. The production of biofilms gives these microorganisms a physiological advantage, being animportant virulence factor, as it confers their resistance to the action of disinfectants used in hygiene procedures. Therefore,the objective of this study was to evaluate the capacity of CPS, isolated from milk samples from cases of bovine subclinicalmastitis in the Alagoas dairy basin, biofilm production and its sensitivity to disinfectant agents used before and after dipping.Materials, Methods & Results: In the Alagoas Milk Basin Region, 10 properties were selected from the municipalities ofBatalha, Cacimbinhas, Major Isidoro and Minador do Negrão, which used a mechanical milking system and supplied milk toa dairy under federal inspection. The California Mastitis Test (CMT) was performed on 1155 cows. The animals diagnosedwith subclinical mastitis were classified as: mild (+); moderate (++) and severe (+++). CMT positive samples were collectedfrom 2+, totaling 891 samples. After cleaning the papillary ostium, 2 to 5 mL of milk was collected from each fourth breastbefore milking began. Subsequently, 0.5 µL of each sample was streak plated by exhaustion on 5% Sheep Blood Agar andincubated at 37°C, with readings at 24 and 48 h. The macroscopic and microscopic characteristics of the colonies, catalaseand coagulase test were evaluated. Biofilm formation was determined by the microplate adhesion test. The efficacy of thesanitizers was evaluated with the active ingredients: lactic acid (2%), allantoin (0.05%), iodine (0.5%), chlorhexidine (2.0%),chlorine (2.5%), at times 15, 30 and 60 seconds. It was observed that 91.2% of the samples were able to form biofilm; Of these,22.2%, 29.6% and 48.2% were strong, moderate and weak producers, respectively. The sensitivity profile of chlorhexidine...

Produção de biofilme e susceptibilidade in vitro a desinfetantes de Staphylococcus coagulase positiva isolados de leite bovino / Biofilm production and in vitro susceptibility to disinfectants of Staphylococcus coagulase positive isolated from bovine milk

Background: Mastitis is one of the diseases that cause damage to dairy cattle, and coagulase positive Staphylococcus (CPS),are its main etiological agents. The production of biofilms gives these microorganisms a physiological advantage, being animportant virulence factor, as it confers their resistance to the action of disinfectants used in hygiene procedures. Therefore,the objective of this study was to evaluate the capacity of CPS, isolated from milk samples from cases of bovine subclinicalmastitis in the Alagoas dairy basin, biofilm production and its sensitivity to disinfectant agents used before and after dipping.Materials, Methods & Results: In the Alagoas Milk Basin Region, 10 properties were selected from the municipalities ofBatalha, Cacimbinhas, Major Isidoro and Minador do Negrão, which used a mechanical milking system and supplied milk toa dairy under federal inspection. The California Mastitis Test (CMT) was performed on 1155 cows. The animals diagnosedwith subclinical mastitis were classified as: mild (+); moderate (++) and severe (+++). CMT positive samples were collectedfrom 2+, totaling 891 samples. After cleaning the papillary ostium, 2 to 5 mL of milk was collected from each fourth breastbefore milking began. Subsequently, 0.5 µL of each sample was streak plated by exhaustion on 5% Sheep Blood Agar andincubated at 37°C, with readings at 24 and 48 h. The macroscopic and microscopic characteristics of the colonies, catalaseand coagulase test were evaluated. Biofilm formation was determined by the microplate adhesion test. The efficacy of thesanitizers was evaluated with the active ingredients: lactic acid (2%), allantoin (0.05%), iodine (0.5%), chlorhexidine (2.0%),chlorine (2.5%), at times 15, 30 and 60 seconds. It was observed that 91.2% of the samples were able to form biofilm; Of these,22.2%, 29.6% and 48.2% were strong, moderate and weak producers, respectively. The sensitivity profile of chlorhexidine...(AU)

Suppresses of Astragalus Polysaccharide on E. coli-Induced Injured Intestinal Microvascular through TLR4-NF-B Signal Pathways in Chickens

