3M Petrifilm Yeast Mold Count Interpretation Guide

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InterpretationguideThe 3M Petrifilm Yeast and MoldCount Plate is a sample-ready culturemedium system which containsnutrients supplemented with antibiotics,a cold-water-soluble gelling agent,and an indicator that facilitates yeastand mold enumeration. Petrifilm Yeastand Mold count plates are used for theenumeration of yeast and mould in thefood and beverage industries.YMYeast and Mold Count Plate1

Figure 1Total count 20Yeast count 16Mould count 43M Petrifilm Yeast and Mold Count Plate contains both yeastcolonies and mold colonies.Figure 3Estimated total count 500Estimated yeast count 480Estimated mould count 21When colonies number more than 150, estimate the count.Gridlines are visible with the use of a backlight to assist withestimated enumeration. Determine the average number ofcolonies in one square (1 cm2) and multiply it by 30 to obtainthe total count per plate. The inoculated area is approximately30 cm2. Yeast colonies may range in colour from tan (as in thisexample) to pink to blue-green.Figure 2Yeast and mould count 0Petrifilm Yeast and Mold count plate without yeast ormolds. Gridlines are visible with the use of a backlight toassist with estimated enumeration.Figure 4Estimated yeast count TNTCPetrifilm Yeast and Mold count plate containing yeast coloniestoo numerous to count (TNTC). The small, blue colonies at theedge of the plate (highlighted in the box) are present throughoutthe entire plate although less visible.For a more accurate count, further dilution of the sample may benecessary.For a more accurate count, further dilution of the sample may benecessary.Yeast and Mold Count Plate2

Figure 5Figure 6aFigure 6bEstimated mould count 64Mould count TNTCMold colonies are beginning to crowd and overlap each otheron the plate. Count each colony margin or focus. The plate canbe divided into sections to assist in counting. In this example,approximately 1/4 of the plate was counted, then the number ofcolonies counted was multiplied by 4 to get the estimated counton the plate. The section shown has 16 molds.Plates in Figures 6a and 6b are the same sample. Figure 6a is a1:10 dilution and has colonies that are small, faint and numerous,making it difficult to count. Figure 6b is a 1:100 dilution andshows how diluting a sample to obtain a colony count of lessthan 150 colonies makes counting easier. As with most growthmedia, in a highly competitive environment (such as Figure 6a),typical colony growth will be inhibited. For heavily contaminatedsamples such as these, further dilutions are recommended fora more accurate count and more typical colony growth (as inFigure 6b).For a more accurate count, further dilution of the sample may benecessary.Mould count 643

Macroscopic differentiationFigure 7Figure 8Yeast count 43Mould count 29Figure 9 shows typical yeast colonies. Characteristicstypical of yeast include:Figure 10 shows typical mould colonies. Characteristics typicalof mould include: Colony grows large Colony has diffuse edges Colony colour may vary as moulds produce a varietyof pigments (i.e., brown, beige, orange, blue-green) Colony appears flat Colony usually has a center focus (i.e., usually darker in colour,may also be different colour)Colony is smallColony has defined edgesColony colour can range from pink-tan to blue-greenColony may appear raisedColony typically is uniform in colour, no center focus(dark center)Microscopic differentiationYeasts and moulds are closely related and cannot always be distinguished from each other withoutmicroscopic examination.Figure 9Figure 10Figure 11Figure 12Figure 13To isolate colonies forfurther identification,lift the top film and pickfrom the colony withinthe gel using a loopor similar device.Transfer the colonyto a drop of sterilewater on a microscopeslide, cover witha coverslip, and viewunder a microscope.Yeast typically appearoval and may showbudding.Mould typicallyappear as branching orthread-like filaments(mycelium).Moulds shown aboveare in various stagesof germination.4

