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J. Dairy Sci. 87:1151-1157
© American Dairy Science Association, 2004.

Predominant Microflora of Downgraded Danish Bulk Tank Milk

C. Holm1, L. Jepsen1, M. Larsen2 and L. Jespersen3

1 Danish Dairy Board, Department of Veterinary and Milk Quality, Frederiks Allé 22, DK-8000 Århus C, Denmark
2 Steins Laboratory A/S, Hjaltesvej 8, DK-7500 Holstebro, Denmark
3 The Royal Veterinary and Agricultural University (KVL), Department of Dairy and Food Science, Food Microbiology, Rolighedsvej 30, DK-1958 Frederiksberg C, Denmark

Corresponding author: L. Jespersen; e-mail: lj{at}kvl.dk.


   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 
The microflora of downgraded Danish bulk tank milk was examined to identify the main causes of increased microbial counts. Seventy-five representative samples with a microbial count exceeding 3.0 x 104 cfu/mL were selected for a more detailed microbial examination. A total of 1237 isolates from these samples were identified. Gram-negative, oxidase-positive bacteria were found in 72% of the samples. Coliforms were found in 20% of the samples, and non-coliforms were found in 49% of the samples. Coryneforms, other gram-positive rods, Lactococcus spp., Micrococcus spp., and coagulase-negative Staphylococcus spp. were found in 28 to 53% of the samples. Bacillus spp., Enterococcus spp., Staphylococcus aureus, Streptococcus dysgalactiae, Streptococcus uberis, and yeasts were found in <25% of the samples. Additionally, the isolates were divided into 3 groups, based on the main cause of an elevated microbial count. Microorganisms primarily associated with poor hygiene dominated the microflora in 64% of the samples; bacteria also related to poor hygiene, but in addition associated with growth at low temperatures (psychrotrophic bacteria) dominated the microflora in 28% of the samples; and bacteria mainly associated with mastitis dominated the microflora in 8% of the samples. A bulk tank milk storage period of 48 h instead of 24 h did not affect the proportion of downgraded milk samples and could not be associated with a specific group of microorganisms. Further, no relationship was found between somatic cell counts and the presence of mastitis bacteria.

Key Words: hygiene • psychrotrophic • mastitis • microflora


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 
During the milking procedure, milk may be contaminated from 3 main sources: the milking equipment, the teat surface, and from within the udder. When the milk is collected and stored in a bulk tank, the cooling capacity, holding temperature, and storage time will influence the ability of the contaminants to grow (Bramley and McKinnon, 1990; Slaghuis, 1996).

Bedding material, manure, mud, and feeds are typically found in the environment of cows and will inevitably contaminate the teat surfaces. Used bedding material has been reported to contain 108 to 1010 cfu/g, and milk from unwashed, heavily soiled cows may have microbial counts >105 cfu/mL (Bramley and McKinnon, 1990). Contaminating bacteria from teat surfaces have been reported as Micrococcus spp., coagulase-negative Staphylococcus spp., Enterococcus spp., coryneforms, Bacillus spp., coliforms, and other gram-negative rods (Hogan et al., 1989; Bramley and McKinnon, 1990).

Milk deposits or remaining washing water on insufficiently cleaned milking equipment might promote the growth of most microorganisms, but will typically select for faster-growing bacteria such as Lactococcus spp., Pseudomonas spp., and coliforms (Murphy and Boor, 2000). When the milk enters the bulk tank, it is cooled to <5°C at a speed depending on the efficiency of the cooler. During the cooling period and especially if the milk does not reach the intended temperature, some microorganisms may multiply. Especially fast-growing psychrotrophic bacteria, such as Pseudomonas fluorescens, will be favored in a refrigerated bulk tank (Gennari and Dragotto, 1992; Ternstroem et al., 1993; Heeschen, 1996).

