Bench Scale Tests

Bacteria Reduction: A series of experiments was designed to quantify bacteria reduction in raw milk due to UV treatment. In all three protocols, milk was inoculated to ensure high bacteria counts for all major categories of bacteria common in raw milk. The inoculants contained strains of bacteria that had been isolated from raw milk samples taken from Wisconsin dairy farms. Two different strains of each of the following bacteria types were used for the inoculants.

 Streptococcus uberis  S. dysgalctia  S. thermophilus  Lactobacillus casei  Escherichia coli  Staphylococcus aureus  Bacillus sp.  Pseudomonas sp.  Enterobacteria sp.  Citrobacter freundi  Serratia marcesceas  Enterobacter cloacae  Enterobacter faecalis

Bacteria colonies were removed from agar plates using a milk sample loop. Each loop was immersed in a container of 250 mL of milk and the loops were vigorously stirred in the milk. This solution was then allowed to incubate at room temperature for 12 hours before being added to the experimental milk samples. Treatment protocol three allowed additional incubation time to further increase bacteria counts. The bacteria counts at the start of all treatments were therefore higher than would be expected in raw milk. The following three treatment protocols, representative of on- farm treatment scenarios were investigated.

Treatment 1. UV treatment immediately after milk harvest (milk temperature 36°C). A sample of 20 litres of raw milk was collected from the UW milking parlour by milking a cow directly into a collection bucket. This milk was transported to the UW milking lab, inoculated and UV treatment was applied within ½ hour of milk collection (Initial counts: SPC 6.8 x 106 cfu.mL-1 psych. 6.8 x 106 cfu.mL-1, therm. 1.6 x 104 cfu.mL-1, coli. 7.0 x 104 cfu.mL-1). This treatment simulated a scenario in which milk might be treated ‘in-line’ as it is transferred from the receiver in a milking parlour to a bulk storage tank.

Treatment 2. UV treatment after milk refrigeration (Milk temperature 4°C). A sample of 20 litres each of raw milk was collected from the refrigerated bulk tank in the UW milking parlour. This milk was transported to the UW milking lab, inoculated and UV treatment applied about 2 hours after milk collection to allow time for the milk to be chilled to storage temperature (Initial counts: SPC 9.9 x 106 cfu.mL-1, psych. 1.0 x 106 cfu.mL-1, therm. 8.4 x 102 cfu.mL-1, coli 2.0 x 104 cfu.mL-1). This treatment simulated a scenario in which milk might be treated by recirculating cooled milk in a refrigerated bulk storage tank.

Treatment 3. UV treatment after un-refrigerated storage for 6 hours (Milk temperature 25°C). A sample of 20 litres of raw milk was collected from the UW milking parlour by milking a cow directly into a collection bucket. This milk was transported to the UW milking lab, inoculated and then allowed to incubate for 6 hours before UV treatment was applied (Initial counts: SPC 2.7 x 107 cfu.mL-1, psychrotrophs 2.0 x 107

cfu.mL-1, therm. 3.5 x 104 cfu.mL-1, coli 2.3 x 104 cfu.mL-1) simulating a situation in which refrigerated storage is not used on a farm.

UV treatment has been characterized in the literature by an energy delivered per surface area of the treatment device (e.g. mJ.cm-2, Matak et al., 2005). This characterization is difficult to apply for situations which may have very different depths of treatment liquid. An alternate treatment method is to characterize the amount of UV energy delivered per volume of milk treated as follows.

For batch operations in which the liquid is re-circulated through a reactor:

Duv = (Puv x t ) / V (Equation 1)

For single pass, continuous flow operations;

Duv = Puv / Q (Equation 2)

Where:

Duv = UV dose (Joules / liter)

Puv = UV power delivered (Watts)

= Lamp output (Watts) x Transmission efficiency t = Treatment time (s)

V = Treated Volume of liquid (litres) Q = liquid flow rate (litres.s-1)

This method of specifying the UV dose is less ambiguous than energy delivered per surface area especially when the depth of liquid film is variable. As will be shown in the following data the relationship between the UV dose rate as described above and the log reduction in bacteria count per mL was approximately linear over a broad range of bacteria counts. The slope of the regression between log bacteria count per mL and the UV dose in J.L-1 was used to describe a ‘kill rate’ for each bacteria type (log reduction per kJ.L-1 UV dose).

A sample volume of 22 litres of inoculated milk was placed into a milking machine receiver and circulated through the UV treatment unit (wavelength = 254 nm) using a speed controlled milk pump to achieve a flow rate of 1.1 l.s-1. The milk receiver was placed under partial vacuum, typical of that used during milking. An in-line sampler was used to extract milk from the flow stream without stopping the treatment process to avoid excessive exposure of milk in the UV reactor during sampling. Samples were taken after 1, 2, 4, 8, 16, 32 and 64 passes of the sample volume through the UV reactor, corresponding to dose rates of 0.23, 0.46, 0.93, 1.9, 3.7, 7.4 and 1.5 kJ.L-1.

Milk samples were analysed for standard plate count, coliform, psychrotrophs and thermoduric bacteria. Samples were taken on the raw milk, immediately following UV exposure (UV). Duplicate milk samples were taken and each plated in duplicate. Standard plate counts were determined by plating on aerobic plate count Petrifilm (3M, St. Paul, MN) and incubating at 32°C for 48 hours. Thermoduric bacteria counts were conducted by heating 5mL aliquots of milk samples to 62.8°C for 30 minutes and immediately cooling to 10°C in an ice bath. Cooled milk samples were then plated on aerobic plate count Petrifilm with incubation at 32°C for 48 hours. Pyschrotrophic bacteria were enumerated using the modified psychrotrophic bacteria count. Milk was plated on aerobic plate count Petrifilm and incubated at 21°C for 48hours. Coliform counts were conducted by plating samples on E.coli/coliform petrifilm and incubating at 35°C for 24 hours.