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)

Effects of different concentrations of sodium azide and chloramphenicol on the preservation of raw milk samples / Efeitos de diferentes concentrações de azida sódica e cloranfenicol na preservação de amostras de leite cru

The preservation of milk samples for microbiological analyses by the Brazilian Network of Milk Quality Control Laboratories requires the addition of preservatives to maintain the microbiota from the time of sample collection to the moment of analysis. The number of microorganisms can change as a result of the active ingredients and concentration of the preservative, as well as due to interactions between the preservatives, incubation time, and packaging temperature. The objective of this research was to evaluate the conservation potential of different concentrations of sodium azide and chloramphenicol on the analytical shelf life of milk samples. Two farms were selected, one with a low bacterial count and one with a high bacterial count. The milk was dispensed into sterile vials and tested after the addition of the usual concentrations of sodium azide and chloramphenicol, doubled concentrations, tripled concentrations, and as a control, without preservatives. The samples were incubated at 3 ± 1 °C, 6 ± 1 °C, and 9 ± 1 °C for 14 days and analyzed daily for their bacterial count by flow cytometry. The tripled preservative concentrations improved conservation, increasing the timespan of the analytical viability of the samples without altering the results.(AU)

A conservação das amostras de leite destinadas para análises microbiológicas pela Rede Brasileira de Laboratórios de Controle da Qualidade do Leite requer adição de conservantes para a preservação da microbiota existente desde o momento da coleta até as análises. O número de microrganismos pode apresentar alterações decorrentes do princípio ativo e concentração do conservante, e ainda entre as interações conservante, tempo de incubação e temperatura de acondicionamento. O objetivo deste trabalho foi avaliar o potencial de conservação de diferentes concentrações de azida sódica e cloranfenicol sobre a vida útil analítica de amostras de leite. Foram selecionadas duas fazendas, sendo uma com baixa contagem bacteriana e outra com alta contagem bacteriana. O leite foi fracionado em frascos estéreis e testado nas seguintes condições: pastilhas com a concentração usual de azida sódica e cloranfenicol, com dupla concentração, com tripla concentração e sem a adição do conservante. As amostras foram incubadas por quatorze dias a 3 ± 1 °C, 6 ± 1 °C e 9 ± 1 °C, e analisadas diariamente por citometria de fluxo para a determinação da contagem bacteriana. A tripla concentração do conservante demonstrou maior conservação, possibilitando o aumento da viabilidade analítica das amostras sem alteração nos resultados.(AU)

Biofilm formation by Salmonella Enteritidis at different incubation temperatures

Background: The genus Salmonella, associated with poultry products, is considered the leading cause of foodborne outbreaks in humans in many countries. In Brazil, Salmonella Enteritidis (SE) is the serovar remains as one most frequentlyisolated from humans, and it is also a major serovar found in animals, food, animal feed, and environmental samples, despiteall the efforts to control this pathogen. Also, the bacterium is able to form biofilms on different surfaces, protecting cellsfrom both cleaning and sanitizing procedures in the food industries. This study aimed to verify the ability of SalmonellaEnteritidis isolates to form biofilm on polystyrene at different incubation temperatures.Materials, Methods & Results: A total of 171 SE samples were isolated from foodborne outbreaks (foods and stool cultures) and poultry products between 2003 and 2010. The biofilm-forming ability of samples was measured at four differenttemperatures (3°C, 9ºC, 25ºC, and 36ºC), for 24 h, simulating temperatures usually found in poultry slaughterhouses. Later,200 μL of each bacterial suspension was inoculated, in triplicate, onto 96-well, flat-bottomed sterile polystyrene microtiterplates, washed, after that, the biofilm was fixed with methanol. The plates were dried at ambient temperature, stained with2% Hucker’s crystal violet. Afterwards, absorbance was read using an ELISA plate reader and the optical density (OD)of each isolate was obtained by the arithmetic mean of the absorbance of three wells and this value was compared withthe mean absorbance of negative controls (ODnc). The following classification was used for the determination of biofilmformation: no biofilm production, weak biofilm production, moderate biofilm production and strong biofilm production.Results demonstrated all isolates from stool cultures and foods involved in foodborne outbreaks, at least one of the fourtemperatures tested, were able to form biofilm...