To investigate the hypothesis that APS can attenuate E. coli-induced microvascular dysfunction in chicken intestine, 60 0-day old male chickens were divided into three groups with 5 replications of 4 chicks. Chicken in the APS group were treated with 15 mg APS daily while the others were given 0 mg APS for 6 days. Then all 7-day old chicken were injected intraperitoneally by E. coli, except for the chicken in the control group. After 4 h, all chickens intestinal samples were collected to detect gene expressions involved in inflammatory factors and adhesion molecules. Results showed that APS attenuated the signs of edema and hemorrhage in 7-day old chicken intestinal mucosa which were induced by E. coli injection. Consistently, APS significantly reduced the increasing mRNA levels of inflammatory factors such as Tumor necrosis factor-a (TNF-a), interleukin (IL) -1 and inducible nitric oxide synthase (iNOS) (p 0.05), the same results were observed in vascular adhesion molecules such as E-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Moreover, we observed that APS supplementation in water suppressed significantly (p 0.05) the decline of reparative factors such as epithelial growth factor (EGF) and basic fibroblast growth factor (bFGF) in E. coli injected group. Furthermore, supplementation with APS substantially blocked (p 0.05) the increase in Toll-like receptor-4 (TLR4) and Nuclear factor (NF)-B mRNA abundance (p=0.087) induced by E. coli infection. This study indicated that microvascular injured chicken intestine induced by E. coli would be attenuated with feeding APS, and the mechanism of repairing were probably mediated through TLR4-NF-B signal pathways.(AU)

Suppresses of Astragalus Polysaccharide on E. coli-Induced Injured Intestinal Microvascular through TLR4-NF-B Signal Pathways in Chickens

To investigate the hypothesis that APS can attenuate E. coli-induced microvascular dysfunction in chicken intestine, 60 0-day old male chickens were divided into three groups with 5 replications of 4 chicks. Chicken in the APS group were treated with 15 mg APS daily while the others were given 0 mg APS for 6 days. Then all 7-day old chicken were injected intraperitoneally by E. coli, except for the chicken in the control group. After 4 h, all chickens intestinal samples were collected to detect gene expressions involved in inflammatory factors and adhesion molecules. Results showed that APS attenuated the signs of edema and hemorrhage in 7-day old chicken intestinal mucosa which were induced by E. coli injection. Consistently, APS significantly reduced the increasing mRNA levels of inflammatory factors such as Tumor necrosis factor-a (TNF-a), interleukin (IL) -1 and inducible nitric oxide synthase (iNOS) (p 0.05), the same results were observed in vascular adhesion molecules such as E-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Moreover, we observed that APS supplementation in water suppressed significantly (p 0.05) the decline of reparative factors such as epithelial growth factor (EGF) and basic fibroblast growth factor (bFGF) in E. coli injected group. Furthermore, supplementation with APS substantially blocked (p 0.05) the increase in Toll-like receptor-4 (TLR4) and Nuclear factor (NF)-B mRNA abundance (p=0.087) induced by E. coli infection. This study indicated that microvascular injured chicken intestine induced by E. coli would be attenuated with feeding APS, and the mechanism of repairing were probably mediated through TLR4-NF-B signal pathways.

Carrier frequency of autosomal recessive disorders (BC, BLAD, FXID and CVM) in Holstein cows in Jalisco, Mexico

The hereditary autosomal recessive disorders bovine citrullinemia (BC), bovine leukocyte adhesion deficiency (BLAD), factor XI deficiency (FXID), and complex vertebral malformation (CVM) have affected dairy cattle breeding significantly around the world. This study examined the carrier frequency of BC, BLAD, FXID, and CVM autosomal recessive disorders in Bos taurus Holstein cows bred in the Altos Norte region of the state of Jalisco, Mexico. We extracted DNA from 408 random samples of peripheral blood, and then used polymerase chain reaction (PCR) to identify insertion mutations for FXID, and PCR with restriction fragment length polymorphism (PCR-RFLP) for CVM, BC and BLAD. We visualized the PCR products using agarose gel electrophoresis stained with GelRed®. We found that 100% of wild-type (N/N) allele homozygous animals for genes CD18, ASS, and FXI were free of the mutations for BLAD, BC and FXID respectively. For gene SLC35A3 we estimated total carrier frequency of 10.3% and allele frequency of 5%.(AU)