In bulk tank milk, bacteria originating from cows with subclinical or clinical mastitis (udder infections) may be present. The number and type of bacteria from cows with subclinical mastitis differ from cows with clinical mastitis. In Denmark, Staphylococcus aureus or coagulase-negative Staphylococcus spp. were found in 76% of the milk samples of subclinical mastitis cases, and Streptococcus uberis or Streptococcus dysgalactiae were found in most other milk samples from cows with subclinical mastitis (Aagaard, 2001). In samples from cows with clinical mastitis, Strep. uberis was found in 24% of the samples, coliform bacteria in 14%, Staphylococcus spp. in 12%, Strep. dysgalactiae in 7%, and Lactococcus spp., Enterococcus spp., hemolytic Streptococcus spp., and Arcanobacterium pyogenes were each present in <5% of the samples; no growth was observed in 24% of the milk samples (Aagaard, 2001). Although inclusion of milk from cows with subclinical mastitis has been reported to contribute <104 cfu/mL to the bulk tank milk, the inclusion of milk from cows with clinical mastitis may elevate the bulk tank milk to 105 cfu/mL (Bramley and McKinnon, 1990). However, because the milk from cows with clinical mastitis has a clotted appearance, it is only accidentally included in the bulk tank milk (Bramley and McKinnon, 1990).

In Denmark, the microbial count is determined once a week for all farmers by use of a BactoScan 8000S instrument (Sørensen et al., 1996). A noteworthy part of these samples has a microbial count exceeding the grading level of 3.0 x 104 cfu/mL (Aagaard, 2001). Despite the economical consequences, little is known about the predominant microflora of the downgraded samples.

The aim of the present study was to identify the predominant microflora in downgraded Danish bulk tank milk in order to identify the main causes of elevated microbial counts. The isolates from 75 representative samples were initially identified to the group, genus, or species level depending on the organism. Isolates were then divided into 3 groups indicating the main cause of the elevated microbial count. Furthermore, the influence on the microflora of storing milk in the bulk tank for a period of 48 h instead of 24 h was examined. Finally, the relationship between mastitis bacteria and somatic cells was evaluated.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 
Sampling
Samples taken weekly from the bulk tank of 5923 farmers were initially included in the examination in order to choose 75 samples for further microbial examination. All collected samples were analyzed for microbial count using BactoScan 8000S (FOSS Analytical A/S, Hillerød, Denmark), and the SCC was obtained with a Fossomatic 5000 instrument (FOSS Analytical A/S). The BactoScan 8000S expresses the microbial count in individual bacterial counts (IBC)/mL, which, in Denmark, are converted to cfu by use of the formula: log cfu = log IBC x 0.923 – 0.087 (Sørensen et al., 1996). The BactoScan 8000S and Fossomatic analyses were performed according to the supplier’s manual. On 3 occasions (May 27, June 12, and June 20, 2002), 25 random samples with >3.0 x 104 cfu/mL were selected for microbial examination. To test the effect of milk storage time on microflora, the milk storage times of the farms were obtained for 73 of the 75 selected samples (for unknown reasons the storage times of 2 samples could not be provided by the dairy).

Enumeration of Microorganisms
For each milk sample, the dilutions 10–2, 10–3, and 10–4 were prepared according to the IDF standard (Anonymous, 1996). Inoculation, incubation, and enumeration of the samples were performed according to the IDF standard (Anonymous, 1991). One milliliter of the dilution was, in duplicate, transferred to Petri dishes, and 12 to 15 mL plate count agar containing 5% reconstituted nonfat dried milk produced according to the IDF standard (Anonymous, 1991) were poured into each Petri dish. The inoculum and the medium were carefully mixed and allowed to solidify. The Petri dishes were inverted and incubated at 30 ± 1°C for 72 ± 3 h. After incubation, the colonies on the Petri dishes were counted, and this count was divided by the dilution factor to obtain the cfu/mL.

Identification and Enumeration of Isolates
For each sample, a Petri dish with 20 to 200 colonies was randomly chosen. From each dish, 20 colonies were randomly selected by drawing a slice including exactly 20 colonies per Petri dish. All colonies in this area were streaked on plate count agar containing 5% reconstituted nonfat dried milk and incubated at 30°C for 24 h. If there were no visible colonies present on the Petri dish after 24 h, the incubation period was prolonged up to 72 h. In total, 1500 isolates were picked; of these, 1237 (82.5%) were able to grow after streaking.

The procedure used for identification of the isolates is described subsequently. The isolates were initially grouped in gram-positive rods, gram-positive cocci, gram-negative bacteria, and yeasts by microscopy and the KOH method for identifying gram-negative bacteria (Gregersen, 1978; Ryu, 1938).

