RESUMO: Biocidas e/ou antibióticos em concentrações sub-inibitórias podem promover o desenvolvimento de resistência adaptativa ou mesmo resistência cruzada nos micro-organismos. Entretanto, estudos destas respostas após o tratamento com a prata são escassos na literatura. Compostos a base de prata, incluindo as nanopartículas de prata (Ag-NPs), podem ser uma alternativa na prevenção e/ou tratamento de mastite bovina. Assim, este trabalho objetivou determinar o efeito de doses sub-inibitórias de Ag-NPs e dos íons Ag+ provenientes do nitrato de prata (AgNO3) sobre isolados de Staphylococcus aureus e de Escherichia coli, provenientes de leite de vacas com mastite. As Ag-NPs foram sintetizadas por redução química utilizando AgNO3 e citrato de sódio e a Concentração Mínima Inibitória (CMI) das Ag-NPs e íons Ag+ nos patógenos da mastite foi determinada. Os isolados foram expostos a concentrações sub-inibitórias de Ag-NPs ou de AgNO3 por 10 dias consecutivos para verificar o desenvolvimento de resistência adaptativa à prata pela mudança no valor da CMI, e de resistência cruzada com antibióticos pela mudança no perfil de sensibilidade em relação ao controle. AgNO3 apresentou-se mais efetivo contra todos os isolados. Não foi verificada alteração no valor da CMI nem do perfil de sensibilidade aos antibióticos, indicando que não houve seleção de resistência adaptativa à prata e de resistência cruzada aos antibióticos pelos micro-organismos estudados. O uso potencial da prata apresentado nos resultados é favorável à continuidade dos estudos objetivando a elaboração de terapias à base de prata para o tratamento da mastite bovina.

ABSTRACT: Biocides and/or antibiotics used in subinhibitory concentrations can promote the development of adaptive resistance or even cross-resistance in microorganisms. However, studies on these responses following silver treatments are scarce in the literature. Silver-based compounds, including silver nanoparticles (Ag-NPs), can be an alternative in the prevention and treatment of bovine mastitis. Thus, this research evaluated the effect of subinhibitory dosages of Ag-NPs and Ag+ ions from silver nitrate (AgNO3) on Staphylococcus aureus and Escherichia coli isolated from milk of cows with mastitis. Ag-NPs were synthesized by chemical reduction using AgNO3 and sodium citrate and the minimum inhibitory concentration (MIC) of Ag-NPs and Ag+ ions on the mastitis pathogens were determined. Isolates were exposed to subinhibitory concentrations of Ag-NPs or AgNO3 for 10 consecutive days to verify the development of adaptive resistance evaluated by changes in the MIC values. The development of cross-resistance with antibiotics was also studied, being verified by comparing the sensitivity profile of treated cells with non-treated cells. AgNO3 was more effective against all isolates. There was no change in the MIC values or in the antibiotic sensitivity profile for both bacteria following consecutive exposure to subinhibitory dosages of Ag-NPs or AgNO3, indicating that silver was not able to select adaptive resistance or cross resistance to the tested antibiotics. The potential of silver presented by these results is favorable to the continuity of studies aiming to elaborate silver-based therapies for the treatment of bovine mastitis.

ABSTRACT The objective of this study was to evaluate the quality of water purification system and identify the bacteria this system, predict bacterial adherence according to the hydrophobicity of these microorganisms and of the polypropylene distribution loop for purified water. The assessment of drinking water that supplies the purification system allowed good-quality physical, chemical, and microbiological specifications. The physicochemical specifications of the distributed purified water were approved, but the heterotrophic bacteria count was higher than allowed (>2 log CFU mL-1).The sanitation of the storage tank with chlorine decreased the number of bacteria adhered to the surface (4.34 cycles log). By sequencing of the 16SrDNA genes, six species of bacteria were identified. The contact angle was determined and polypropylene surface and all bacteria were considered to be hydrophilic, and adhesion was thermodynamically unfavorable. This case study showed the importance of monitoring the water quality in the purified water systems and the importance of sanitization with chemical agents. The count of heterotrophic bacteria on the polypropylene surface was consistent with the predicted thermodynamics results because the number of adhered cells reached approximate values of 5 log CFU cm-2.