Quantificação de Staphylococcus aureus e bactérias mesófilas aeróbias para avaliar higienização de equipamentos de ordenha / Staphylococcus aureus and mesophilic aerobic bacteria quantification in hygienization process of milking equipment

Background: Milk’s composition is an excellent substrate for microorganism’s multiplication. Presence of Staphylococcusaureus and aerobic mesophilic bacteria are one of the most common problems in dairy farms. On dairy industry’s and milkfarms Clean in Place (CIP) system higyenization are commonly used, then the cleaning occurs as a closed process, forbetter results sanitizans are applied, in order to obtain a safety food. This project aim to evaluate Staphylococcus aureusand aerobic mesophilic bacteria reduction after two milking higyenization process.Materials, Methods & Results: This research was done on a Rio Grande do Sul North Milk farm, with mechanized milkingand Clean in Place system for cleaning. For liners and CIP tubes higyenization commercial products as Sodium Hipoclorite3% and phosphoric acid 11.3% are used for detergency, and peracetic acid 5% for sanitization. Milk bunk tank are higyenized with sodium hypoclorite 3.8% alcalin detergent. After higyenization steps liners, CIP’s water process, bulk milk tankand milk set were collected. At process 1, liners and CIP water were collected after milking, detergency and sanitizationthat occurred immediately at the detergency’s finish, while process 2 the sanitization was realized 8 h after detergency,before following milking. Cooling milk bulk tank was collected before and after detergency, and milk set after milkingsConvencional microbiology were used to count and results in log10 UFC.cm-2...

Biofilm formation by Salmonella Enteritidis at different incubation temperatures

Background: The genus Salmonella, associated with poultry products, is considered the leading cause of foodborne outbreaks in humans in many countries. In Brazil, Salmonella Enteritidis (SE) is the serovar remains as one most frequentlyisolated from humans, and it is also a major serovar found in animals, food, animal feed, and environmental samples, despiteall the efforts to control this pathogen. Also, the bacterium is able to form biofilms on different surfaces, protecting cellsfrom both cleaning and sanitizing procedures in the food industries. This study aimed to verify the ability of SalmonellaEnteritidis isolates to form biofilm on polystyrene at different incubation temperatures.Materials, Methods & Results: A total of 171 SE samples were isolated from foodborne outbreaks (foods and stool cultures) and poultry products between 2003 and 2010. The biofilm-forming ability of samples was measured at four differenttemperatures (3°C, 9ºC, 25ºC, and 36ºC), for 24 h, simulating temperatures usually found in poultry slaughterhouses. Later,200 μL of each bacterial suspension was inoculated, in triplicate, onto 96-well, flat-bottomed sterile polystyrene microtiterplates, washed, after that, the biofilm was fixed with methanol. The plates were dried at ambient temperature, stained with2% Huckers crystal violet. Afterwards, absorbance was read using an ELISA plate reader and the optical density (OD)of each isolate was obtained by the arithmetic mean of the absorbance of three wells and this value was compared withthe mean absorbance of negative controls (ODnc). The following classification was used for the determination of biofilmformation: no biofilm production, weak biofilm production, moderate biofilm production and strong biofilm production.Results demonstrated all isolates from stool cultures and foods involved in foodborne outbreaks, at least one of the fourtemperatures tested, were able to form biofilm...(AU)

Quantificação de Staphylococcus aureus e bactérias mesófilas aeróbias para avaliar higienização de equipamentos de ordenha / Staphylococcus aureus and mesophilic aerobic bacteria quantification in hygienization process of milking equipment

