Fifty-eight meat and poultry plants in Virginia were assessed during spring and summer of 2000. These assessments were all conducted in the presence of state inspection and were designed to be non-regulatory. The audit team included N.G. Marriott, M.A. Tolbert and B.P. Quinn. The audits consisted of a tour of the facility and a review of SSOPs and all HACCP related documentation. To assist in these audits, a HACCP check sheet was developed and utilized to indicate suggestions or deficiencies. Most of the plants had an understanding of how to implement HACCP properly. The majority of the suggestions that were noted were not so much about the HACCP concept, but more with regards to the legality of a HACCP document. The most noted deficiency was improper cross-outs. If there is a correction, one line should be drawn through the error and then must be initialed. With respect to the HACCP plan, most deficiencies were related to the hazards and the critical control points. During these audits, two microbial determination methods (Standard Plate Count and Bioluminescence) were used to evaluate processing equipment. Typically, three pieces of equipment were tested at each plant. When the data were collected, the two microbial determination methods were correlated. The "corr" function in SAS resulted in a correlation coefficient of .4478, which is low and indicates a poor correlation. A pass/fail method similar to one done by Illsley et. al. resulted in a 48.9% agreement between the methods in this research. / Master of Science

Heterotrophic plate counts (HPCs) are commonly used to assess the general microbiological quality of drinking water. Drinking water quality specifications world-wide recommend HPC limits from 100 to 500 cfu.m1-1. However, a number of recent studies revealed evidence that commonly used indicator bacteria may not be as harmless as generally accepted. It appears that immuno-compromised individuals, which represent increasing components of many consumer populations, are particularly at risk. This would include the very young and very old, patients with diseases such as AIDS, and patients on therapy after organ transplantations and cancer treatment. Since, epidemiological and animal infectivity studies are complex and difficult to control, attempts have been made by researchers to examine HPCs directly in order to assess health risks. These analyses included: cytotoxicity, invasiveness, enzyme analyses, antibiotic susceptibility and identification. In this study, 339 bacterial colonies were isolated at random from selected drinking water supplies in South Africa using heterotrophic plate count tests. In a first step to screen for potentially pathogenic properties, 188 (55.5%) of the isolates showed α- and β-haemolysis on human- and horse-blood agar media. Subsequent analysis of the haemolytic isolates for enzymatic properties associated with pathogenicity revealed the presence of chondroitinase in 5.3% of the isolates, coagulase in 16.0%, DNase in 60.6%, elastase in 33 .0%, fibrinolysin in 53.7%, gelatinase in 62.2%, hyaluronidase in 21. 3 %, lecithinase in 47.9%, lipase in 54.8%, and proteinase in 64.4%. Fluorescein and pyocyanin were not produced by any of the isolates. The Kirby-Bauer quality controlled disc diffusion method was applied in the demonstration of antibiotic resistance by the HPC isolates. Among the haemolytic isolates 77.7% were resistant to oxacillin (1 µg), 59.6% to penicillin G (2 units), 47.3% to penicillin G (10 units), 54.3% to ampicillin (10 µg) and 43.1% to ampicillin (25 µg). Cell culture studies revealed that 96% of haemolytic isolates were cytotoxic to HEp-2 cells and 98.9% of the 181 cytotoxic isolates adhered to HEp-2 or Caco-2 cells. Gram-negative isolates tended to adhere in larger numbers than gram-positive isolates. The average index of adherence for Gram-negative bacteria was 20-30 bacteria per HEp-2 