The artisanal Minas cheese is produced from raw cow's milk and wooden utensils were employed in its manufacture, which were replaced by other materials at the request of local laws. This substitution caused changes in the traditional characteristics of cheese. Due to the absence of scientific studies indicating the microbial composition of biofilms formed on wooden forms, tables and shelves used in these cheese production, the present work evaluated the counts of Staphylococcus aureus, Escherichia coli, coliforms at 32 °C, yeasts, presumptive mesophilic Lactobacillus spp. and Lactococcus spp. in these biofilms, milk, whey endogenous culture and ripened cheese in two traditional regions: Serro and Serra da Canastra. Also, we checked for the presence of Salmonella sp. and Listeria monocytogenes in the ripened cheeses. The ultra structure of the biofilms was also assessed. Counts above legislation (> 2 log cfu/mL) for the pathogens evaluated were found in milk samples from both regions. Only one shelf and one form from Serro were above limits proposed (5 cfu/cm² for S. aureus and E. coli and 25 cfu/cm² for coliforms) in this study for contaminants evaluated. In Canastra, few utensils presented safe counting of pathogens. There was no Salmonella sp. and Listeria monocytogenes in the cheeses after ripening. Thus, the quality of the cheese is related to improving the microbiological quality of milk, implementation and maintenance of good manufacturing practices, correct cleaning of wooden utensils, and not its replacement.

The adhesion of the solids presents in food can difficult the process of surface cleaning and promotes the bacterial adhesion process and can trigger health problems. In our study, we used UHT whole milk, chocolate based milk and infant formula to evaluate the adhesion of Enterobacter sakazakii on stainless steel coupons, and we determine the work of adhesion by measuring the contact angle as well as measured the interfacial tension of the samples. Inaddition we evaluated the hydrophobicity of stainless steel after pre-conditioning with milk samples mentioned. E. sakazakii was able to adhere to stainless steel in large numbers in the presence of dairy products. The chocolate based milk obtained the lower contact angle with stainless steel surface, higher interfacial tension and consequently higher adhesion work. It was verified a tendency of decreasing the interfacial tension as a function of the increasing of protein content. The pre-conditioning of the stainless steel coupons with milk samples changed the hydrophobic characteristics of the surfaces and became them hydrophilic. Therefore, variations in the composition of the milk products affect parameters important that can influence the procedure of hygiene in surface used in food industry.

The interaction between the surface of stainless steel and Bacillus cereus was studied in terms of the characteristics of interfacial interaction determined from the measurement of the contact angle of the surface of B. cereus and stainless steel in the presence or absence of B. cereus adherence. The microtopographies and the roughness of the surface of stainless steel and stainless steel adhered by B. cereus were evaluated with the help of atomic force microscopy and perfilometry. The strain of B. cereus studied was considered hydrophilic, whereas the stainless steel was considered hydrophobic. The adhesion was not thermodynamically favorable (ΔGadhesion > 0) between the stainless steel and the strain of B. cereus studied. Thus, the interaction between them was not favored by the thermodynamic aspect of adhesion. There was no difference (p > 0.05) in the roughness of the surfaces of stainless steel adhered by B. cereus when analyzed by atomic force microscope and perfilometry.

The ability of bacteria to attach to surface and develop into a biofilm has been of considerable interest to food industry. Electrostatic, Lifhistz-van der Waals and Lewis acid-base forces are usually considered responsible for the interactions at the interface of the bacterial adhesion. The study of microbial adhesion thermodynamic is important because it represents the reflection of microbial surface and food processing surface physicochemical characteristics. This review examines the most important aspects involved in bacterial attachment to a surface with emphases in thermodynamics of adhesion process.

There is an increasing tendency to add natural antimicrobials of plant origin into food. The objective of this work was to develop a microbial sachet incorporated with allyl isothiocyanate (AIT), a volatile compound of plant origin, and to test its efficiency against growth of yeasts and molds, Staphylococcus sp. and psychrotrophic bacteria on sliced mozzarella cheese. Another objective was to quantify the concentration of AIT in the headspace of cheese packaging. A reduction of 3.6 log cycles was observed in yeasts and molds counts in the mozzarella packed with the antimicrobial sachet over 15-day storage time. The sachet also showed an antibacterial effect on Staphylococcus sp., reducing 2.4 log cycles after 12-day storage. Psychrotrophic bacteria species were the most resistant to the antimicrobial action. The highest concentration of AIT (0.08µg.mL-1) inside the active packaging system was observed at the 6-day of storage at 12 ºC ± 2 ºC. At the end of the storage time, AIT concentration decreased to only 10% of the initial concentration. Active packaging containing antimicrobial sachet has a potential use for sliced mozzarella, with molds and yeasts being the most sensitive to the antimicrobial effects.