Carrier frequency of autosomal recessive disorders (BC, BLAD, FXID and CVM) in Holstein cows in Jalisco, Mexico

The hereditary autosomal recessive disorders bovine citrullinemia (BC), bovine leukocyte adhesion deficiency (BLAD), factor XI deficiency (FXID), and complex vertebral malformation (CVM) have affected dairy cattle breeding significantly around the world. This study examined the carrier frequency of BC, BLAD, FXID, and CVM autosomal recessive disorders in Bos taurus Holstein cows bred in the Altos Norte region of the state of Jalisco, Mexico. We extracted DNA from 408 random samples of peripheral blood, and then used polymerase chain reaction (PCR) to identify insertion mutations for FXID, and PCR with restriction fragment length polymorphism (PCR-RFLP) for CVM, BC and BLAD. We visualized the PCR products using agarose gel electrophoresis stained with GelRed®. We found that 100% of wild-type (N/N) allele homozygous animals for genes CD18, ASS, and FXI were free of the mutations for BLAD, BC and FXID respectively. For gene SLC35A3 we estimated total carrier frequency of 10.3% and allele frequency of 5%.(AU)

Carrier frequency of autosomal recessive disorders (BC, BLAD, FXID and CVM) in Holstein cows in Jalisco, Mexico

ABSTRACT: The hereditary autosomal recessive disorders bovine citrullinemia (BC), bovine leukocyte adhesion deficiency (BLAD), factor XI deficiency (FXID), and complex vertebral malformation (CVM) have affected dairy cattle breeding significantly around the world. This study examined the carrier frequency of BC, BLAD, FXID, and CVM autosomal recessive disorders in Bos taurus Holstein cows bred in the Altos Norte region of the state of Jalisco, Mexico. We extracted DNA from 408 random samples of peripheral blood, and then used polymerase chain reaction (PCR) to identify insertion mutations for FXID, and PCR with restriction fragment length polymorphism (PCR-RFLP) for CVM, BC and BLAD. We visualized the PCR products using agarose gel electrophoresis stained with GelRed®. We found that 100% of wild-type (N/N) allele homozygous animals for genes CD18, ASS, and FXI were free of the mutations for BLAD, BC and FXID respectively. For gene SLC35A3 we estimated total carrier frequency of 10.3% and allele frequency of 5%.

Adhesion of pathogenic bacteria to polystyrene, skin and gut mucus of gilthead seabream, infectious capacity and antibiotics susceptibility / Adesão de bactérias patogênicas a poliestireno, pele e muco intestinal de gilthead seabream, capacidade infecciosa e susceptibilidade a antibióticos

Linking proprieties of adhesion, infectious capacities and antibiotic resistance of pathogen bacteria could help to treat fish diseases. Adhesions of ten fish pathogenic bacteria were tested in microtiter plates vacant, coated with skin or gut mucus, fixed with methanol, stained with 2% crystal-violet and revealed by colorimetric method. Infectious capacity was performed by exposing gilthead seabream fibroblast cell line (SAF-1) to 107 -108 CFUmL-1 of pathogen bacteria. Cell viability was measured 3h, 9h and 20 hours post-infection. The sensitivity to antibiotics was executed by disk diffusion. Data showed that all the bacteria tested adhere to polystyrene. For skin mucus, Vibrio harveyi, Vibrio alginolyticus, Halomonas venusta, and Aeromonas bivalvium were moderately adherent. Dietzia maris was strongly adherent. For gut mucus, 60% of tested bacteria were weakly adherent and 40% were non adherent. For infection, D. maris, V. harveyi and A. bivalvium decreased the cells viability to 89% after only 3h. After 20h, the viability percentage ranged between 1% and 32%. All isolates presented resistance to 1000 mg ml-1 of sulphonamide, 60% were resistant to sulfonamide and penicillin G. Present findings could be relevant in fish aquaculture and underscore the importance of the linkage between adhesion, infectious capacity, and antibiotic susceptibility of pathogen bacteria to avoid fish diseases.