Background: Milks composition is an excellent substrate for microorganisms multiplication. Presence of Staphylococcusaureus and aerobic mesophilic bacteria are one of the most common problems in dairy farms. On dairy industrys and milkfarms Clean in Place (CIP) system higyenization are commonly used, then the cleaning occurs as a closed process, forbetter results sanitizans are applied, in order to obtain a safety food. This project aim to evaluate Staphylococcus aureusand aerobic mesophilic bacteria reduction after two milking higyenization process.Materials, Methods & Results: This research was done on a Rio Grande do Sul North Milk farm, with mechanized milkingand Clean in Place system for cleaning. For liners and CIP tubes higyenization commercial products as Sodium Hipoclorite3% and phosphoric acid 11.3% are used for detergency, and peracetic acid 5% for sanitization. Milk bunk tank are higyenized with sodium hypoclorite 3.8% alcalin detergent. After higyenization steps liners, CIPs water process, bulk milk tankand milk set were collected. At process 1, liners and CIP water were collected after milking, detergency and sanitizationthat occurred immediately at the detergencys finish, while process 2 the sanitization was realized 8 h after detergency,before following milking. Cooling milk bulk tank was collected before and after detergency, and milk set after milkingsConvencional microbiology were used to count and results in log10 UFC.cm-2...(AU)

Salmonella Enteritidis forms biofilm under low temperatures on different food industry surfaces / Salmonella Enteritidis forma biofilme sob baixas temperaturas em diferentes superfícies da indústria de alimentos

We evaluated the influence of temperature on the ability of Salmonella Enteritidis (SE) to form biofilms on stainless steel, polyethylene, and polyurethane surfaces under different hygiene procedures. These materials were placed on SE culture and incubated at 42±1 ºC, 36±1 ºC, 25±1 ºC, 9±1 ºC, and 3±1 ºC for 4, 8, 12, and 24 h. Hot water at 45 ºC and 85 ºC, 0.5% peracetic acid solution, and 1% quaternary ammonia were used for hygienization. Biofilm formation occurred at all temperatures evaluated, highlighting at 3 ºC which has not been reported as an ideal temperature for the adhesion of SE to these materials. The SE adhered more often to polyethylene surfaces than to polyurethane and stainless steel surfaces (P<0.05). Peracetic acid and water at 85 ºC had similar hygienization efficiency (P<0.05) followed by quaternary ammonia whereas water at 45 ºC was not effective. SE adhered to these materials under low temperatures which to date have been deemed safe for food preservation.(AU)

Avaliou-se o efeito da temperatura na capacidade de Salmonella Enteritidis (SE) formar biofilme em superfícies de aço inoxidável, polietileno e poliuretano e diferentes processos de higienização. Corpos de prova destes materiais foram postos frente a culturas de SE e incubados a 42±1 ºC, 36±1 ºC, 25±1 ºC, 9±1 ºC e 3±1 ºC por 4, 8, 12 e 24 horas. Para a higienização foram testados água aquecida a 45ºC e 85 ºC e soluções de ácido peracético 0,5% e amônia quaternária 1%. Verificou-se a formação de biofilmes em todas as temperaturas avaliadas, ressaltando-se a 3 ºC, ainda não citada como propícia para adesão de SE. Houve maior adesão ao polietileno do que ao poliuretano e ao aço inoxidável (P<0.05). Para higienização, o ácido peracético e a água a 85 ºC tiveram ação semelhante (P<0.05), seguidos por amônia quaternária, enquanto que a água a 45 ºC não foi eficaz. Todos os materiais avaliados propiciaram a aderência de SE, mesmo sob temperaturas baixas, consideradas até então seguras para a conservação dos alimentos.(AU)

ATP-Bioluminescence and conventional microbiology for hygiene evaluation of cutting room surfaces in poultry slaughterhouse