Gram-positive rods were divided in Bacillus spp., coryneforms, and other gram-positive rods by macro- and micromorphological examinations. Gram-positive cocci were subjected to a catalase test by adding a colony to a drop of 3% hydrogen peroxide solution. Catalase-positive isolates were inoculated in a modified Hugh and Leifson’s medium (Oxoid) and incubated at 30°C for 24 h to differentiate between fermentative Staphylococcus spp. and oxidative Micrococcus spp. (Aalbaek et al., 1981). The fermentative isolates were streaked onto blood agar plates and incubated at 30°C for 24 h (Nylin, 1996) and tested for coagulase activity by incubating colony mass in rabbit coagulase plasma with EDTA (BD, Franklin Lakes, NJ) at 37°C for 4 h. Hemolytic and coagulase-positive strains were identified as Staph. aureus and coagulase-negative Staphylococcus spp. were grouped as such. Catalase-negative isolates were streaked onto blood agar plates with aesculin and onto blood agar plates with 2 different concentrations of penicillin (0.1 and 1.0 IU/mL) and incubated at 30°C for 24 h. After incubation, the catalase-negative isolates were identified according to their hemolytic properties, their ability to hydrolyze aesculin, and their ability to grow in the presence of penicillin (Nylin, 1996). Hemolytic isolates were interpreted as hemolytic Streptococcus spp. (including Streptococcus agalactiae). Nonhemolytic and aesculin-negative isolates were interpreted as Strep. dysgalactiae. Non-hemolytic and aesculin-positive isolates with no growth on blood plates with penicillin were interpreted as Strep. uberis. Non-hemolytic and aesculin-positive isolates growing on blood plates with 0.1 IU/mL penicillin were interpreted as Lactococcus spp. Non-hemolytic and aesculin-positive isolates growing on blood plates with 1.0 IU/mL penicillin were interpreted as Enterococcus spp.

Gram-negative rods were tested for oxidase activity using Bactident oxidase test strips (Merck, Whitehouse Station, NJ). Of the oxidase-positive isolates, 16 randomly selected isolates were further identified to the species level by use of API 20 NE (bioMerieux, Marcy l’Etoile, France), which were incubated at 30°C for 24 to 48 h before reading. The API 20 NE profiles were interpreted by the APILAB PLUS software v.3.3.3 (bioMerieux). Oxidase-negative isolates were inoculated in MacConkey broth at 30°C for 24 h to differentiate coliforms from non-coliforms (Aalbaek et al., 1981).

In total, 62 randomly selected yeast isolates were identified by determination of their macro- and micromorphological characteristics as described by Yarrow (1998) and by use of the API ID32C kit (bioMerieux), which were incubated at 25°C for 48 to 72 h according to the supplier’s manual. The API ID32C profiles were interpreted by the APILAB PLUS software v.3.3.3 (bioMerieux). The isolates were finally identified to the species level according to the description given by Kurtzman and Fell (1998).

For each sample, the counts for the different types of microorganisms were calculated by dividing the number of a given type of microorganism with the number of recovered isolates (maximum of 20) and multiplying it with the total colony count of the sample. The geometric mean was calculated for each type of microorganism by including only the samples from where the particular type of microorganism was identified.


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 
Table 1Go shows the geometric means of the microbial count and the percentages of downgraded Danish milk samples during the previous 3 yr (2000 to 2002). The geometric mean has been slightly decreasing over the years, and the relative proportion of samples with microbial count >3.0 x 104 cfu/mL has been stable and accounts for <7%. In Table 2Go, the geometric means of the microbial count and percentages of samples >3.0 x 104 cfu/mL on the 3 sampling days are shown. These results show that samples investigated in this study were taken on days resembling the yearly geometric means with counts between 7.3 and 7.5 x 103 cfu/mL and 6 to 7% of the samples >3.0 x 104 cfu/mL.


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  		Table 1. Microbial counts of Danish bulk tank milk (2000–2002).
 

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  		Table 2. Microbial counts of Danish bulk tank milk on sampling days.
 
Table 3Go shows the results of the identification of 1237 representative isolates from 75 samples with >3.0 x 104 cfu/mL. Gram-negative, oxidase-positive bacteria were found in 72% of the samples with a geometric mean of 2.5 x 104 cfu/mL. Coliforms were found in 20% of the samples with a geometric mean of 1.7 x 104 cfu/mL, and non-coliforms were found in 49% of the samples with a geometric mean of 1.3 x 104 cfu/mL. Coryneforms, other gram-positive rods, Lactococcus spp., Micrococcus spp., and coagulase-negative Staphylococcus spp. were found in 28 to 53% of the samples with geometric means between 0.8 and 1.5 x 104 cfu/mL. Bacillus spp., Enterococcus spp., Staph. aureus, Strep. Dysgalactiae, and yeasts were found in <25% of the samples with geometric means <1.0 x 104 cfu/mL. Streptococcus uberis was found in 15% of samples with a geometric mean of 3.4 x 104 cfu/mL.