Estudar o link entre propriedades de adesão, capacidades infecciosas e resistência a antibióticos de bactérias patogênicas pode ajudar a tratar doenças de peixes. A adesão de dez bactérias patogênicas foi testada em placas de microtitulação vazias, revestidas com muco da pele ou do intestino, fixadas com metanol, coradas com 2% de violeta cristal e reveladas pelo modo colorimétrico. A capacidade infecciosa foi realizada expondo a linha celular de fibroblasto de dourada (SAF-1) a 107-108 CFUmL-1 de bactérias patogénicas. A viabilidade celular foi medida 3h, 9h e 20 horas após a infecção. A sensibilidade aos antibióticos foi executada por difusão em disco. Os dados mostram que todas as bactérias testadas aderem ao poliestireno. Para o muco cutâneo, Vibrio harveyi, Vibrio alginolyticus, Halomonas venusta e Aeromonas bivalvium foram moderadamente aderentes. Dietzia maris foi fortemente aderente. Para o muco intestinal, 60% das bactérias testadas eram fracamente aderentes e 40% não aderentes. Para infecção, D. maris, V. harveyi e A. bivalvium diminuíram a viabilidade celular para 89% após apenas 3h. Após 20h, o percentual de viabilidade variou entre 1% e 32%. Todos os isolados apresentaram resistência a 1000 mg mL-1 de sulfonamida, 60% foram resistentes à sulfonamida e à penicilina G. Os achados atuais podem ser relevantes na aqüicultura e ressaltam a importância da ligação entre adesão, capacidade infecciosa e suscetibilidade a antibióticos de bactérias patogênicas para evitar doenças em peixes.

Adhesion of pathogenic bacteria to polystyrene, skin and gut mucus of gilthead seabream, infectious capacity and antibiotics susceptibility / Adesão de bactérias patogênicas a poliestireno, pele e muco intestinal de gilthead seabream, capacidade infecciosa e susceptibilidade a antibióticos

Linking proprieties of adhesion, infectious capacities and antibiotic resistance of pathogen bacteria could help to treat fish diseases. Adhesions of ten fish pathogenic bacteria were tested in microtiter plates vacant, coated with skin or gut mucus, fixed with methanol, stained with 2% crystal-violet and revealed by colorimetric method. Infectious capacity was performed by exposing gilthead seabream fibroblast cell line (SAF-1) to 107 -108 CFUmL-1 of pathogen bacteria. Cell viability was measured 3h, 9h and 20 hours post-infection. The sensitivity to antibiotics was executed by disk diffusion. Data showed that all the bacteria tested adhere to polystyrene. For skin mucus, Vibrio harveyi, Vibrio alginolyticus, Halomonas venusta, and Aeromonas bivalvium were moderately adherent. Dietzia maris was strongly adherent. For gut mucus, 60% of tested bacteria were weakly adherent and 40% were non adherent. For infection, D. maris, V. harveyi and A. bivalvium decreased the cells viability to 89% after only 3h. After 20h, the viability percentage ranged between 1% and 32%. All isolates presented resistance to 1000 mg ml-1 of sulphonamide, 60% were resistant to sulfonamide and penicillin G. Present findings could be relevant in fish aquaculture and underscore the importance of the linkage between adhesion, infectious capacity, and antibiotic susceptibility of pathogen bacteria to avoid fish diseases.(AU)

Estudar o link entre propriedades de adesão, capacidades infecciosas e resistência a antibióticos de bactérias patogênicas pode ajudar a tratar doenças de peixes. A adesão de dez bactérias patogênicas foi testada em placas de microtitulação vazias, revestidas com muco da pele ou do intestino, fixadas com metanol, coradas com 2% de violeta cristal e reveladas pelo modo colorimétrico. A capacidade infecciosa foi realizada expondo a linha celular de fibroblasto de dourada (SAF-1) a 107-108 CFUmL-1 de bactérias patogénicas. A viabilidade celular foi medida 3h, 9h e 20 horas após a infecção. A sensibilidade aos antibióticos foi executada por difusão em disco. Os dados mostram que todas as bactérias testadas aderem ao poliestireno. Para o muco cutâneo, Vibrio harveyi, Vibrio alginolyticus, Halomonas venusta e Aeromonas bivalvium foram moderadamente aderentes. Dietzia maris foi fortemente aderente. Para o muco intestinal, 60% das bactérias testadas eram fracamente aderentes e 40% não aderentes. Para infecção, D. maris, V. harveyi e A. bivalvium diminuíram a viabilidade celular para 89% após apenas 3h. Após 20h, o percentual de viabilidade variou entre 1% e 32%. Todos os isolados apresentaram resistência a 1000 mg mL-1 de sulfonamida, 60% foram resistentes à sulfonamida e à penicilina G. Os achados atuais podem ser relevantes na aqüicultura e ressaltam a importância da ligação entre adesão, capacidade infecciosa e suscetibilidade a antibióticos de bactérias patogênicas para evitar doenças em peixes.(AU)