cell, compared to 3-7 for Gram-positive bacteria. HEp-2 cells were invaded by 43.6% and Caco-2 cells by 49.7% of the 181 cytotoxic isolates. The invasion index on HEp-2 cells was 1.9xlO-1 to 8.9xl0-6, compared to 7.7xl0-2 to 8.3xlO-6 on Caco-2 cells. The most commonly isolated genera showing potentially pathogenic features were: Aeromonas, Acinetobacter, Aureobacterium, Bacillus, Chryseobacterium, Corynebacterium, Klebsiella, Moraxella, Pseudomonas, Staphylococcus, Tsukamurella and Vibrio. All these genera are known to contain opportunistic pathogens. Our results support earlier findings on potentially pathogenic features of bacteria detected by heterotrophic plate counts on drinking water. These findings seem to be in agreement with some epidemiological studies, which indicated an association between HPCs of drinking water and the incidence of gastroenteritis in consumers. However, the extent of the health risk concerned needs to be defined in detail for meaningful revision of quality guidelines for HPCs in drinking water. / Dissertation (MSc (Medical Virology))--University of Pretoria, 2005. / Medical Virology / unrestricted

Foram avaliados 25 reservatórios de água dos equipos odontológicos da Clínica de Dentística/Endodontia da FOB/USP com relação à presença de micro-organismos e a ação do ultrassom (US) na remoção do biofilme. Amostras de 10ml de água foram obtidas e alíquotas de 25l in natura e diluída até 10-4 foram semeadas pela técnica da gota nos meios: R2A Agar (R2A), Plate Count Agar (PCA), Peptona Diluída (PD) e Sabouraud Dextrose Agar com cloranfenicol a 1% (SDA), incubadas a 24º C por 72 horas. A água dos reservatórios foi descartada e 500 ml de água destilada esterilizada foi adicionada, sendo submetidos à ação do ultrassom (US) por 15 minutos, seguidos do mesmo procedimento descrito anteriormente. As colônias de bactérias foram quantificadas e os fungos foram identificados por micro-cultivo. A média da detecção de UFC/ml antes e após o US foi de 173.787 e 15.841 para o R2A, 104.873 e 3.034 para o PCA e de 245.824 e 8.231 para o PD. A média de fungos foi de 52,4 antes e 19,2 UFC/ml após ação do US. Fungos foram detectados em 20 reservatórios antes e em 12 deles após uso do US. O Penicillium sp apresentou prevalência de 36% nos reservatórios de água avaliados. Os resultados obtidos permitem concluir que o US foi eficiente em desestruturar o biofilme, embora não o elimine por completo, apresentando maior efetividade na desestruturação de bactérias. / A total of 25 waterline unit reservoirs of the odontological sets from the Dentistry/Endodontic Clinic of FOB/USP were assessed, in relation to the presence of microorganisms and the ultrasound action (US) on the biofilm removal. Waterline samples of 10ml were obtained from aliquots of 25l in natura and diluted until 10-4, then, they were spread using the dripping technique on the means: R2A Agar (R2A), Plate Count Agar (PCA), diluted Peptone (DP) and Sabouraud Dextrose Agar with cloranfenicol at 1% (SDA), being incubated at 24º C for 72 hours. The waterline units of the reservoirs were discarded and 500 ml of sterilized distilled water was added, submitted to ultrasound action (US) for 15 minutes, following the same procedure described afore. The bacteria colonies were quantified and the fungi were identified through micro-culture. The average of detection of UFC/ml before and after US was 173.787 and 15.841 for R2A, 104.873 and 3.034 for PCA and of 245.824 and 8.231 for PD. The fungi average was 52,4 before and 19,2 UFC/ml after the action of US. Fungi were detected in 20 reservoirs before and 12 after using US. Penicillium sp showed a prevalence of 36% in the waterline reservoirs assessed. The results obtained, led to the conclusion that US was efficient to break the structure of the biofilm, although it did not eliminate it completely, showing more effectiveness to break the bacteria structure.