A adesão de substâncias orgânicas e de microrganismos em cupons de aço inoxidável ASI 304, #4 foi avaliada. A medida de Unidades Relativas de Luz (RLU) foi afetada em menor ou maior nível pelo tipo e concentração das substâncias orgânicas e suas combinações. Entretanto, se as amostras fossem oriundas de superfícies de processamento de alimentos, elas seriam consideradas em boas condições higiênicas pois as medidas de log URL ficaram abaixo de 2,18 log (< 150 URL) com ou sem adição das substâncias orgânicas e independente da concentração estudada, as células vegetativas apresentaram número de log URL para S. carnosus maior (p<0,05) do que para os esporos de B. subtilis. Amostras de superfícies aderidas com caseína, lipídio e sacarose e 2.9x10¹ CDM.cm-2 ou 2.9x10² CDM.ml-1 não apresentaram diferença significativa (p>0.05) na medida de ATP-bioluminescência. Entretanto, superfícies aderidas com 2.9x10³ CDM.cm-2 de esporos de B. subtilis e as substâncias orgânicas mostraram um aumento no log URL comparado com as superfícies contendo somente substâncias orgânicas.

Adhesion of organic substances and microorganisms on stainless steel surface was evaluated. The log RLU measurement was affected at a lower or higher degree, by type or concentration of the organic substances (casein, lipid, sucrose and their combinations) adhered to surface. However, if the samples analyzed were from the food processing surfaces, they would have been considered to be under satisfactory hygienic conditions with log RLU < 2.18 (< 150 RLU). The log RLU measurements of Staphylococcus carnosus-adhered surfaces, combined with or without the organic substances, were higher (p<0.05) than those of the Bacillus subtilis spores-adhered surface, regardless of the concentration. The samples taken from the surfaces adhered with the casein, lipid, sucrose and 2.9x10¹ DMC.cm-2 and 2.9x10² DMC.ml-1 did not have any significant differences (p>0.05) on ATP-bioluminescence measurements. However, the surfaces adhered with 2.9x10³ DMC.cm-2 of B. subtilis spores and organic substances showed an increase in the log RLU measurement compared to surfaces containing only the organic substances.

O objetivo principal deste trabalho foi avaliar a eficiência de sachês absorvedores de oxigênio a 75%, 80% e 85% de umidade relativa e diferentes temperaturas, 10±2 ºC e 25±2 ºC. O experimento consiste em determinar a absorção de O2 sob essas condições. Um sachê foi colocado dentro de um dessecador contendo um sistema de homogeneização do ar interno. Alíquotas de ar são retiradas dos dessecadores em intervalos de tempos pré-estabelecido e seu conteúdo de oxigênio analisado. Os resultados mostraram que a absorção de oxigênio pelos saches aumentaram com o aumento da temperatura para ambas as temperaturas. No entanto, os sachês mostraram uma maior eficiência para 85% de umidade relativa e 25±2ºC de temperatura. Na condição ambiente (25±2ºC/75%RH), a taxa de absorção dos sachês foi de 50 mL/dia e 18,5 mL/dia para 10±2ºC. O experimento foi conduzido com delineamento experimental inteiramente casualizado, com três repetições.

The main objective of this work was to evaluate the efficiency of oxygen - absorbing sachets at relative humidity of 75%, 80% and 85% and different temperatures, 10±2 ºC and 25±2 ºC. The experiment consisted in determining the O2 absorption under these conditions. A sachet was placed in desiccators with an internal air homogenization system. Aliquots of air were removed at pre-established time intervals and analyzed for oxygen content. The results showed that oxygen absorption by the sachet increased as the relative humidity increased for both temperature. Therefore the oxygen - absorbing sachets were most active under 25±2ºC and 85% relative humidity. At ambient condition (25±2ºC/75%RH) the rate of oxygen absorbed was 50 mL/day and 18,5 mL/day for 10±2ºC. It was used a totally casualized design with three replicates.

O objetivo deste trabalho foi estabelecer uma proposta para o processamento do queijo Coalho regional seguindo-se os procedimentos adequados para a obtenção de um produto final de qualidade. Empregou-se leite de vaca padronizado e pasteurizado e culturas láticas endógenas. Foram isoladas, identificadas e caracterizadas culturas láticas endógenas de leite de vaca cru e de queijos Coalho artesanais. Algumas culturas foram avaliadas para a fabricação do queijo Coalho, seguindo-se o fluxograma de fabricação estabelecido neste estudo. O produto final foi submetido a análises físico-químicas, teste de aceitação e teste de fritura. Os resultados obtidos mostraram que é viável a padronização do produto tradicional, proporcionando redução de custos, qualidade microbiológica e a manutenção das características sensoriais do queijo Coalho regional, com o uso de culturas láticas nacionais.