Background: The hygiene procedures in poultry slaughterhouses consist in the use of hot water, detergent and sanitizing, configuring Sanitation Standard Operating Procedure (SSOP). These actions control contamination in food processing environments, especially by pathogenic microorganisms, which cause diseases with impact on public health and economic losses. The microbiological control of aerobic mesophiles, Staphylococcus aureus and Escherichia coli, are used as indicators of contamination. The hygienic-sanitary conditions on the surfaces of the poultry slaughterhouse cuttting room were evaluated, before and after cleaning and sanitizing procedures.Materials, Methods & Results: Conventional microbiology (Rodac plates and sponge for quantification of aerobic mesophiles, Staphylococcus aureus and Escherichia coli) and ATP-Bioluminescence were used to analyze the action of hot water and the active principles peracetic acid, quaternary ammonia and biguanide in the standard pre-operational hygiene procedure in the cutting room of the poultry slaughterhouse under Federal Inspection with slaughter capacity of more than 20.000 birds/h. The evaluations were performed on three lines of chicken thigh cuts at the same time and in a completely randomized manner on stainless steel surfaces, polyurethane belts and polyethylene boards. Samples were made in four replicates at the three surface totaling 108 assay for each microorganism. The samples were collected at the end of the cutting process, before and after washing the surfaces with hot water (between 45 and 50ºC) and after sanitization with 0.5% peracetic acid, 2% quaternary ammonia and 1% biguanide. The ATP-Bioluminescence method detected organic matter at all collected points and Rodac plates allowed a better recovery of microorganisms than sponges for quantification of aerobic mesophiles, E. coli and S. aureus.[...]

ATP-Bioluminescence and conventional microbiology for hygiene evaluation of cutting room surfaces in poultry slaughterhouse

Background: The hygiene procedures in poultry slaughterhouses consist in the use of hot water, detergent and sanitizing, configuring Sanitation Standard Operating Procedure (SSOP). These actions control contamination in food processing environments, especially by pathogenic microorganisms, which cause diseases with impact on public health and economic losses. The microbiological control of aerobic mesophiles, Staphylococcus aureus and Escherichia coli, are used as indicators of contamination. The hygienic-sanitary conditions on the surfaces of the poultry slaughterhouse cuttting room were evaluated, before and after cleaning and sanitizing procedures.Materials, Methods & Results: Conventional microbiology (Rodac plates and sponge for quantification of aerobic mesophiles, Staphylococcus aureus and Escherichia coli) and ATP-Bioluminescence were used to analyze the action of hot water and the active principles peracetic acid, quaternary ammonia and biguanide in the standard pre-operational hygiene procedure in the cutting room of the poultry slaughterhouse under Federal Inspection with slaughter capacity of more than 20.000 birds/h. The evaluations were performed on three lines of chicken thigh cuts at the same time and in a completely randomized manner on stainless steel surfaces, polyurethane belts and polyethylene boards. Samples were made in four replicates at the three surface totaling 108 assay for each microorganism. The samples were collected at the end of the cutting process, before and after washing the surfaces with hot water (between 45 and 50ºC) and after sanitization with 0.5% peracetic acid, 2% quaternary ammonia and 1% biguanide. The ATP-Bioluminescence method detected organic matter at all collected points and Rodac plates allowed a better recovery of microorganisms than sponges for quantification of aerobic mesophiles, E. coli and S. aureus.[...](AU)

Salmonella spp. isolated by miniaturized most probable number and conventional microbiology in poultry slaughterhouses

Background: Salmonella spp. are frequently isolated from fowls, and their detection in poultry products varies according to the breeding system and the slaughtering process, bringing risks to the consumer and compromising the marketability. The control of Salmonella in poultry slaughterhouses is based on the detection of bacteria, but the quantification of the agent would be important in assessing risk, as well as in obtaining data to determine the capacity of each step of the process to decrease or increase bacterial contamination. The aims of this study were to propose a method for the quantification of Salmonella in poultry slaughterhouses, frequency of isolation and serovars identified. Materials, Methods & Results: Twenty-one broiler flocks from seven federally inspected slaughterhouses in southern Brazil, totaling 1,071 samples, were assessed by miniaturized most probable number (mMPN) and conventional microbiology. The samples were collected in triplicate at 17 points, which included cloacae, transportation cages before and after sanitization, water (scald tank, supply, pre-chiller and chiller), and carcasses (before and after scalding, defeathering, rinsing, evisceration, final rinsing, chilling at 4ºC, and freezing at -12C for 24 h, 30 and 60 days). Typical Salmonella colonies were submitted to TSI, LIA, SIM, urea, and polyvalent anti-O antiserum tests [...](AU)

Salmonella spp. isolated by miniaturized most probable number and conventional microbiology in poultry slaughterhouses