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  		Table 3. Occurrence and number (cfu/mL) of microorganisms in Danish bulk tank milk with >3.0 x 104 cfu/mL.
 
In Table 4Go, the identification of gram-negative, oxidase-positive isolates is presented. The identification is based on 16 randomly selected isolates. For all of the isolates, the reliability of the identifications as performed by the API 20 NE (bioMerieux,) was >90%. Of these isolates, 7 were identified as P. fluorescens, 2 as Chryseobacterium indologenes, 1 as Alcaligenes odorens, 1 as Comamonas acidovorans, 1 as Pasteurella heamolytica, 1 as Pseudomonas pseudomallei, and 3 isolates could only be identified to the Pseudomonas spp. level. The 62 representative yeast isolates were identified based on both their morphology and the results obtained by the API ID 32C kit. Of the isolates, 61% were identified as Debaryomyces hansenii (Candida famata), 24% as Kluyveromyces marxianus (Candida kefyr), 3% as Debaryomyces polymorphus, 3% as Cryptococcus curvatus, and 1.6% each as Candida sake, Pichia fermentans, Candida zeylanoides, Dipodascus capitatus, and Trichosporon asahii. The identifications based on the API ID 32C kit alone resulted in identification values from 84 to 100%.


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  		Table 4. Identification of gram-negative, oxidase-positive bacteria isolated from Danish bulk tank milk with >3.0 x 104 cfu/mL1.
 
The proportion of downgraded bulk tank milk samples dominated (i.e., >50% of the isolates) by one type (group, genus, or species depending on the level of identification) of microorganism is shown in Table 5Go. For 66% of the samples, one type of microorganism dominated the microflora. Gram-negative, oxidase-positive bacteria dominated in 28% of the samples. In 8% of the samples, gram-positive rods other than Bacillus spp. and coryneforms dominated, and Lactococcus spp. dominated in 7% of the samples. Other types of microorganisms were found to be dominant in <=4% of the samples.


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  		Table 5. Proportion of downgraded Danish bulk tank milk samples with >=50% of a single microorganism present.
 
The identified isolates were grouped in 3 different categories indicating the main causes of the elevated microbial counts. Bacillus spp., coliforms, coryneforms, Enterococcus spp., Lactococcus spp., Micrococcus spp., non-coliforms, other gram-positive rods, and yeasts are considered mainly to be associated with poor udder hygiene and equipment cleaning and, therefore, are grouped as hygiene microorganisms. The gram-negative, oxidase-positive bacteria, which are dominated by P. fluorescens, are also related to poor hygiene, but in addition are associated with growth at low temperatures and, therefore, are grouped as psychrotrophic hygiene bacteria. Staphylococcus spp. and Streptococcus spp. are the most common bacteria found in milk from cows with mastitis and, thus, are grouped as mastitis bacteria. As shown in Table 6Go, microorganisms associated with poor hygiene were found in 95% of the samples with a geometric mean of 4.7 x 104 cfu/mL, psychrotrophic hygiene bacteria were found in 72% of the samples with a geometric mean of 2.5 x 104 cfu/mL, and mastitis bacteria were found in 48% of the samples with a geometric mean of 1.5 x 104 cfu/mL. Using the previously described groups, 64% of the samples with >3.0 x 104 cfu/mL were dominated by microorganisms associated with poor hygiene, 28% of the samples were dominated by psychrotrophic bacteria, and in 8% of the samples, the majority of the isolates belonged to the mastitis group.


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  		Table 6. Microbial groups indicating the main causes of elevated microbial counts and their number (cfu/mL) in Danish bulk tank milk >3.0 x 104 cfu/mL.
 