Foram avaliados 25 reservatórios de água dos equipos odontológicos da Clínica de Dentística/Endodontia da FOB/USP com relação à presença de micro-organismos e a ação do ultrassom (US) na remoção do biofilme. Amostras de 10ml de água foram obtidas e alíquotas de 25l in natura e diluída até 10-4 foram semeadas pela técnica da gota nos meios: R2A Agar (R2A), Plate Count Agar (PCA), Peptona Diluída (PD) e Sabouraud Dextrose Agar com cloranfenicol a 1% (SDA), incubadas a 24º C por 72 horas. A água dos reservatórios foi descartada e 500 ml de água destilada esterilizada foi adicionada, sendo submetidos à ação do ultrassom (US) por 15 minutos, seguidos do mesmo procedimento descrito anteriormente. As colônias de bactérias foram quantificadas e os fungos foram identificados por micro-cultivo. A média da detecção de UFC/ml antes e após o US foi de 173.787 e 15.841 para o R2A, 104.873 e 3.034 para o PCA e de 245.824 e 8.231 para o PD. A média de fungos foi de 52,4 antes e 19,2 UFC/ml após ação do US. Fungos foram detectados em 20 reservatórios antes e em 12 deles após uso do US. O Penicillium sp apresentou prevalência de 36% nos reservatórios de água avaliados. Os resultados obtidos permitem concluir que o US foi eficiente em desestruturar o biofilme, embora não o elimine por completo, apresentando maior efetividade na desestruturação de bactérias. / A total of 25 waterline unit reservoirs of the odontological sets from the Dentistry/Endodontic Clinic of FOB/USP were assessed, in relation to the presence of microorganisms and the ultrasound action (US) on the biofilm removal. Waterline samples of 10ml were obtained from aliquots of 25l in natura and diluted until 10-4, then, they were spread using the dripping technique on the means: R2A Agar (R2A), Plate Count Agar (PCA), diluted Peptone (DP) and Sabouraud Dextrose Agar with cloranfenicol at 1% (SDA), being incubated at 24º C for 72 hours. The waterline units of the reservoirs were discarded and 500 ml of sterilized distilled water was added, submitted to ultrasound action (US) for 15 minutes, following the same procedure described afore. The bacteria colonies were quantified and the fungi were identified through micro-culture. The average of detection of UFC/ml before and after US was 173.787 and 15.841 for R2A, 104.873 and 3.034 for PCA and of 245.824 and 8.231 for PD. The fungi average was 52,4 before and 19,2 UFC/ml after the action of US. Fungi were detected in 20 reservoirs before and 12 after using US. Penicillium sp showed a prevalence of 36% in the waterline reservoirs assessed. The results obtained, led to the conclusion that US was efficient to break the structure of the biofilm, although it did not eliminate it completely, showing more effectiveness to break the bacteria structure.

The presence of heterotrophic plate count (HPC) bacteria in drinking water distribution systems is usually not considered harmful to the general consumer. However, precautions must be taken regarding the immunocompromised. All water supply authorities in South Africa are lawfully required to provide consumers with high-quality drinking water that complies with South African-and international standards. This study mainly focused on the isolation, identification and characterization of HPC and other bacteria from biofilm-and bulk water samples from two sampling points located within the Potchefstroom drinking water distribution system. Based on five main objectives set out in this study, results indicated that the bulk water at the J.S. van der Merwe building was of ideal quality fit for lifetime consumption. Application of enrichment-and selective media allowed for the isolation of 12 different bacterial morphotypes. These were identified by way of biochemical-and molecular methods as Bacillus cereus, Bacillus subtilis, Brevundimonas spp., Clostridiaceae, Corynebacterium renale, Flavobacteriaceae, Kytococcus sedentarius, Leuconostoc lactic, Lysinibacillus sphaericus, Pseudomonas spp., Staphylococcus aureus and Staphylococcus capitis. The greatest diversity of bacteria was detected early autumn 2008, while the lowest diversity occurred during mid-winter 2007. Bacillus cereus, Kytococcus sedentarius and Staphylococcus capitis displayed potential pathogenic properties on blood agar. Kytococcus sedentarius could be classified as potentially the most pathogenic among the isolates. All isolates displayed multiple-resistant patterns towards tested antibiotics. Corynebacterium renale and Staphylococcus aureus were least resistant bacterial species and Lysinibacillus sphaericus the most resistant. All isolates were susceptible to ciprofloxacin (CIP) and streptomycin (S), but most were resistant to erythromycin (E). Transmission electron microscopy (TEM) allowed for detailed examination of Brevundimonas spp., Pseudomonas spp. and Staphylococcus spp. The capability of Brevundimonas spp. to produce slime and store nutrients within inclusion bodies, suggests the ability of this bacterium to form biofilm and persist in the drinking water for prolonged periods. Despite the inhibitory or toxic effect of copper against bacterial growth, scanning electron microscopy (SEM) revealed the presence of biofilms as well as diatoms on red-copper coupons. Biofilm activity was also observed on reverse-osmosis (RO) filters. Since corrosion was evident on red-copper coupons, it is recommended that prospective studies also look into the significance of microbial induced corrosion (MIC) within the Potchefstroom drinking water distribution system. Other prospects include determining minimum inhibitory concentrations of isolates against antibiotics and the application of culture independent methods such as SSCP and DGGE to investigate biofilm development. The use of diatoms as an index of the drinking water quality is also suggested. / Thesis (M.Sc. (Environmental Science))--North-West University, Potchefstroom Campus, 2010.