Background: Salmonella spp. are frequently isolated from fowls, and their detection in poultry products varies according to the breeding system and the slaughtering process, bringing risks to the consumer and compromising the marketability. The control of Salmonella in poultry slaughterhouses is based on the detection of bacteria, but the quantification of the agent would be important in assessing risk, as well as in obtaining data to determine the capacity of each step of the process to decrease or increase bacterial contamination. The aims of this study were to propose a method for the quantification of Salmonella in poultry slaughterhouses, frequency of isolation and serovars identified. Materials, Methods & Results: Twenty-one broiler flocks from seven federally inspected slaughterhouses in southern Brazil, totaling 1,071 samples, were assessed by miniaturized most probable number (mMPN) and conventional microbiology. The samples were collected in triplicate at 17 points, which included cloacae, transportation cages before and after sanitization, water (scald tank, supply, pre-chiller and chiller), and carcasses (before and after scalding, defeathering, rinsing, evisceration, final rinsing, chilling at 4ºC, and freezing at -12C for 24 h, 30 and 60 days). Typical Salmonella colonies were submitted to TSI, LIA, SIM, urea, and polyvalent anti-O antiserum tests [...]

The use of vortex and ultrasound techniques for the in vitro removal of Salmonella spp. Biofilms

Background: The presence of biofilm is common to all types of surfaces, such as stainless steel. Once formed, biofilms act as a point of constant contamination releasing fragments or planktonic cells of microorganisms, such as Salmonella spp., and may impair the microbiological quality of products. The laboratory methods are being used and tested in vitro for removal and the subsequent quantification of biofilms, including the use of vortex and ultrasound. The aim of the study was to evaluate the effectiveness of these two laboratory methods for removing Salmonella spp. biofilms, in vitro cultured on stainless steel surface from the food industry.Materials, Methods & Results: Three strains were analyzed for biofilm formation by Salmonella spp., and they are S. Typhimurium ATCC 14028, S. Enteritidis ATCC 13076 and S. Enteritidis isolated from drag swab on poultry farm and genetically confirmed by Microarray, denominated P106. Coupons of stainless steel AISI 316, with an area of 1 cm² were used. These materials used for the coupons were obtained from the equipments at the cutting room in the poultry slaughterhouse. Biofilms were formed using TSB broth without glucose and incubated at 36°C for 24 h. Six replicates were performed for each microorganism in each removal method. After the biofilm formation, two methods were used in the removal stage, the vortexing, performed for 2 min, and the sonication method, with coupons maintained for 10 min in an ultrasound bath, at a frequency of 40 kHz and potency 81 W. Serial dilutions were made and transferred to PCA agar for quantification in log10CFU.mL-1. The microtopography was performed by scanning electron microscopy (SEM) of surfaces before the removal step. Statistical analysis of the results showed no significant difference between the two removal methods and between the three strains studied (P > 0.05).[...](AU)

The use of vortex and ultrasound techniques for the in vitro removal of Salmonella spp. Biofilms

Background: The presence of biofilm is common to all types of surfaces, such as stainless steel. Once formed, biofilms act as a point of constant contamination releasing fragments or planktonic cells of microorganisms, such as Salmonella spp., and may impair the microbiological quality of products. The laboratory methods are being used and tested in vitro for removal and the subsequent quantification of biofilms, including the use of vortex and ultrasound. The aim of the study was to evaluate the effectiveness of these two laboratory methods for removing Salmonella spp. biofilms, in vitro cultured on stainless steel surface from the food industry.Materials, Methods & Results: Three strains were analyzed for biofilm formation by Salmonella spp., and they are S. Typhimurium ATCC 14028, S. Enteritidis ATCC 13076 and S. Enteritidis isolated from drag swab on poultry farm and genetically confirmed by Microarray, denominated P106. Coupons of stainless steel AISI 316, with an area of 1 cm² were used. These materials used for the coupons were obtained from the equipments at the cutting room in the poultry slaughterhouse. Biofilms were formed using TSB broth without glucose and incubated at 36°C for 24 h. Six replicates were performed for each microorganism in each removal method. After the biofilm formation, two methods were used in the removal stage, the vortexing, performed for 2 min, and the sonication method, with coupons maintained for 10 min in an ultrasound bath, at a frequency of 40 kHz and potency 81 W. Serial dilutions were made and transferred to PCA agar for quantification in log10CFU.mL-1. The microtopography was performed by scanning electron microscopy (SEM) of surfaces before the removal step. Statistical analysis of the results showed no significant difference between the two removal methods and between the three strains studied (P > 0.05).[...]