Surgery-induced changes in erythrocyte and plasma lipid peroxidation, enzymatic and non-enzymatic antioxidants of female rats: protective role of heparin and pentoxifylline

Background: Intra abdominal adhesions are a complication that occurs after many abdominal surgical procedures, especially gynecological operations. These complications occur by irritating the peritoneum due to such circumstances as infection or surgical trauma, and are considered a pathological part of the healing process of peritoneal injury. It manifests itself with symptoms such as pain, intestinal or urethral obstruction and abdominal abscesses. Oxidative stress due to adhesions plays an important role on adhesion formation. In addition to many researches done at the point of prevention of adhesion and decreasing stress parameters, in this study, it was planned to determine the effect of Heparin (H) and Pentoxifylline (PTX) on malondialdehyde (MDA) and some antioxidant values.Materials, Methods & Results: This study was performed on rats and thirty-seven female Sprague-Dawley rats were randomized into four groups. The first group was sham (Sh) group (n = 7) and laparotomy was performed and 2 mL of 0.9 % NaCl was applied. For all other rats (n = 30) the small intestine was withdrawn and the uterus was uncovered and the anti mesenteric surfaces of the left uterine horn and left abdominal wall were superficially tilted until slight bleeding was seen. Lesion areas have been covered. Two mL 0.9 % NaCl to control (C) group (n = 10), 500 IU heparin to group H, and 25 mg / kg Pentoxifylline to group PTX (n = 10) have been given and then the abdominal incision was closed. The adhesion score of group Sh was found to be more important than C and PTX groups (P < 0.05). The adhesion score of group C was determined to be more significant than group H (P < 0.05). In group Sh, erythrocyte reduced glutathione (GSH) levels were found to be more significant (P < 0.01) than C, H and PTX groups whereas it was found that group C was more significant than group H (P < 0.01).[...]

Surgery-induced changes in erythrocyte and plasma lipid peroxidation, enzymatic and non-enzymatic antioxidants of female rats: protective role of heparin and pentoxifylline

Background: Intra abdominal adhesions are a complication that occurs after many abdominal surgical procedures, especially gynecological operations. These complications occur by irritating the peritoneum due to such circumstances as infection or surgical trauma, and are considered a pathological part of the healing process of peritoneal injury. It manifests itself with symptoms such as pain, intestinal or urethral obstruction and abdominal abscesses. Oxidative stress due to adhesions plays an important role on adhesion formation. In addition to many researches done at the point of prevention of adhesion and decreasing stress parameters, in this study, it was planned to determine the effect of Heparin (H) and Pentoxifylline (PTX) on malondialdehyde (MDA) and some antioxidant values.Materials, Methods & Results: This study was performed on rats and thirty-seven female Sprague-Dawley rats were randomized into four groups. The first group was sham (Sh) group (n = 7) and laparotomy was performed and 2 mL of 0.9 % NaCl was applied. For all other rats (n = 30) the small intestine was withdrawn and the uterus was uncovered and the anti mesenteric surfaces of the left uterine horn and left abdominal wall were superficially tilted until slight bleeding was seen. Lesion areas have been covered. Two mL 0.9 % NaCl to control (C) group (n = 10), 500 IU heparin to group H, and 25 mg / kg Pentoxifylline to group PTX (n = 10) have been given and then the abdominal incision was closed. The adhesion score of group Sh was found to be more important than C and PTX groups (P < 0.05). The adhesion score of group C was determined to be more significant than group H (P < 0.05). In group Sh, erythrocyte reduced glutathione (GSH) levels were found to be more significant (P < 0.01) than C, H and PTX groups whereas it was found that group C was more significant than group H (P < 0.01).[...](AU)