The presence of heterotrophic plate count (HPC) bacteria in drinking water distribution systems is usually not considered harmful to the general consumer. However, precautions must be taken regarding the immunocompromised. All water supply authorities in South Africa are lawfully required to provide consumers with high-quality drinking water that complies with South African-and international standards. This study mainly focused on the isolation, identification and characterization of HPC and other bacteria from biofilm-and bulk water samples from two sampling points located within the Potchefstroom drinking water distribution system. Based on five main objectives set out in this study, results indicated that the bulk water at the J.S. van der Merwe building was of ideal quality fit for lifetime consumption. Application of enrichment-and selective media allowed for the isolation of 12 different bacterial morphotypes. These were identified by way of biochemical-and molecular methods as Bacillus cereus, Bacillus subtilis, Brevundimonas spp., Clostridiaceae, Corynebacterium renale, Flavobacteriaceae, Kytococcus sedentarius, Leuconostoc lactic, Lysinibacillus sphaericus, Pseudomonas spp., Staphylococcus aureus and Staphylococcus capitis. The greatest diversity of bacteria was detected early autumn 2008, while the lowest diversity occurred during mid-winter 2007. Bacillus cereus, Kytococcus sedentarius and Staphylococcus capitis displayed potential pathogenic properties on blood agar. Kytococcus sedentarius could be classified as potentially the most pathogenic among the isolates. All isolates displayed multiple-resistant patterns towards tested antibiotics. Corynebacterium renale and Staphylococcus aureus were least resistant bacterial species and Lysinibacillus sphaericus the most resistant. All isolates were susceptible to ciprofloxacin (CIP) and streptomycin (S), but most were resistant to erythromycin (E). Transmission electron microscopy (TEM) allowed for detailed examination of Brevundimonas spp., Pseudomonas spp. and Staphylococcus spp. The capability of Brevundimonas spp. to produce slime and store nutrients within inclusion bodies, suggests the ability of this bacterium to form biofilm and persist in the drinking water for prolonged periods. Despite the inhibitory or toxic effect of copper against bacterial growth, scanning electron microscopy (SEM) revealed the presence of biofilms as well as diatoms on red-copper coupons. Biofilm activity was also observed on reverse-osmosis (RO) filters. Since corrosion was evident on red-copper coupons, it is recommended that prospective studies also look into the significance of microbial induced corrosion (MIC) within the Potchefstroom drinking water distribution system. Other prospects include determining minimum inhibitory concentrations of isolates against antibiotics and the application of culture independent methods such as SSCP and DGGE to investigate biofilm development. The use of diatoms as an index of the drinking water quality is also suggested. / Thesis (M.Sc. (Environmental Science))--North-West University, Potchefstroom Campus, 2010.