Antimicrobial resistance profile of Salmonella Heidelberg isolated from a poultry slaughterhouse in 2005 and 2009 / Perfil de resistência a antimicrobianos por Salmonella Heidelberg isoladas de abatedouro avícola em 2005 e 2009

Background: Salmonella spp. is recognized as being one of the most common bacterial causes of food-borne illness spreadon poultry production due to easy adaptation to environment and diffi cult to eradicate. In poultry production system antimicrobials are added in feed as growth promoters in continuous and sub-therapeutic doses, inducing a selective pressure andconsequent antimicrobial resistance. This management causes public health problems to disseminate resistant pathogensthrough food chain and reduce the options of treatment of bacterial infections.Materials, Methods & Results: The samples were isolated in a poultry slaughterhouse under Federal Inspection in amonitoring, research and quantifi cation project of Salmonella spp in critical control points in slaughterhouse. Adaptedmethodology was used for quantifi cation of Salmonella as follows: swabs and cages were placed in 50 mL of peptonewater buffered 1% (PW 1%) and incubated at 37ºC for 12 h; for the analysis of water 100 mL were inoculated in 50 mL ofpeptone water buffered in triple concentration and incubated at 37ºC for 12 h; the chickens and carcasses were packed insterile bags with a capacity of 4000 mL, added 150 mL of peptone water buffered 1%, agitated manually for one minuteand the rinsing broth incubated by 12 h at 37ºC. After hatching were made decimal dilutions Rapapport Vassiliadis broth(RV), inoculated 1 mL in 9 mL of RV broth until 10-3 dilution and incubation for 12 h at 41°C in a water bath with agitation. After this period 100 µL of RV broth were seeded in Agar Rambach and Agar XLD and the plates incubated at 37ºCfor 12 h. Salmonella-like growth were placed in Agar Rambach and confi rmed as Salmonella to biochemical tests (TSI,LIA, urea broth) and assayed for polyvalent antiserum to Salmonella. The fi nal identifi cation of the samples was carriedout by the Polymerase Chain Reaction (PCR, Premi® Test Salmonella DSM). Were selected 20 samples of Salmonella...(AU)

Isolamento de Salmonella na tecnologia de abate de suínos como subsídio para o sistema appcc / Salmonella isolation in a pig slaughtering technology as allowance for the system haccp / Aislamiento de Salmonella en la tecnología de matanza de porcinos como subsidio para el sistema appcc

A Salmonella pode estar presente em carcaças suínas ao final da tecnologia de abate, sendo que não existem procedimentos de inspeção especificamente direcionados para o controle desta bactéria, consideradas um risco potencial para a saúde pública. O objetivo do trabalho foi verificar a presença de Salmonella em carcaças suínas e caracterizar os pontos críticos de controle (PCCs) da tecnologia de abate como subsídio ao sistema APPCC. Foram realizados swabs de carcaças após a escaldagem/depilação, antes da evisceração, após evisceração e serragem da carcaça e após 24 horas de refrigeração, gerando 120 amostras (30 carcaças de cinco lotes). Isolou-se Salmonella em 53,3% dos PCs analisados, sendo 40% antes da eviscerarão e 30% após a escaldagem/depilação e evisceração/serragem. Nas carcaças resfriadas por 24 horas não foi isolada Salmonella. Os sorovares identificados foram Typhimurium (96,7 %) e Panama (3,22%). O isolamento de Salmonella reforça a necessidade de programas de monitoria da bactéria, desde as granjas até os abatedouros, enquanto o não isolamento nas carcaças resfriadas indica a importância da lavagem final e adequação da cadeia do frio, o que minimiza a contaminação pela intensificação das medidas de higiene e controle de pontos críticos para subsidiar o sistema APPCC.(AU)