Phosphatase reactionFigure 14Figure 15Yeast and mould count 0Yeast and mould count 0The 3M Petrifilm Yeast and Mold Count Plates utilise a phosphatase indicator dye. All living cells contain phosphatase; therefore, naturalphosphatase in samples can cause the indicator to react. Two types of colour reactions are sometimes seen: a uniform blue backgroundcolour or intense, blue spots. Figure 7 shows uniform blue background color and Figure 8 shows intense blue spots which are often seen withspices or granulated products. Figure 8 also shows food particles that yielded phosphatase. To reduce a phosphatase reaction, follow one or more of these techniques:1. Dilute Sample: Further sample dilution will minimise blue background colour or reduce the number of intense blue spots.2. Sample Preparation: Mix sample and let settle for 3–5 minutes before plating. Draw sample from centre portion of sample container or usefiltered homogenizer bag to avoid plating large particles.3. Check and Note: Observe plates within 24-48 hours of incubation and make note of any colour change to aid in final interpretation.Reminders for useStorageInoculation1 mL 8 C1Store the unopened 3M Petrifilm Yeast and Mold Count Plate pouches atrefrigerated or frozen at temperatures 8 C ( 46 F). Use before expirationdate on package. Just prior to use, allowunopened pouches to come to roomtemperature before opening.2Yeast and Mold Count PlateSeal by folding the end of the pouchover and applying adhesive tape. Toprevent exposure to moisture, do notrefrigerate opened pouches. Storeresealed pouches in a cool, dry place forno longer than one month.3Place the Petrifilm Yeast and Mold countplate on a flat level surface. Lift the topfilm and with a pipette perpendicular tothe inoculation area, dispense 1 mL ofsample suspension onto the center of thebottom film.5

4Drop the top film down onto the sample.5Place the 3M Petrifilm Yeast and MoldSpreader on the center of the plate.Incubation7Lift the spreader and leave the plateundisturbed for at least one minute topermit the gel to form.86Gently apply pressure on the spreaderto distribute the inoculum over circulararea. Do not twist or slide the spreader.InterpretationIncubate plates with clear side up instacks of up to 20. Please refer to productinstructions for third party validatedmethods. Because some molds may growquickly, it may be useful to read and countplates at 3 days as smaller colonies maybe obscured by larger, overgrown moldsat 5 days. If this happens, the 3 day countmay be used; however, it should be reportedas an estimated count.9Petrifilm Yeast and Mold count platescan be counted using a standard colonycounter or other illuminated magnifier.Use appropriate sterile diluentsButterfield’s phosphate buffer, 0.1% peptone water, peptone salt diluent, saline solution (0.85-0.90%),bisulphite-free letheen broth or distilled water.Do not use diluents containing citrate, bisulphite or thiosulfate with 3M Petrifilm Yeast and Mold CountPlates; they can inhibit growth.If citrate buffer is indicated in the standard procedure, substitute with one of the buffers listed above,warmed to 40–45 C.3M Food Safety offers a fullline of products to accomplisha variety of your microbialtesting needs. For more productinformation, visit us at3M.com/foodsafety/PetrifilmUser’s Responsibilities: 3M Petrifilm Plate performance has not been evaluated with allcombinations of microbial flora, incubation conditions and food matrices. It is the user’s responsibilityto determine that any test methods and results meet the user’s requirements. Should re-printing of thisInterpretation Guide be necessary, user’s print settings may impact picture and colour quality.3M UK PLCCharnwood Campus10 Bakewell RoadLoughborough LE11 5RBFor detailed CAUTIONS, DISCLAIMER OF WARRANTIES/LIMITED REMEDYand LIMITATION OF 3M LIABILITY, STORAGE AND DISPOSAL informationand INSTRUCTIONS FOR USE, see Product’s package ty3M and Petrifilm are trademarks of the 3M company. 3M 2019. All rights reserved. 70-2008-4258-4. J451005.6

1. Dilute Sample: Further sample dilution will minimise blue background colour or reduce the number of intense blue spots. 2. Sample Preparation: Mix sample and let settle for 3-5 minutes before plating. Draw sample from centre portion of sample container or use filtered homogenizer bag to avoid plating large particles. 3.

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