The influence of milk storage time on the distribution and count of downgraded milk was examined. The storage time of 73 samples >3.0 x 104 cfu/mL was compared with the storage time distribution of all Danish farmers. Of the 73 samples, 16.4% had 24-h storage and 83.6% had 48-h storage of the milk before it was collected, which was found not to be significant (P = 0.24) using the binomial distribution among Danish farmers (20.5% of all Danish milk producers have 24-h storage and 79.5% have 48-h storage of the milk before it is collected) for calculation. The average count of the samples that had been stored for 24 h was 7.9 x 104 cfu/mL compared with 8.7 x 104 cfu/mL for samples that had been stored for 24 h. This difference was not found to be significant (P = 0.88). The storage time could not be associated with a specific group of microorganisms or their origin. Of the samples dominated by microorganisms associated with poor hygiene, 17.4% had 24-h storage and 82.6% had 48-h storage of the milk before it was collected. Of the samples dominated by psychrotrophic hygiene bacteria, 14.3% had 24-h storage and 85.7% had 48-h storage of the milk before it was collected. Of the samples dominated by mastitis bacteria, 16.7% had 24-h storage and 83.3% had 48-h storage. Using the binomial distribution of all Danish farmers for calculation, none of these distributions were found to be significant (P = 0.38, 0.35, and 0.64, respectively).

The average SCC of the 5923 samples included in the study was 2.7 x 105 cells/mL and varied between 2.0 x 104 and 1.2 x 106 cells/mL. The average SCC of the selected 75 samples with >3.0 x 104 cfu/mL was 2.9 x 105 cells/mL and varied between 6.6 x 104 and 5.1 x 105 cells/mL. Analysis of variance showed no significant difference between these 2 averages. In 8% of the 75 samples with >3.0 x 104 cfu/mL, mastitis bacteria were found to dominate the samples. The average SCC of these samples was 3.6 x 105 cells/mL and varied between 1.5 x 105 and 5.0 x 105 cells/mL, but no significant (P = 0.053) difference from the average of all samples was found using analysis of variance.


   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
ACKNOWLEDGEMENTS
 REFERENCES
 
The geometric mean of the microbial count of Danish milk and the fraction of downgraded samples are low and comparable with other industrialized countries (Rombaut et al., 2002). When samples are downgraded, the present study shows that, in most samples, a single type (group, genus, or species depending on the level of identification) of microorganism causes the increased microbial count. Gram-negative, oxidase-positive bacteria were found to be the most common type of bacteria present in downgraded Danish milk. The dominance of one type of microorganism is supported by a Scottish study in which samples with >4.5 x 104 cfu/mL were examined. Approximately 80% of these samples were found to be dominated by a single type of microorganism. Contrary to the present study where gram-negative, oxidase-positive bacteria were found to be the most common type, Strep. uberis was found to be the most common single type of microorganism causing elevated microbial counts in Scottish milk (Jeffrey and Wilson, 1987).

Microorganisms belonging to the Bacillus spp., coliforms, coryneforms, Enterococcus spp., Lactococcus spp., Micrococcus spp., non-coliforms, other gram-positive rods, and yeasts were individually found at a low level and only dominanted in few samples. However, as these microorganisms are reported to be associated with poor hygiene, it was shown that poor hygiene is the dominating cause of elevated counts in Danish milk. A subgroup of the microorganisms associated with poor hygiene, consisting of thermoduric bacteria such as Bacillus spp. and Micrococcus spp., are normally found in more persistent milk deposits in cracked rubber parts (Bramley and McKinnon, 1990). Thus, the level of Bacillus spp. and Micrococcus spp. found in this study indicates the presence of persistent milk deposits on milking equipment. The majority of the yeast isolates were identified as D. hansenii, the presence of which has previously been reported in milk (Fleet and Mian, 1987; van den Tempel and Jakobsen, 1998). In a previous Danish study, 37 yeast isolates taken from the raw milk silo at 4 different Danablu dairies were identified. The isolates included D. hansenii, but, contrary to the present study, no isolates of K. marxianus were found (van den Tempel and Jakobsen, 1998).

The results show that gram-negative, oxidase-positive bacteria were found in 72% of the tested milk samples and dominated the microflora in 28% of the samples. The identification of 16 of these isolates indicated that P. fluorescens and Pseudomonas spp. are the most common species in this group; species such as C. indologenes, A. odorens, Com. acidovorans, and Pasteurella heamolytica may occur. Psychrotrophic bacteria including Pseudomonas spp., Chryseobacterium spp., and Alcaligenes spp. are ambient in the cow’s environment; the teats are known to harbor high numbers especially (Morse et al., 1968; Bramley and McKinnon, 1990). Pseudomonas fluorescens has the ability to produce exopolysaccharides, which enables it to produce biofilm on milking equipment and in the bulk tank (Read and Costerton, 1987; Jayarao and Wang, 1999). Additionally, P. fluorescens is a fast-growing organism with a generation time of 7 h at 4 to 6°C (Ingraham and Stokes, 1959). Thus, the finding of P. fluorescens and other psychrotrophic isolates in this study suggests a selective growth in the refrigerated bulk tank and during the time from milk collection until sample analysis. A high prevalence of P. fluorescens has previously been reported in randomly selected bulk milk samples and should not be considered unique for samples >3.0 x 104 cfu/mL (Gennari and Dragotto, 1992; Ternstörm et al., 1993; Desmasures et al., 1997; Michel et al., 2001).