Salmonella can be present in swine carcasses at the end of slaughtering and there is no specifically guided inspection procedure for the control of this bacterium, considered a potential risk for public health. The objective of this paper is to verify the presence of Salmonella in carcasses and to characterize the critical points of control (CPCs) of the slaughtering technology as a subsidy to the HACCP system. The carcasses were swabbed at different points during the slaughtering process, ant after 24 hours of refrigeration, generating 120 samples (30 carcasses from five different batches). Salmonella was isolated in 53.3% of the CPs analyzed, being 40% before evisceration and 30% after scalding/depilation and evisceration/ sawing. No Salmonella was isolated in the carcasses refrigerated for 24 hours. Typhimurium (96.7%) and Panama (3.22%) were the serovars identified. Salmonella isolation indicates the need of programs to monitor the bacterium from the farms to the slaughterhouse, while the non-isolation in the refrigerated carcasses shows the relevance of the cold chain, which minimizes the contamination by the intensification of hygiene measures and the control of critical points to subsidize the HACCP system.(AU)

La Salmonella puede estar presente en carcasas porcinas al final de la matanza, siendo que no existen procedimientos de inspección específicamente direccionados para el control de esta bacteria, consideradas un riesgo potencial a la salud pública. El objetivo del trabajo ha sido verificar la presencia de Salmonella en carcasas porcinas y caracterizar los puntos críticos de control (PCCs) en la tecnología de matanza como subsidio al sistema APPCC. Se han realizado swabs de carcasas tras escaldadura/depilación, antes de evisceración, después de la evisceración y serrín de la carcasa y después de 24 horas de refrigeración, generando 120 muestras (30 carcasas de cinco lotes) Se ha aislado Salmonella en 53,3% de los PCs analizados, siendo 40% antes de evisceración y 30% después de la escaldadura/depilación y evisceración/serrín. En las carcasas enfriadas por 24 horas no se ha aislado Salmonella. Los serovares identificados fueron Typhimurium (96,7%) y Panama (3,22%). El aislamiento de Salmonella refuerza la necesidad de programas de monitoria de la bacteria, desde las granjas hasta los mataderos, el no aislamiento en las carcasas enfriadas indica la importancia del lavado final y adecuación de la cadena del frío, lo que minimiza la contaminación por la intensificación de las medidas de higiene y control de puntos críticos para subsidiar el sistema APPCC(AU)

Antimicrobial resistance profile of Salmonella Heidelberg isolated from a poultry slaughterhouse in 2005 and 2009 / Perfil de resistência a antimicrobianos por Salmonella Heidelberg isoladas de abatedouro avícola em 2005 e 2009

Background: Salmonella spp. is recognized as being one of the most common bacterial causes of food-borne illness spreadon poultry production due to easy adaptation to environment and diffi cult to eradicate. In poultry production system antimicrobials are added in feed as growth promoters in continuous and sub-therapeutic doses, inducing a selective pressure andconsequent antimicrobial resistance. This management causes public health problems to disseminate resistant pathogensthrough food chain and reduce the options of treatment of bacterial infections.Materials, Methods & Results: The samples were isolated in a poultry slaughterhouse under Federal Inspection in amonitoring, research and quantifi cation project of Salmonella spp in critical control points in slaughterhouse. Adaptedmethodology was used for quantifi cation of Salmonella as follows: swabs and cages were placed in 50 mL of peptonewater buffered 1% (PW 1%) and incubated at 37ºC for 12 h; for the analysis of water 100 mL were inoculated in 50 mL ofpeptone water buffered in triple concentration and incubated at 37ºC for 12 h; the chickens and carcasses were packed insterile bags with a capacity of 4000 mL, added 150 mL of peptone water buffered 1%, agitated manually for one minuteand the rinsing broth incubated by 12 h at 37ºC. After hatching were made decimal dilutions Rapapport Vassiliadis broth(RV), inoculated 1 mL in 9 mL of RV broth until 10-3 dilution and incubation for 12 h at 41°C in a water bath with agitation. After this period 100 µL of RV broth were seeded in Agar Rambach and Agar XLD and the plates incubated at 37ºCfor 12 h. Salmonella-like growth were placed in Agar Rambach and confi rmed as Salmonella to biochemical tests (TSI,LIA, urea broth) and assayed for polyvalent antiserum to Salmonella. The fi nal identifi cation of the samples was carriedout by the Polymerase Chain Reaction (PCR, Premi® Test Salmonella DSM). Were selected 20 samples of Salmonella...