Cell migration inhibition activity of a non-RGD disintegrin from Crotalus durissus collilineatus venom

In recent decades, snake venom disintegrins have received special attention due to their potential use in anticancer therapy. Disintegrins are small and cysteine-rich proteins present in snake venoms and can interact with specific integrins to inhibit their activities in cell-cell and cell-ECM interactions. These molecules, known to inhibit platelet aggregation, are also capable of interacting with certain cancer-related integrins, and may interfere in important processes involved in carcinogenesis. Therefore, disintegrin from Crotalus durissus collilineatus venom was isolated, structurally characterized and evaluated for its toxicity and ability to interfere with cell proliferation and migration in MDA-MB-231, a human breast cancer cell line. Methods: Based on previous studies, disintegrin was isolated by FPLC, through two chromatographic steps, both on reversed phase C-18 columns. The isolated disintegrin was structurally characterized by Tris-TricineSDS-PAGE, mass spectrometry and N-terminal sequencing. For the functional assays, MTT and wound-healing assays were performed in order to investigate cytotoxicity and effect on cell migration in vitro, respectively. Results: Disintegrin presented a molecular mass of 7287.4 Da and its amino acid sequence shared similarity with the disintegrin domain of P-II metalloproteases. Using functional assays, the disintegrin showed low cytotoxicity (15% and 17%, at 3 and 6 µg/mL, respectively) after 24 h of incubation and in the wound-healing assay, the disintegrin (3 µg/mL) was able to significantly inhibit cell migration (24%, p < 0.05), compared to negative control. Conclusion: Thus, our results demonstrate that non-RGD disintegrin from C. d. collilineatus induces low cytotoxicity and inhibits migration of human breast cancer cells. Therefore, it may be a very useful molecular tool for understanding ECM-cell interaction cancer-related mechanisms involved in an important integrin family that highlights molecular aspects of tumorigenesis. Also, non-RGD disintegrin has potential to serve as an agent in anticancer therapy or adjuvant component combined with other anticancer drugs.(AU)

Growth kinetics of Escherichia coli O157: H7 on the epicarp of fresh vegetables and fruits

Despite the increasing reports on the incidence of fresh vegetables and fruits as a possible vehicle for human pathogens, there is currently limited knowledge on the growth potential of Escherichia coli O157:H7 on different plant substrates. This study analyzed the selective adhesion and growth of E. coli O157:H7 on chili habanero (Capsicum chinense L.), cucumber (Cucumis sativus), radish (Raphanus sativus), tomato (Lycopersicon esculentum), beet (Beta vulgaris subsp. vulgaris), and onion (Allium cepa L.) under laboratory conditions. The Gompertz parameters were used to determine the growth kinetics. Scanning electron microscopy was used to visualize the adhesion of E. coli O157:H7 on the epicarp of the samples. Predictive models were constructed to compare the growth of E. coli O157:H7 on the samples with different intrinsic factors and to demonstrate the low selectivity of the pathogen. No significant difference was observed in the lag-phase duration (LPD), generation time (GT), and exponential growth rate (EGR) of the pathogen adhered to the samples. The interaction between the microorganism and the substrate was less supportive to the growth of E. coli O157:H7 for onion, whereas for tomato and cucumber, the time for the microorganism to attain the maximum growth rate (M) was significantly longer than that recorded for other samples.(AU)

Cell migration inhibition activity of a non-RGD disintegrin from Crotalus durissus collilineatus venom