The main objective of this work is to propose a manufacturing procedure to make a safe artisan Coalho type cheese. Pasteurized and standardized cow´s milk added with endogenous lactic acid culture was used in the procedure. Endogenous lactic cultures were identified and characterized after being isolated from raw cow´s milk and artisan cheeses. The selected cultures were used in the manufacturing procedure designed for Coalho type cheese. Cheese made with added endogenous lactic acid culture underwent physic-chemical analysis, sensory acceptance and fritter tests. Results showed that it is feasible to make such cheese in a standardized process, with improved microbiological quality, keeping sensory properties of traditional cheese made with raw milk and a reduced cost using suppression on imports of starter cultures.

Foi avaliado o sistema de absorvedor de oxigênio para inibição de microrganismos em massa fresca de lasagna durante estocagem a 10±2ºC. A massa de lasagna foi produzida com e sem sorbato de potássio e acondicionada em embalagens com alta barreira a oxigênio contendo um sachê absorvedor em seu interior. Foram avaliados três tratamentos: massa com sorbato de potássio (1), massa sem sorbato de potássio e com sachê absorvedor de oxigênio (2) e massa sem sorbato de potássio e sem sachê absorvedor de oxigênio (3). Em todos os tratamentos, a massa foi envasada a vácuo e distribuída aleatoriamente sob temperatura de 10±2ºC e avaliada microbiologicamente quanto o crescimento de bolores e leveduras, Staphylococcus spp, coliformes totais e Escherichia coli. Todos os tratamentos foram efetivos na inibição do crescimento dos microrganismos avaliados durante o período de 30 dias de armazenamento. O tratamento 2 inibiu o crescimento de bolores e leveduras e Staphylococcus spp, em 1 e 1,5 ciclos logarítmicos, respectivamente. Não houve diferença significativa para coliformes em todos os tratamentos.

O2 absorbent system was evaluated on the inhibition of microorganisms growth in fresh lasagna pasta during storage at 10±2ºC. Fresh lasagna pasta was produced with and without potassium sorbate and acondicionated in high O2 barrier bags containing a O2 - absorber sachet in the head-space. Three treatments were obtained: pasta with potassium sorbate, pasta without potassium sorbate packed with sachet and pasta without potassium sorbate packed without sachet. In all treatments, the pasta were vacuum packed, randomly distributed at temperature of 10±2ºC and microbiologically evaluated for molds and yeast, Staphylococcus spp, total coliforms and Escherichia coli countings. All the treatments were effective in inhibiting the growth of the microorganisms evaluated during 30 storage days. The treatment using O2-absorber in the pasta without preservative, inhibition of 1 and 1.5 logarithmic cycles was observed for the molds and yeasts and Staphylococcus spp, respectively. No differences were observed among treatments for coliforms countings.

A técnica do ATP-bioluminescência avaliou os procedimentos de higienização de superfícies em aço inoxidável dos tanques de transporte de leite cru, de estocagem de leite cru resfriado, de equilíbrio de um pasteurizador, de estocagem de leite pasteurizado e para embalagem de leite pasteurizado e, ainda, a superfície interna de uma centrífuga de leite quanto à determinação de Unidades Relativas de Luz (URL) e da contagem de mesófilos aeróbios (UFC.cm-2). Não houve diferença significativa (p>0.05) na medida de URL nas superfícies antes e depois do procedimento de higienização. Também, não houve diferença significativa (p>0.05) no número de UFC.cm-2 nas superfícies antes da higienização. No entanto, contagens bacterianas diferentes foram constatadas após este procedimento. As superfícies da centrífuga e dos tanques de leite resfriado e para embalagem de leite apresentaram as maiores contagens após a higienização, atingindo 2,21, 2,18, e 1,21 log10, respectivamente. Os resultados mostram que não há concordância entre os métodos na classificação das condições higiênicas das superfícies, indicando que não há relação direta entre RLU e UFC.cm-2. A técnica do ATP-bioluminescência indicou que 100% das superfícies estavam em condições higiênicas inadequadas, enquanto a contagem bacteriana indicou apenas 50% de acordo com as recomendações da APHA e 33% de acordo com OMS. Variações elevadas nas medidas de URL sugerem a necessidade de se realizar mais de uma análise na mesma superfície. A técnica do ATP-bioluminescência não deve simplesmente substituir os testes microbiológicos mas ser usada para se obter informações complementares para avaliar as condições higiênicas das superfícies para o processamento de leite.