In this study, the bacterial growth dynamics of 14 pilot drinking water distribution systems were studied in order to observe water quality changes due to corrosion inhibitor addition. Empirical models were developed to quantity the effect of inhibitor type and dose on bacterial growth (biofilm and bulk water). Water and pipe coupon samples were taken and examined during the experiments. The coupons were exposed to drinking water at approximately 20°C for at least 5 weeks to allow the formation of a measurable quasi- steady-state biofilm. Bulk water samples were taken every week. In this study, two simple but practical empirical models were created. Sensitivity analysis for the bulk HPC model (for all 14 of the PDSs) showed that maintaining a chloramine residual at 2.6 mg/L instead of 1.1 mg/L would decrease bulk HPC by anywhere from 0.5 to 0.9 log, which was greater than the increase in bulk HPC from inhibitor addition at 0.31 to 0.42 log for Si and P based inhibitors respectively. This means that maintaining higher residual levels can counteract the relatively modest increases due to inhibitors. BF HPC was affected by pipe material, effluent residual and temperature in addition to a small increase due to inhibitor addition. Biofilm density was most affected by material type, with polyvinyl chloride (PVC) biofilm density consistently much lower than other materials (0.66, 0.92, and 1.22 log lower than lined cast iron (LCI), unlined cast iron (UCI), and galvanized steel (G), respectively). Temperature had a significant effect on both biofilm and bulk HPC levels but it is not practical to alter temperature for public drinking water distribution systems so temperature is not a management tool like residual. This study evaluated the effects of four different corrosion inhibitors (i.e. based on either phosphate or silica) on drinking water distribution system biofilms and bulk water HPC levels. Four different pipe materials were used in the pilot scale experiments, polyvinyl chloride (PVC), lined cast iron (LCI), unlined cast iron (UCI), and galvanized steel (G). Three kinds of phosphate based and one silica based corrosion inhibitors were added at concentrations typically applied in a drinking water distribution system for corrosion control. The data showed that there was a statistically significant increase of 0.34 log in biofilm bacterial densities (measured as HPC) with the addition of any of the phosphate based inhibitors (ortho-phosphorus, blended ortho-poly-phosphate, and zinc ortho-phosphate). A silica based inhibitor resulted in an increase of 0.36 log. The biological data also showed that there was a statistically significant increase in bulk water bacterial densities (measured as heterotrophic plates count, HPC) with the addition of any of the four inhibitors. For bulk HPC this increase was relatively small, being 15.4% (0.42 log) when using phosphate based inhibitors, and 11.0% (0.31 log) for the silica based inhibitor. Experiments with PDS influent spiked with phosphate salts, phosphate based inhibitors, and the silicate inhibitor showed that the growth response of P17 and NOx in the AOC test was increased by addition of these inorganic compounds. For this source water and the PDSs there was more than one limiting nutrient. In addition to organic compounds phosphorus was identified as a nutrient stimulating growth, and there was also an unidentified nutrient in the silica based inhibitor. However since the percentage increases due to inhibitors were no greater than 15% it is unlikely that this change would be significant for the bulk water microbial quality. In addition it was shown that increasing the chloramines residual could offset any additional growth and that the inhibitors could help compliance with the lead and copper rule. However corrosion inhibitors might result in an increase in monitoring and maintenance requirements, particularly in dead ends, reaches with long HRTs, and possibly storage facilities. In addition it is unknown what the effect of corrosion inhibitors are on the growth of coliform bacteria and opportunistic pathogens relative to ordinary heterotrophs. A method was developed to monitor precision for heterotrophic plate count (HPC) using both blind duplicates and lab replicates as part of a project looking at pilot drinking water distribution systems. Precision control charts were used to monitor for changes in assay variability with time just as they are used for chemical assays. In adapting these control charts for the HPC assay, it was determined that only plate counts ≥ 30 cfu per plate could be used for Quality Assurance (QA) purposes. In addition, four dilutions were used for all known Quality Control (QC) samples to insure counts usable for QC purposes would be obtained. As a result there was a 50% increase in the required labor for a given number of samples when blind duplicates and lab replicates were run in parallel with the samples. For bulk water HPCs the distributions of the duplicate and replicate data were found to be significantly different and separate control charts were used. A probability based analysis for setting up the warning limit (WL) and control limit (CL) was compared with the method following National Institute of Standard and Technology (NIST) guidelines.

No description available.