Biofilm former Salmonella enteritidis are multiresistant to antibiotics / Salmonella Enteritidis formadoras de biofilmes são multirresistentes a antimicrobianos

Background: The Salmonella Enteritidis is one of the most isolated pathogens in outbreaks of foodborne illness, whichcan occur due to various factors such as cooking temperature, inadequate storage and cross-contamination. The choice ofthe appropriate disinfectant in food industries is essential to prevent the spread of contamination and control of biofilmson surfaces. It is also extremely important the concern with resistance to antimicrobials used both as growth promotersand in human and animal treatments, which may generate a selective pressure favoring the emergence of resistant bacteria.Materials, Methods & Results: Twenty samples of Salmonella Enteritidis were tested, 10 from outbreaks of foodbornediseases and 10 of poultry origin, as for the formation of biofilms, antibiotic resistance and sanitizers. The samples werestored frozen in BHI with 20% glycerol. For reactivation were incubated in BHI broth, plated on XLD agar and subsequentlyperformed biochemical tests to check purity. Firstly were evaluated for biofilm formation on polystyrene at temperature of36 ± 1ºC. We tested the sanitizing resistance to biguanide concentrations 0.6%, 1.0% and 1.5%, peracetic acid at concentrations 0.1%, 0.5% and 1.0%, and quaternary ammonia at concentrations of 0.3%, 1.0% and 2.0%. For tests of antimicrobialresistance the cultures were evaluated front 10 μg ampicillin, 30 μg cephalexin, 30 μg chloramphenicol, 5 μg enrofloxacin,15 μg erythromycin, 30 μg neomycin, 25 μg sulphazotrim, 300 μg sulfonamides. According to the results, 25% of sampleswere strongly biofilm formers, 35% moderately formers, 35% weakly formers and 10% not biofilm formers. In sanitizers, quaternary ammonia and peracetic acid were effective at all concentrations and at all times, but tests with biguanide...(AU)

Biofilm former Salmonella enteritidis are multiresistant to antibiotics / Salmonella Enteritidis formadoras de biofilmes são multirresistentes a antimicrobianos

Background: The Salmonella Enteritidis is one of the most isolated pathogens in outbreaks of foodborne illness, whichcan occur due to various factors such as cooking temperature, inadequate storage and cross-contamination. The choice ofthe appropriate disinfectant in food industries is essential to prevent the spread of contamination and control of biofilmson surfaces. It is also extremely important the concern with resistance to antimicrobials used both as growth promotersand in human and animal treatments, which may generate a selective pressure favoring the emergence of resistant bacteria.Materials, Methods & Results: Twenty samples of Salmonella Enteritidis were tested, 10 from outbreaks of foodbornediseases and 10 of poultry origin, as for the formation of biofilms, antibiotic resistance and sanitizers. The samples werestored frozen in BHI with 20% glycerol. For reactivation were incubated in BHI broth, plated on XLD agar and subsequentlyperformed biochemical tests to check purity. Firstly were evaluated for biofilm formation on polystyrene at temperature of36 ± 1ºC. We tested the sanitizing resistance to biguanide concentrations 0.6%, 1.0% and 1.5%, peracetic acid at concentrations 0.1%, 0.5% and 1.0%, and quaternary ammonia at concentrations of 0.3%, 1.0% and 2.0%. For tests of antimicrobialresistance the cultures were evaluated front 10 μg ampicillin, 30 μg cephalexin, 30 μg chloramphenicol, 5 μg enrofloxacin,15 μg erythromycin, 30 μg neomycin, 25 μg sulphazotrim, 300 μg sulfonamides. According to the results, 25% of sampleswere strongly biofilm formers, 35% moderately formers, 35% weakly formers and 10% not biofilm formers. In sanitizers, quaternary ammonia and peracetic acid were effective at all concentrations and at all times, but tests with biguanide...