ATP-bioluminescence assay was used to evaluate the cleaning and sanitizing procedures of stainless steel milk contact surfaces. Surfaces of a raw milk transportation tank, a raw milk cooling storage tank, an equalization pasteurizing tank, a pasteurized milk storage tank and a pasteurized milk packaging tank, as well as a milk centrifuge were evaluated. The Relative Light Unit (RLU) using a commercial luminometer and mesophilic aerobic counts (CFU.cm-2) on surfaces were determined. There were no significant differences (p>0.05) in RLU on the assessed surfaces before and after the cleaning and sanitizing procedures. Also, there were no significant differences (p>0.05) in CFU.cm-2 on the surfaces before the cleaning and sanitizing procedures, whereas different bacterial numbers were found on surfaces after the hygiene procedures. The surfaces of the milk centrifuge, raw milk cooling tank and milk packaging tank showed the largest mesophilic aerobic numbers after the hygiene, reaching 2.21, 2.18, and 1.21 log10, respectively. The results demonstrated that there is no agreement among the techniques for classification of hygienic conditions of the evaluated surfaces, showing a non-direct relation between RLU and CFU.cm-2. The ATP-bioluminescence method showed that 100% of the surfaces were under inadequate hygiene conditions, while the plate count method detected only 50%, based on the APHA's recommendation, and 33%, based on the WHO's recommendation. High variations in RLU measurements were observed, suggesting the need for more than only one surface analysis. The ATP-bioluminescence is not a direct replacement for microbiological testing, but provides complementary information and should be used as part of an integrated strategy.

É objetivo deste estudo desenvolver um sistema multimídia com a finalidade de subsidiar a melhoria de procedimentos de higiene em Unidades de Alimentação e Nutrição. O sistema Clean Up, composto de várias bases de conhecimento e informações interligadas e organizadas em módulos foi avaliado por 30 usuários que responderam ao teste de aceitação baseado na Escala Likert modificada, com os atributos próprios de escala hedônica, com notas de 1 a 9, para classificação que varia de "extremamente bom" a "extremamente ruim". A partir dos dados coletados foram calculadas a moda e os valores máximo e mínimo das notas para cada atributo julgado pelos avaliadores. A nota 8 foi a mais freqüente, classificando os módulos como "muito bom", segundo a escala Likert. É também função do multimídia entreter o usuário. As cores, sons, filmes, imagens devem surpreender e criar expectativas positivas, aumentando a vontade de explorar o conteúdo durante a aquisição do conhecimento.

The purpose of this study was to develop a multimedia system to help improve hygiene practices in food facilities.The Clean Up system, composed of many knowledge bases and interconnected information organized in modules, was tested by 30 users who answered an acceptability test based on the modified Likert Scale, with typical hedonic scale attributes and grades ranging from 1 to 9, in a classification that varied from "extremely good" to "extremely bad." The mode and the maximum and minimum values of the grades given to each attribute judged by the users were calculated from the collected data. The most frequent grade given in the quantitative assessment of the modules was 8, that is, "very good" according to the Likert Scale. The multimedia should also entertain the user. Colors, sounds, films and images must surprise the user and create positive expectations, increasing the user's will to explore its contents during learning.

O manjericão (Ocimum basilicum L.) é empregado tanto na indústria culinária quanto fitoterápica e na medicina tradicional, devido ao teor e composição de seu óleo essencial. No entanto, o manjericão não pode ser conservado por longo período após a colheita e sua qualidade pode ser prejudicada. O objetivo deste trabalho foi verificar a influência da época e do horário de colheita sobre a conservação de manjericão armazenado por diferentes períodos. O manjericão foi colhido às 8h e às 16h em agosto/1999 e em janeiro/2000. Os ramos foram acondicionados em filmes de PVC e armazenados por 3, 6 e 9 dias. Durante o armazenamento, o teor de clorofila, e o teor e a composição do óleo essencial foram determinados e foram conduzidas análises microbiológicas. A época de colheita e os dias de armazenamento influenciaram o teor final de óleo essencial. Houve decréscimo linear dos teores de óleo essencial e de clorofila e do número de colônias de fungos e bactérias ao longo do armazenamento. Não houve efeito da época de colheita ou do horário de colheita sobre a composição do óleo essencial, mas os teores de eugenol e linalol aumentaram durante o armazenamento. O número de coliformes ficou abaixo de 0,3 NMP g-1 e o número de Staphylococcus aureus, abaixo de 1,0x10² UFC g-1.

