1. Assessment of Sanitation Practices Michael S. Ramsey Teaching Laboratory Manager UCD mramsey@ucdavis.edu

2. Talk today assumes risk assessment and design and implementation of Winery Sanitation and Quality Program The link to my talk is still up but brutal to list Google “Michael Ramsey” and “HACCP”

3. Objectives Discuss “Quality” Methods to Assess Sanitation Practices –Other industries –Non-microbiological –Microbiological Cell swabbings ATP Bioluminescence

4. What is Wine Quality? “Quality... you know what it is, yet you don’t know what it is.” –Robert Pirsig (1974); Zen and the Art of Motorcycle Maintenance; pg 178

5. What is Wine Quality All sectors of manufacturing debate the definition of “quality” To the ancient Greeks, “quality” meant “truth” –They would not have understood our usage of the word in our modern wine making dilemma –Aristotle said “Quality (Truth) is not an act, it is a habit” The Oxford English Dictionary (OED) defines “quality”: – “the degree of excellence of the thing”

6. What is Wine Quality? Why isn’t wine quality as easy to define as, say, golf ball quality? “You can’t fake quality any more than you can fake a good meal” - William S. Burroughs

7. What is Wine Quality? Example: the ester, ethyl acetate –Produced by some microbes that can be present during wine fermentation –This character is found at high concentration in wine vinegar – It is also a principle aroma compound of many nail polish removers. People who associate this character with nail polish remover tend to consider it quite objectionable in wines People who associate it with vinegar do not find it as unpalatable

8. What is Wine Quality? Another example is rosemary character This herb character is found in soap products and commonly used in cooking. Individuals who associate the character with a soap find it quite offensive in foods and beverages Those who recognize it as the herb rosemary do not object to its presence.

9. What is Wine Quality? As Pirsig implied, we all have different understandings of “quality” and that these understandings are inherently subjective

10. What is Wine Quality? In other manufacturing sectors, “quality” is about minimizing variability and meeting clear specifications – or “zero defects” OED definition of defect: a shortcoming, imperfection, or lack Origin: late Middle English (as a noun, influenced by Old French defect 'deficiency'): from Latin defectus, past participle of deficere 'desert or fail', from de- (expressing reversal) + facere 'do'

11. What is Wine Quality? If “quality” means “zero defects” then a defective wine is one that is purchased by your consumer in any condition other than how you intended for it to be in the bottle. Wine “Quality” is ultimately how you – or your customer - define it. “Quality means doing it right when no one is looking.” –Henry Ford

12. How does “quality”, then, impact one’s assessment of my sanitation practices?

13. Questions to Ask Yourself What should be the main emphasis of my definition of quality? What are the important areas of risk and sanitation in my winery to look into and why? How much risk am I willing to accept? Which method(s) of assessment are most suitable for my vision of Quality and Quality Assurance?

14. Traditionally, a combination of three methods are used to assess efficacy of sanitation methods –Education and Training –Inspections (visual assessments) –Microbiological testing “We are stuck with technology when what we really want is just stuff that works.” –Douglas Adams, The Salmon of Doubt (2002)

15. Education and Training Requires continuous work due to employee turnover –Which is rarely done

16. Visual Assessments Limited Often, a lack of specificity (terms like satisfactory, acceptable, etc.) leaves the interpretation to the inspector, who may place too much emphasis on relatively unimportant matters and thus increase costs without reducing hazards. Not always reliable to detect sanitary conditions

17. Microbiological Methods of Assessment in Food Molding surfaces with agar Staining surfaces and image analysis with epifluorescence microscope Bacterial impedance (i.e., BacTrac 4300) –A sample is incubated and tested for changes in impedance in a specific medium

18. Microbial Methods in Wineries Microbial Swabbing ATP Bioluminescence

19. Microbiological Swabbing Apply sterile swab over 4 in. x 4 in. surface area Swab is transferred to sterile liquid & vortexed Liquid is put on selective plates which allow microbes to grow

20. Microbiological Swabs

21. Positive attributes –Possible to quantify number and type of microbes present –Can be used to trend sanitary condition of facility/equipment. Technology presumes there's just one right way to do things and there never is. –Robert M. Pirsig, Zen and the Art of Motorcycle Maintenance (1974), p. 166

22. Microbiological Swabs Microbiological testing also has some limitations –Results not available for days –TVC (Total Viable Counts) is not really “total”. It is only a measure of the fraction of the microbes able to produce colonies in the medium used under the conditions of incubation. Difficulties related to sampling –Training, swab material, and technique

23. ATP Bioluminescence Widespread use in dairy, food, and beer industries since the early 1990s ATP is short for adenosine triphosphate. –This compound is the basic currency for energy in all living cells.

24. ATP Bioluminescence The presence of this molecule may be assayed simply using an enzyme and coenzyme complex (luciferase/luciferin) found in the tail of the firefly, Photinus pyralis.

25. ATP Bioluminescence The reaction is essentially the stoichiometric conversion of ATP to photons of light in the presence of the enzyme luciferase

26. ATP Bioluminescence luciferin + ATP + O2 + Mg 2+ oxyluciferin + AMP + PPi + CO2 + light The breakthrough idea was to use ATP from all biological residues on a surface as an indicator of cleanliness, rather than try to ascertain the number of microorganisms present.

27. ATP Bioluminescence The total ATP present on a swabbed surface can be extracted, assayed, and potentially uploaded to a computer database extremely rapidly (within five minutes).

28. ATP Bioluminescence The light given off in this reaction is assayed as Relative Light Units (RLUs) – a measure of the light produced by the ATPase assay relative to a control sample. RLU readings above the number you have chosen (based on the sum of your interferences) would be “FAIL”events in your cleaning/sanitation program.

29. ATP Bioluminescence Since the results are “relative”, we can not compare results from one system to another. However, these assays are sensitive, direct, objective tests of cleaning efficiency and risk.

30. ATP Bioluminescence Although the ATP bioluminescence assay is very rapid, it is not very specific. –The results tell us nothing about the nature of the contaminant, whether it is juice, wine, random organic matter, microorganisms or combinations. There is no direct correlation between ATP results and microbial counts: ATP amounts can vary widely between microbes.

31. ATP Bioluminescence If your QA strategy is to provide quick and sensitive answers to your winery cleaning efficiency, or your bottling line sanitation requires “zero” contaminants regardless of the nature of that contamination, ATP Bioluminescence assays might just be for you.

32. Issues With Both Methods Cell culture and ATP methods tend to under count remaining cells Training and technique There is a limit of detection with swabbing –Depending on area swabbed and swab material All ATP-Bioluminescence systems also have a limit of detection –No system can presently detect 100% of all ATP present. The lower the limit of detection, the more sensitive a system will be. Generally, LODs decrease as the price of the unit increases. You will need to decide how much sensitivity fits into your vision of quality and your budget.

33. Conclusions A combination of three methods are used to assess efficacy of sanitation methods –Education and Training –Inspections (visual assessments) –Microbiological testing For further discussion of ATP methods in the winery, refer to Ramsey, M. S. and Mills, D. A. 2013. Winery Biofilms: A source for contamination throughout the winery. Practical Winery and Vineyard Journal. Winter Issue.Winter Issue

34. Thank you “Making an art out of your technological life is the way to solve the problem of technology.” –Robert Pirsig,1974