O objetivo do trabalho foi avaliar a utilização da metodologia de citometria de fluxo na determinação da contagem bacteriana total (CBT) do leite cru. No primeiro estudo foi avaliado o efeito da temperatura de armazenamento, da idade da amostra e do tipo conservante sobre a CBT. Também foi estudada a possibilidade de se utilizar uma única amostra de leite para a realização das análises previstas na Instrução Normativa 51 (IN-51). Foram testadas, três temperaturas de armazenamento (0oC - congelado, 7oC - resfriamento e 24 oC - ambiente), três conservantes (bronopol, azidiol e sem conservante) e quatro tempos entre a coleta e a análise (idade da amostra) (um (D1), três (D3), cinco (D5) e sete (D7) dias). Foi considerado tratamento controle para análise de CBT, amostras refrigeradas, com azidiol e com um dia de idade. Para as análises de composição e CCS, o tratamento controle foram amostras refrigeradas, com bronopol e com um dia de idade. Os resultados indicaram que será necessária a coleta de duas amostras, uma destinada à determinação de CCS e composição, contendo bronopol e, outra, para CBT, contendo azidiol. A amostra para CBT poderá ser analisada em até sete dias após a coleta, desde que mantida sob refrigeração à 7ºC. Deve-se evitar o aquecimento ou o congelamento da amostra para CBT, bem como garantir a adição do azidiol. O segundo estudo teve como objetivo estabelecer a correlação entre os métodos de referência e de citometria de fluxo na determinação da CBT. Amostras coletadas nos meses de junho à setembro (n=155) foram agrupadas considerando-se estação da seca e as coletadas nos meses de novembro e dezembro (n=68), estação das águas. Foram realizadas análises simultâneas pelos métodos de referência (contagem padrão em placas) e de citometria de fluxo (equipamento Bactocount), sendo os resultados expressos em unidades formadoras de colônia (UFC) e contagem individual de bactérias (CIB), respectivamente. As equações lineares de correlação entre a CIB e UFC foram semelhantes nas estações, indicando que uma única equação pode ser utilizada ao longo do ano para transformar os resultados de CIB para UFC. A equação linear obtida foi: log(UFC) = log(CIB) x 0,7224 + 1,4617 com coeficiente de correlação de 0,8125. A acurácia do equipamento Bactocount na estimativa do valor de referência, expressa pelo erro padrão (s(y,x)), foi de 0,309 log UFC/mL. Os resultados mostraram que o equipamento Bactocount pode ser calibrado para expressar os resultados em UFC e com isso ser utilizado no monitoramento da qualidade do leite. / The objective of this study was to evaluate the utilization of electronic flow cytometry to determine total bacterial count (TBC) of raw milk. In the first experiment, the effect of storage temperature, sample age and milk preservative type on TBC were evaluated. Additionally, the use of a single milk sample to performance regulatory milk analysis under the Normative Instruction 51 (NI-51) was tested. Effects were standardized as: three storage temperatures (0ºC - freezer, 7ºC - refrigerator and 24ºC - room temperature), four sample ages (1 (D1), 3 (D3), 5 (D5) and 7 (D7) days) and three milk preservatives (bronopol, azidiol and no preservative). Control treatment for TBC analysis was defined as refrigerated milk samples containing azidiol with 1 day of storage. For determination of milk components and somatic cell count (SCC), control treatment was defined as refrigerated milk samples containing bronopol with 1 day of storage. Results of the first experiment showed that two milk samples are necessary to performance regulatory milk analysis under the NI-51; one containing bronopol should be used for determination of milk components and SCC, and other containing azidiol for TBC. Milk samples used for TBC can be tested until 7 days after sampling when they are kept at 7ºC. Freezing or heating milk samples for TBC analysis should be avoided and addition of azidiol is always necessary. The second experiment was designed to determine a correlation between two methods of TBC, electronic flow cytometry and standard plate count. Milk samples collected from June to September (n = 155) were named as dry season samples and milk samples collected in November and December (n = 68) were named as rainy season samples. Each milk sample was used to run both methods of TBC. Results were expressed as individual bacterial count (IBC) and colony forming unit (CFU) for electronic flow cytometry (Bactocount) and standard plate count, respectively. The linear equations of correlation between IBC and CFU had similar patterns in both seasons, dry and rainy, indicating that a single equation can be used to transform IBC results in CFU along the year. The linear equation was defined as log(CFU) = 0.7224 x log(IBC) + 1.4617 with coefficient of correlation of 0.8125. The accuracy of Bactocount in estimating reference values, denoted by the standard error (s(y,x)), was 0.309 log CFU/mL. The results showed that Bactocount can be calibrated to express TBC readings in CFU and, consequently, be used to monitor milk quality.