Fresh basil (Ocimum basilicum L.) is used in food, phytotherapic industry, and in traditional therapeutic, due to its essential oil content and composition. Nevertheless basil can not be kept for long periods after harvest and its quality can be reduced. This work aimed to assess the influence of the season and harvest time in the postharvest conservation of basil stored for different periods. Basil was harvested at 8 am and 4 pm both in August/1999 and January/2000. Cuttings were conditioned in PVC packages and stored for 3, 6, and 9 days. During storage, chlorophyll content, essential oil content and composition were determined as well as microbiological analyses were carried out. Harvest season and the days of storage influenced the final content of essential oil. There was a linear decrease in the content of essential oil, in the chlorophyll content and in the number of mold and yeast colonies during storage. There was no effect of cropping season or harvest hour on essential oil composition, but the eugenol and linalool content increased during storage. Coliforms were under 0.3 MPN g-1 and the number of Staphylococcus aureus was under 1.0x10² UFC g-1.

Evidências que o leite produzido e consumido no Brasil nem sempre apresenta a qualidade desejada têm gerado a discussão e desenvolvimento de novas políticas de incentivo à produção leiteira, resultando no desenvolvimento do Programa Nacional de Melhoria da Qualidade do Leite. Em complementação, em 2002 o Ministério da Agricultura publicou a Instrução Normativa 51 (IN51), com importantes inovações em relação à conservação e transporte do leite cru, além de estabelecimento de um padrão de qualidade para esse tipo de leite (10(6) UFC/mL), a ser implantado em diferentes prazos nas diferentes regiões do país, a partir de 2005. O presente trabalho teve como objetivo verificar se o leite cru produzido em quatro áreas de quatro estados produtores de leite no Brasil estaria, nesse momento, em condições de cumprir o estabelecido na IN 51, especialmente quanto ao atendimento dos padrões microbiológicos previstos. Amostras de leite cru, coletadas em 210 diferentes propriedades nas regiões de Viçosa, MG (47), Pelotas, RS (50), Londrina, PR (63) e Botucatu, SP (50), foram analisadas quanto aos níveis de contaminação por aeróbios mesófilos, utilizando o PetrifilmTM AC. Parcela significativa das amostras (48,6%) apresentaram contagens acima do determinado pela IN51, sendo 21,3% na região de Viçosa (MG), 56,0% na região de Pelotas (RS), 47,6% na região de Londrina (PR) e 68,0% na região de Botucatu (SP). Considerando as diferenças de cada região, foi possível observar a importância da refrigeração na conservação e transporte da produção, bem como da implantação de boas práticas e assistência técnica nas propriedades. Os resultados obtidos permitem concluir que a adequação às normas estabelecidas pela IN51 pode ser mais difícil em algumas regiões do que em outras, sendo fundamental a adoção da refrigeração na conservação e no transporte da produção, e de programas regionais de assistência a produtores leiteiros.

The need to improve quality and safety of milk produced and consumed in Brazil resulted in development of new governmental policies to support the Brazilian milk-producing sector. Among them, the most relevant is the PNMQL (National Program to Improve Milk Quality). In 2002, the Brazilian Ministry of Agriculture created the Instrução Normativa 51 (Normative Instruction 51, IN51), with important innovations related to maintenance and transport of raw milk, besides establishment of microbiological standards to be implemented in different periods of time, according to the producing region, starting in 2005. The present study aimed to trace if raw milk produced in four important milk-producing regions in the country would be capable to be in accordance to the microbiological standards established by IN51. Raw milk samples, collected in 210 different milk-producing farms, located in vicinities of Viçosa (Minas Gerais state), Pelotas (Rio Grande do Sul state), Londrina (Paraná state) and Botucatu (Sao Paulo state) were submitted to counts of mesophilic aerobes using PetrifilmTM AC plates. In a significant number of samples, the counts were above the standards, being 21.3% from Viçosa, 56.0% from Pelotas, 47.6% from Londrina and 68.0% from Botucatu. Taking into account the differences in milk-producing practices adopted in the four regions, the study evidenced the importance of cooling during maintenance and transport of raw milk. The relevance of adoption of good hygiene practices and access to technical support was also evidenced. The conclusion of the study is that the difficulties to achieve the standards established by IN51 may vary from region to region. Cooling of milk for maintenance and transport is fundamental, but regional programs to assist the milk producers are also needed.