Mastitis bacteria were found in 48% of the samples and were found to be the main cause of elevated microbial count in 8% of the samples. The isolation of coagulase-negative Staphylococcus spp., Staph. aureus, Strep. dysgalactiae, and Strep. uberis was expected based on the knowledge of the most frequent mastitis pathogens in Denmark (Aagaard, 2001). Hemolytic Streptococcus spp. mastitis pathogens such as Strep. agalactiae were not found among the identified isolates, which agrees well with the knowledge that hemolytic Streptococcus spp. is only isolated from 2.0% of the subclinical mastitis cases and 3.2% of the clinical mastitis cases in Denmark (Aagaard, 2001). In Scottish milk, 43.8% of the samples >4.5 x 104 cfu/mL have been found to be dominated by mastitis bacteria, with Strep. uberis dominating in 17.8% of the samples and Strep. agalactiae dominating in 13.5% of the samples (Jeffrey and Wilson, 1987).

Storage of milk for 2 d instead of one did not affect the proportion or count of downgraded milk samples and could not be associated with growth of a specific group of microorganisms. Prolonged storage at low temperature is expected to select for psychrotrophic bacteria, and, therefore, a higher percentage of this group in milk stored for 2 d was expected (Ingraham and Stokes, 1959). However, this was not found.

The average SCC of samples from where mastitis bacteria were isolated did not differ significantly from the average SCC of all samples. Neither did the samples dominated by mastitis bacteria. A relationship between mastitis bacteria and the SCC has previously been reported (Fenlon et al., 1995). Based on this knowledge, it was expected that samples dominated by mastitis bacteria and with a microbial count >3.0 x 104 cfu/mL would have an increased number of somatic cells. However, this was not observed in the present study.

Overall, the level of bacteria in Danish bulk tank milk is low and stable. The study also shows that milk with an elevated microbial count is typically dominated by one type of microorganism. Of these microorganisms, gram-negative, oxidase-positive bacteria were found to be the most common type. When microorganisms associated with poor hygiene were grouped, it was found to be the main cause of elevated microbial counts in Danish milk. A tank milk storage period of 48 h instead of 24 h could not be shown to affect the proportion or count of downgraded milk samples or to be associated with a specific group of microorganisms. No relationship was found between SCC and the presence of mastitis bacteria.


   ACKNOWLEDGEMENTS
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
REFERENCES
 
The present work was supported by The Royal Veterinary and Agricultural University, The Danish Dairy Board, FOSS Analytical A/S, and the Danish Academy of Technical Sciences. The competent technicians from Steins Laboratory A/S and Jørn Andreasen from FOSS Analytical A/S are thanked for their skillful technical assistance with identifying the isolates.

Received for publication June 23, 2003. Accepted for publication December 23, 2003.


   REFERENCES
TOP
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 ACKNOWLEDGEMENTS
 REFERENCES
 


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Kurtzman, C. P., and J. W. Fell. 1998. The Yeasts, A Taxonomic Study, 4th ed. Elsevier, Amsterdam.

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Morse, P. M., H. Jackson, C. H. McNaughton, A. G. Leggatt, G. B. Landerkin, and C. K. Johns. 1968. Investigation of factors contributing to the bacteria count of bulk tank milk. II. Bacteria in milk from individual cows. J. Dairy Sci. 51:1188–1191.

Murphy, S. C., and K. J. Boor. 2000. Trouble-shooting sources and causes of high bacteria counts in raw milk. Dairy Food Environ. Sanit. 20:606–611.

Nylin, B. 1996. Esculin—Blood agar as a diagnostic tool in acute mastitis for the practitioner. Pages 270–273 in XIX World Buiatrics Congress, Edinburgh, Scotland.

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