O objetivo do trabalho foi avaliar a utilização da metodologia de citometria de fluxo na determinação da contagem bacteriana total (CBT) do leite cru. No primeiro estudo foi avaliado o efeito da temperatura de armazenamento, da idade da amostra e do tipo conservante sobre a CBT. Também foi estudada a possibilidade de se utilizar uma única amostra de leite para a realização das análises previstas na Instrução Normativa 51 (IN-51). Foram testadas, três temperaturas de armazenamento (0oC – congelado, 7oC – resfriamento e 24 oC – ambiente), três conservantes (bronopol, azidiol e sem conservante) e quatro tempos entre a coleta e a análise (idade da amostra) (um (D1), três (D3), cinco (D5) e sete (D7) dias). Foi considerado tratamento controle para análise de CBT, amostras refrigeradas, com azidiol e com um dia de idade. Para as análises de composição e CCS, o tratamento controle foram amostras refrigeradas, com bronopol e com um dia de idade. Os resultados indicaram que será necessária a coleta de duas amostras, uma destinada à determinação de CCS e composição, contendo bronopol e, outra, para CBT, contendo azidiol. A amostra para CBT poderá ser analisada em até sete dias após a coleta, desde que mantida sob refrigeração à 7ºC. Deve-se evitar o aquecimento ou o congelamento da amostra para CBT, bem como garantir a adição do azidiol. O segundo estudo teve como objetivo estabelecer a correlação entre os métodos de referência e de citometria de fluxo na determinação da CBT. Amostras coletadas nos meses de junho à setembro (n=155) foram agrupadas considerando-se estação da seca e as coletadas nos meses de novembro e dezembro (n=68), estação das águas. Foram realizadas análises simultâneas pelos métodos de referência (contagem padrão em placas) e de citometria de fluxo (equipamento Bactocount), sendo os resultados expressos em unidades formadoras de colônia (UFC) e contagem individual de bactérias (CIB), respectivamente. As equações lineares de correlação entre a CIB e UFC foram semelhantes nas estações, indicando que uma única equação pode ser utilizada ao longo do ano para transformar os resultados de CIB para UFC. A equação linear obtida foi: log(UFC) = log(CIB) x 0,7224 + 1,4617 com coeficiente de correlação de 0,8125. A acurácia do equipamento Bactocount na estimativa do valor de referência, expressa pelo erro padrão (s(y,x)), foi de 0,309 log UFC/mL. Os resultados mostraram que o equipamento Bactocount pode ser calibrado para expressar os resultados em UFC e com isso ser utilizado no monitoramento da qualidade do leite. / The objective of this study was to evaluate the utilization of electronic flow cytometry to determine total bacterial count (TBC) of raw milk. In the first experiment, the effect of storage temperature, sample age and milk preservative type on TBC were evaluated. Additionally, the use of a single milk sample to performance regulatory milk analysis under the Normative Instruction 51 (NI-51) was tested. Effects were standardized as: three storage temperatures (0ºC – freezer, 7ºC – refrigerator and 24ºC – room temperature), four sample ages (1 (D1), 3 (D3), 5 (D5) and 7 (D7) days) and three milk preservatives (bronopol, azidiol and no preservative). Control treatment for TBC analysis was defined as refrigerated milk samples containing azidiol with 1 day of storage. For determination of milk components and somatic cell count (SCC), control treatment was defined as refrigerated milk samples containing bronopol with 1 day of storage. Results of the first experiment showed that two milk samples are necessary to performance regulatory milk analysis under the NI-51; one containing bronopol should be used for determination of milk components and SCC, and other containing azidiol for TBC. Milk samples used for TBC can be tested until 7 days after sampling when they are kept at 7ºC. Freezing or heating milk samples for TBC analysis should be avoided and addition of azidiol is always necessary. The second experiment was designed to determine a correlation between two methods of TBC, electronic flow cytometry and standard plate count. Milk samples collected from June to September (n = 155) were named as dry season samples and milk samples collected in November and December (n = 68) were named as rainy season samples. Each milk sample was used to run both methods of TBC. Results were expressed as individual bacterial count (IBC) and colony forming unit (CFU) for electronic flow cytometry (Bactocount) and standard plate count, respectively. The linear equations of correlation between IBC and CFU had similar patterns in both seasons, dry and rainy, indicating that a single equation can be used to transform IBC results in CFU along the year. The linear equation was defined as log(CFU) = 0.7224 x log(IBC) + 1.4617 with coefficient of correlation of 0.8125. The accuracy of Bactocount in estimating reference values, denoted by the standard error (s(y,x)), was 0.309 log CFU/mL. The results showed that Bactocount can be calibrated to express TBC readings in CFU and, consequently, be used to monitor milk quality.