Background:In recent decades, snake venom disintegrins have received special attention due to their potential use in anticancer therapy. Disintegrins are small and cysteine-rich proteins present in snake venoms and can interact with specific integrins to inhibit their activities in cell-cell and cell-ECM interactions. These molecules, known to inhibit platelet aggregation, are also capable of interacting with certain cancer-related integrins, and may interfere in important processes involved in carcinogenesis. Therefore, disintegrin from Crotalus durissus collilineatus venom was isolated, structurally characterized and evaluated for its toxicity and ability to interfere with cell proliferation and migration in MDA-MB-231, a human breast cancer cell line.Methods:Based on previous studies, disintegrin was isolated by FPLC, through two chromatographic steps, both on reversed phase C-18 columns. The isolated disintegrin was structurally characterized by Tris-Tricine-SDS-PAGE, mass spectrometry and N-terminal sequencing. For the functional assays, MTT and wound-healing assays were performed in order to investigate cytotoxicity and effect on cell migration in vitro, respectively.Results:Disintegrin presented a molecular mass of 7287.4 Da and its amino acid sequence shared similarity with the disintegrin domain of P-II metalloproteases. Using functional assays, the disintegrin showed low cytotoxicity (15% and 17%, at 3 and 6 μg/mL, respectively) after 24 h of incubation and in the wound-healing assay, the disintegrin (3 μg/mL) was able to significantly inhibit cell migration (24%, p < 0.05), compared to negative control.Conclusion:Thus, our results demonstrate that non-RGD disintegrin from C. d. collilineatus induces low cytotoxicity and inhibits migration of human breast cancer cells. Therefore, it may be a very useful molecular tool for understanding ECM-cell interaction cancer-related mechanisms involved in...(AU)

Hyaluronic acid and synovial fluid in preventing adhesion formation after tenorrhaphy: An in vivo study on rabbit Achilles tendon / Ácido hialurônico e líquido sinovial na prevenção da formação de aderências após tenorrafia: Um estudo in vivo sobre o tendão de Aquiles de coelho

Peritendinous adhesions are a significant problem in tendon surgery. One of the most preferred methods for preventing adhesion formation is anti-adhesive applications. This study aimed to investigate the efficacy of natural synovial fluid as an alternative viscoelastic agent for preventing adhesion in the healing period compared to hyaluronic acid. Thirty adult New Zealand rabbits were used in the study to form three experimental models. In all groups, a rupture was induced in the Achilles tendons. Following tenorrhaphy, hyaluronic acid was applied in one group and synovial fluid in the other group for anti-adhesive purposes. In the control group, no viscoelastic application was made. Bandage was applied to the operated extremities for three weeks post-operatively. At the end of five weeks, euthanasia was performed on the animals using pentobarbital sodium. Surgical sites in all groups were macroscopically examined for healing status as well as the presence and extent of adhesions. Biomechanical tests and histopathological examinations were then performed on the tendons in all groups. Findings established once again the positive contribution of hyaluronic acid to preventing adhesion formation as well as to healing and tensile strength in tendon surgery. Although, we found that it is possible to use synovial fluid, which is a natural source of hyaluronic acid, as a viscoelastic material, it is not superior to commercial hyaluronic acid preparation.(AU)

Asderências peritendinares são um problema significativo na cirurgia tendínea. Um dos métodos mais empregados para evitar a formação de adesão são as aplicações anti-adesivas. Este estudo tem como objetivo investigar a eficácia do líquido sinovial natural como um agente viscoelástico alternativo para prevenir a adesão no período de cicatrização comparado ao ácido hialurônico. Trinta coelhos adultos da Nova Zelândia foram utilizados no estudo para formar três modelos experimentais. Em todos os grupos, uma ruptura foi induzida nos tendões de Aquiles. Após tenorrafia, o ácido hialurônico foi aplicado em um grupo e o líquido sinovial no outro grupo para fins anti-adesivos. No grupo controle, não foi feita qualquer aplicação viscoelástica. A bandagem foi aplicada às extremidades operadas por três semanas pós-operatório. No final de cinco semanas, a eutanásia foi realizada nos animais utilizando pentobarbital sódico. Os locais cirúrgicos em todos os grupos foram examinados macroscopicamente quanto ao estado de cura, bem como a presença e extensão das aderências. Testes biomecânicos e exames histopatológicos foram realizados nos tendões em todos os grupos. Os achados estabeleceram mais uma vez a contribuição positiva do ácido hialurônico para prevenir a formação de aderências, bem como para a cicatrização e resistência à tração na cirurgia do tendão. Embora se tenha constatado que é possível utilizar fluido sinovial, que é uma fonte natural de ácido hialurónico, como material viscoelástico, não é superior à preparação de ácido hialurónico comercial.(AU)